Digitized by the Internet Archive in 2007 with funding from IVIicrosoft Corporation http://www.archive.org/details/comparativeelectOOboserich COMPARATIVE ELECTRO-PHYSIOLOGY WORKS BY THE SAME AUTHOR RESPONSE IN THE LIVING AND NON- LIVING. With 117 Illustrations. 8vo. lOs. 6d. 1902. PLANT RESPONSE; as a means of Physio- logical Investigations. Witli 278 Illustrations. 8vo. 21s. 1906. LONGMANS, GREEN & CO., 39 Paternoster Row, London. New York, Bombay, and Calcutta COMPARATIVE ELECTRO-PHYSIOLOGY A PHYSICO-PHYSIOLOGICAL STVDY BY JAGADIS CHUNDER ^OSE, M.A., D.Sc. PROFESSOR, PRESIDENCY COLLEGE, CALCUTTA WITH ILLUSTRATIONS LONGMANS, GREEN, AND CO. 39 PATERNOSTER ROW. LONDON NEW YORK, BOMBAY, AND CALCUTTA 1907 All rights reserved ?fb ~-1 1907 PREFACE I^^ive phenomena in general, which I commenced with the ubHcation of a Memoir ^ at the International Congress of cience, Paris, 1900. In this first of my publications on the bject I undertook to show the similarities of response in organic and living substances. The method which I at at time employed for obtaining my response-records was at of Conductivity Variation. With the object of showing at the similarity of response here demonstrated to exist as due to some fundamental molecular reaction, common matter in general, and therefore to be detected by any ethod of recording response, I next undertook to record the Electro-motive Variation under stimulus. Believing, as I did, in the continuity of these responsive phenomena, I ^Jsed the same experimental devices by which I had already Hwcceeded in obtaining the electric response of inorganic sub- stances, to test whether ordinary plants also, meaning those ually regarded as insensitive, would or would not exhibit excitatory electrical response to stimulus. The stimulation ' ' De la Gen^ralite des Phenomenes Moleculaires produits par rEleciricite sur la Matiere Inorganique et sur la Matiere Vivante ' ( Travatix du Congris International de Physique, Paris, 1900). See also ' On the Similarity of Effects of Electrical Stimulus on Inorganic and Living Substances,' Report Brit. Assoc, Bradford, September 19CX) {Electrician). I VI COMPARATIVE ELECTRO-PHYSIOLOGY employed was mechanical and quantitative, thus obviating many sources of complication. By this method I was able to show that every plant, and every organ of every plant, gave true excitatory electrical response. As observations similar to these were subsequently made by another investigator, I quote here the following summary of my results from the preliminary account which I communicated to the Royal Society, May 7, and afterwards read, with accompanying ex- perimental demonstration, before the Society, on June 6, 1901. * An interesting link, between the response given by inor- ganic substances and the animal tissues, is that given by plant tissues. By methods somewhat resembling that described above, I have obtained from plants a strong electric response to mechanical stimulus. The response is not confined to sensitive plants like Mimosa, but is universally present. I have, for example, obtained such response from the roots, stems, and leaves of, among others, horse-chestnut, vine, white lily, rhubarb, and horse-radish. The ''current of injury" is, generally speaking, from the injured to the uninjured part. A " negative variation " is also produced. I obtained both the single electric twitches and tetanus. Very interesting also are the effects of fatigue, of temperature, of stimulants, and of poison. Definite areas killed by poison exhibit no response, whereas neighbouring unaffected portions show the normal response.' ^ It may be well to point out here that at the time when this communication was made, the view that ordinary plants were excitable, and responded to mechanical stimulus by * A more complete account will be found in the report of my ' Friday Evening Discourse ' before the Royal Institution, May 10, 1901, and in the Journal of the Linnean Society^ vol. xxxv. p. 275. PREFACE vii. definite electro-motive changes, was regarded as highly- controversial. Indeed, in the discussion which followed the reading of my Paper, on June 6, 1901, Sir John Burdon Sanderson went so far as to state that this excitatory response of ordinary plants to mechanical stimulation was an impossibility. My next investigation was directed towards the question whether the responsive effects which I had shown to occur in ordinary plants might not be further exhibited by means iof visible mechanical response, thus finally removing the dis- [tinction commonly assumed to exist between the ' sensitive ' ind supposed non-sensitive. These results were published [in my work on Plant Response,^ where the effects of various [environmental stimuli on the different plant organs were lemonstrated by means of responsive movements. Many anomalous effects hitherto ascribed to specific sensibilities [were here shown to be due to the differential excitability [of anisotropic structures, and to the opposite effects of [external and internal stimuli. Among other things, it was there shown that internal stimulus was in reality derived from ^external sources, and that the term * autonomous response ' was a misnomer, since all movements were due, either to the immediate effects of external stimulus, or to stimulus previously absorbed and held latent in the plant, to find subsequent ex- pression. It was further shown that not gross mechanical movements alone, but also other invisible movements, were initiated by the action of stimulus ; that external stimulus, so far from invariably causing a run-down of energy, more often brought about its accumulation by the plant ; and that the various activities, such as the ascent of sap and growth, ' Plant Response as a Means of Physiological Investigation^ 1906. Vlil COMPARATIVE ELECTRO-PHYSIOLOGY were thus in reality different reactions to the stimulating action of energy supplied by the environment. With regard to these points, my results have been in direct opposition to current views, according to which the effect induced by stimulus is always disproportionately greater than the stimulus. From the plausible analogy of the firing-off of a gun by the pulling of a trigger, or the action of a combustion-engine, it has been customary to suppose that all response to stimulus must be of the nature of an explosive chemical change, accompanied by an inevitable run-down of energy. This supposition, however, overlooks the obvious fact that the plant is not consumed by the incessant and multifarious stimuli of its environment. Rather, as we all know, it is the energy of the environ- ment which is the agent that fashions the microscopic embryo into the gigantic banyan-tree. And it is clear that, for this to be possible, the energy contributed by the blow of external stimulus must have been largely conserved. In the course of the present work, I have not only been able to corroborate, by means of electrical response, the various results which I had already established, with regard to the plant, by mechanical response, but I have also ex- tended the electrical method in various directions, so as to include many more recondite problems in connection with the irritability of living tissues. It was my original inten- tion to confine this investigation to the Electro-physiology of Plants. But, finding that in the results so obtained I pos- ses.sed a key to that of the animal also, I proceeded to apply the same methods of inquiry, and to use the same experi- mental devices, in the one case as in the other. I have thus been able to trace out the gradual differentiation of various PREFACE ix responsive peculiarities, characteristic of given tissues, from their simplest types in the plant to their most complex in the animal. The value of such a comparative method of study, for the elucidation of biological problems in general, is sufficiently obvious. Exception may be taken with regard to the unorthodox point of view from which various ques- tions in animal physiology have been approached. It must be remembered, however, that in this work the attempt has been to explain responsive phenomena in general on the consideration of that fundamental molecular reaction which occurs even in inorganic matter. My mode of investigation has thus been determined by the necessary progression ^from simple to complex, and by my conviction as to the :ontinuity which existed between them. And from this ittempt it will be seen that various results, which, accord- ing to the so-called vitalistic assumption were anomalous, ire, in fact, capable of an increasingly simple and satis- Lctory explanation. It must also be understood that my '^ork deals mainly with the electrical response of plants, ind that its extension into the field of Animal Electro- )hysiology was intended for the demonstration of the con- :inuity between the two. It was therefore impossible, in the short space at my disposal, to make more than the >rief necessary references to the different theories already in vogue concerning the response of various animal tissues, 'hese will be found, in all their detail, in the excellent Lccount given in the standard work of Biedermann.^ For the sake of clearness, however, I shall at this point enumerate a few only of the points of difference between :urrent views and the results, obtained from actual experi- ' Biedermann, Electro-physiology (English translation), 1896. X COMPARATIVE ELECTRO-PHYSIOLOGV ment, which I have set forth in the present volume. The reactions of different tissues have hitherto been re- garded as specifically different. As against this, a continuity has here been shown to exist between them. Thus, nerve was universally regarded as typically non-motile ; its re- sponses were believed to be characteristically different from those of muscle. I have been able to show, however, that nerve is not only indisputably motile, but also that the investigation of its response by the mechanical method is capable of greater delicacy, and freedom from error, than that by the electrical. The characteristic variations in the response of nerve, moreover, are, generally speaking, similar to those of the muscle. It has been customary, again, to regard plants as devoid of the power to conduct true excita- tion. But I have shown that this view is incorrect. Experi- ments have been described, showing that the response of the isolated vegetal nerve is indistinguishable from that of animal nerve, throughout a long series of parallel variations of condition. So complete, indeed, has that similarity between the responses of plant and animal, of which this is an instance, been found, that the discovery of a given responsive characteristic in one case has proved a sure guide to its observation in the other, and the explanation of a phenomenon, under the simpler conditions of the plant, has been found fully sufficient for its elucidation under the more complex circumstances of the animal. Many anomalous conclusions, with regard to the response of certain animal tissues, had arisen from the failure to take account of the differential excitability of anisotropic organs. Now this is a subject which, in the case of the simple plant organ, is capable of very exact investigation. I have been PREFACE xi able to show that this differential excitability is widely- present as a factor in determining the character of special responses, and that it finds its culminating expression in the electrical organs of certain well-known fishes. Few conclusions in Electro-physiology have been sup- posed to rest on securer foundations than the generalisation known as Pflliger's Law of the polar effects of currents. I lave found, however, that this law is not by any means >f such universal application as had been supposed, since, ibove and below a certain range of electromotive intensity, le polar effects of currents are precisely opposite to those enunciated by Pfliiger. Finally, that nervous impulse, which must necessarily form the basis of sensation, was supposed to lie beyond iny conceivable power of visual scrutiny. But it has here )een shown that this impulse is actually attended by change ►f form, and is therefore capable of direct observation. This '^ave of nerve-disturbance, moreover, instead of being single, las been shown to be of two different kinds, in which fact, ^as I have further explained, lies the significance of the two different qualities or tones of sensation. In the concluding portion of the paper which I read >efore the Bradford meeting of the British Association in le year 1900, I said : — In the phenomena described above there is little ►reach of continuity. It is difficult to draw a line and : " Here the physical process ends, and the physiological ►rocess begins " ; or " That is a phenomenon of inorganic latter, and this is a vital phenomenon, peculiar to living )rganisms '' ; or " These are the lines of demarcation that iparate the physical, the physiological, and the beginning Xii COMPARATIVE ELECTRO-PHYSIOLOGY of psychical processes." Such arbitrary lines can hardly be drawn. ' We may explain each of the above classes of phenomena by making numerous and independent assumptions ; or, finding some property of matter common and persistent in the living and non-living substances, attempt from this common underlying property to explain the many phe- nomena which at first appear so different. And for this it may be said that the tendency of science has always been to attempt to find, wherever facts justify it, an under- lying unity in apparent diversity.' It was for the demonstration of this underlying unity that I set out on these investigations seven years ago. And now, in bringing to its close another stage of their publication, I may, perhaps, be permitted to express the hope that by them not only may a deeper perception of this unity have been made attainable, but also that many regions of inquiry may prove to have been opened out, which had at one time been regarded as beyond the scope of experi- mental exploration. I take this opportunity to thank my assistants for their efficient help in these researches. J. C. BOSE. Presidency Collegk, Calcutta : Augnsi 1906. CONTENTS CHAPTER I THE MOLECULAR RESPONSIVENESS OF MATTER sponse to stimulus by change of form — Permeability variation — Variation of solubility— Method of resistivity variation : {a) positive variation; (d) negative variation — Sign of response changed under different molecular modifications — Response of vegetable tissue by variation of electrical resistance — Response by electro-motive variation in inorganic substances — The method of block — Positive and negative responses — Similar responses in living tissues — Effects of fatigue, stimulants, and poisons on inorganic and organic responses — Method of relative depression, or negative variation, so called ........ CHAPTER n THE ELECTRO-MOTIVE RESPONSE OF PLANTS TO DIFFERENT , FORMS OF STIMULATION Historical — Difficulties of investigation — Electrical response of pulvinus of ^ Mimosa — Simultaneous mechanical and electrical records — Division of ^B plants into 'ordinary' and 'sensitive' arbitrary — Mechanical and ^B electrical response of * ordinary ' plants — Direct and transmitted stimu- li lation— All forms of stimulus induce excitatory change of galvanometric CHAPTER HI THE APPLICATION OF QUANTITATIVE STIMULUS AND K ELATION BETWEEN STIMULUS AND RESPONSE Conditions of obtaining uniform response— Torsional vibration as a form of stimulus— Method of block— Effective intensity of stimulus dependent XIV COMPARATIVE ELECTRO-PHYSIOLOGY recorder — Uniform electric responses — List of suitable specimens — Effect of season on excitability — Stimulation by thermal shocks — Thermal stimulator — Second method of confining excitation to one contact — In- creasing response to increasing stimulus— Effect of fatigue— Tetanus . 29 CHAPTER IV OBSERVATION BY RHEOTOME ON ELECTRIC RESPONSE IN PLANTS Response-curve showing general time-relations — Instantaneous mechanical stimulation by electro-magnetic release— Arrangement of the rheotome — Tabular statement of results of rheotomic observations — Rhythmic multiple responses .......... 44 CHAPTER V THE ELECTRICAL INDICATIONS OF POSITIVE AND NEGATIVE TURGIDITY-VARIATIONS Motile responses of opposite signs, characteristic of positive and negative turgidity-variations - Indirect hydrostatic effect of stimulus causes expansion and erection of leaf-Positive and negative work — Wave of increased hydrostatic tension transmitted with relatively greater velocity than wave of true excitation— Method of separating hydro-positive and excitatory effects— Indirect effect of stimulus, causing positive turgidity- variation induces galvanometric positivity — Antagonistic elements in the electrical response— Separation of hydro-positive from true excitatory effect by means of physiological block . . ^ . . , 54 CHAPTER VI EXTERNAL STIMULUS AND INTERNAL ENERGY Hydraulic transmission of energy in plants — True meaning of tonic condi- tion—Opposite expressions of internal energy and external stimulus seen in growth-response— Parallelism between responses of growing and motile organs— Increased internal energy caused by augmentation 01 temperature finds expression in enhanced rate of growth ; erection of motile leaf ; curling movement of spiral tendril ; and galvanometric positivity— External stimulus induces opposite eftect in all these cases- Sudden variation of temperature, acting as a stimulus, induces transient retardation of growth ; depression of motile leaf ; uncurling movement of spiral tendril ; and galvanometric negativity— Laws of mechanical and electrical response 60 CONTENTS XV CHAPTER VII ABSORPTION AND EMISSION OF ENERGY IN RESPONSE PAGE Sign of response determined by latent energy of tissue, and Vjy intensity of external stimulus — Sub-tonic, normal and hyper-tonic conditions— The critical level — Outward manifestation of response possible only when critical level is exceeded — Three typical cases : response greater than ■ stimulus ; response equal to stimulus ; and response less than stimulus — Investigation by growth-response — The sum of work, internal and external, performed by stimulus constant — ^Positive response of tissues characterised by feeble protoplasmic activity or sub-tonicity — Enhance- tment of normal excitability of sub-tonic tissue by absorption of stimulus 76 CHAPTER VIII VARIOUS TYPES OF RESPONSE lemical theory of response — Insufficiency of the theory of assimilation and dissimilation — Similar responsive effects seen in inorganic matter — Modifying influence of molecular condition on response — Five molecular stages, A, B, c, D, E — Staircase effect, uniform response, fatigue — No sharp line of demarcation between physical and chemical phenomena — Volta-chemical effect and by-productions — Phasic alternation —Alter- nating fatigue — ^Rapid fatigue under continuous stimulation — In sub- tonic tissue summated effect of latent components raises tonicity and excitability — Response not always disproportionately greater than stimulus— Instances of stimulus partially held latent : staircase and additive effects, multiple response, renewed growth .... 86 CHAPTER IX DETECTION OF PHYSIOLOGICAL ANISOTROPY BY ELECTRIC RESPONSE inomalies in mechanical and electrical response — Resultant response deter- mined by differential excitability — Responsive current from the more to the less excitable — Laws of response in anisotropic organ — Demonstra- tion by means of mechanical stimulation— Vibrational stimulus — Stimu- lation by pressure — Quantitative stimulation by thermal shocks . . 107 CHAPTER X THE NATURAL CURRENT AND ITS VARIATIONS Fatural current in anisotropic organ from the less to the more excitable — External stimulus induces responsive current in opposite direction — Increase of internal energy induces positive, and decrease negative, xvi COMPARATIVE ELECTRO-PHYSIOLOGY variation of natural current — Effect on natural current of variation of temperature— Effect of sudden variation— Variation of natural current by chemical agents, referred to physiological reaction — Agents which render tissue excitable, induce the positive, and those which cause excita- tion, the negative variation — Action of hydrochloric acid — Action of Na^COg — Effect modified by strength of dose — Effect of CO, and of alcohol vapour— Natural current and its variations— Extreme unrelia- bility of negative variation so-called as a test of excitatory reaction — Reversal of natural current by excessive cold or by stimulation — Re- versal of normal response under sub-tonicity or fatigue . . . .116 CHAPTER XI VARIATIONS OF EXCITABILITY UNDER CHEMICAL REAGENTS Induced variation of excitability studied by two methods: (i) direct (2) transmitted stimulation — Effect of chloroform — Effect of chloral — Effect of formalin —Advantage of the Method of Block over that of negative variation — Effect of KHO — Response unaffected by variation of resistance — Stimulating action of solution of sugar — Of sodium carbon- ate — Effect of doses — Effect of hydrochloric acid — Diphasic response on application of potash— Conversion of normal negative into almormal positive response by abolition of true excitability 1 29 CHAPTER XII VARIATIONS OF EXCITABILITY DETERMINED BY METHOD OF INTERFERENCE Arrangement for interference of excitatory waves — Effect of increasing difference of phase — Interference effects causing change from positive to negative, through intermediate diphasic — Diametric balance — Effect of unilateral application of KHO — Effect of unilateral cooling . . . 141 CHAPTER XIII CURRENT OF INJURY AND NEGATIVE VARIATION Different theories of current of injury — Pre-existence theory of Du Bois- Reymond — Electrical distribution in a muscle-cylinder — Electro-mole- cular theory of Bernstein — Hermann's Alteration Theory— Experiments demonstrating that so-called current of injury is a persistent after-effect of over-stimulation — Residual galvanometric negativity of strongly excited tissue — Distribution of electrical potential in vegetable tissue with one end sectioned — Electrical distribution in plant-cylinder similar to that in muscle-cylinder — True significance of response by negative variation — Apparent abnormalities in so-called current of injury — • Positive ' current of injury 149 CONTENTS xvil CHAPTER XIV CURRENT OF DEATH — RESPONSE BY POSITIVE VARIATION PAGE Anomalous case of response by positive variation — Inquiry into the cause — Electric exploration of dying and dead tissue : death being natural — Determination of electric distribution in tissue with one end killed — Dying tissue shows maximum negativity, and dead tissue, positivity to living — Explanation of this peculiar distribution — Response by negative or positive variation, depending on degree of injury — Three typical cases -Explanation by theory of assimilation and dissimilation misleading — All response finally traceable to simple fundamental reactions . .164 CHAPTER XV EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE leneral observation of effect of temperature on plant — Effect of fall and rise of temperature on autonomous response of Desmodium — Effect of frost in abolition of electrical response — After-effects of application of cold, in Eucharis, Ivy and Holly — Effect of rise of temperature in diminishing height of response — This not probably due to diminution of excitability — Similar effect in autonomous motile response of Desmodium — En- hanced response as after-effect of cyclic variation of temperature— Aboli- tion of response at a critical high temperature ..... 180 CHAPTER XVI THE ELECTRICAL SPASM OF DEATH )ifferent post-mortem symptoms— Accurate methods for determination of death-point — Determination of death-point by abolition or reversal of normal electrical response— Determination of death-point by mechanical death-spasm — From thermo-mechanical inversion — By observation of electrical spasm : {a) in anisotropic organs : {b) in radial organs— Simul- taneous record of electrical inversion and reversal of normal electrical response — Remarkable consistency of results obtained by different methods— Tabulation of observations .192 CHAPTER XVII MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE [Repeated responses under single strong stimulus — Multiple mechanical response in Biophyttim — Multiple electrical responses in various animal and vegetable tissues— Continuity of multiple and autonomous response — Transition from multiple response to autonomous, and vice versa — a xviii COMPARATIVE ELECTRO-PHYSlOLOGY PAGE Autonomous mechanical response of Desmodium gyrans and its time- relations — Simultaneous mechanical and electrical records of automatic pulsations in Desmodium — Double electrical pulsation, principal and subsidiary waves — Electrical pulsation of Desmodiiun leaflet under physical restraint— Growth-pulsation — So- called current of rest in grow- ing plants ............ 207 CHAPTER XVIII RESPONSE OF LEAVES Observations of Burdon Sanderson on leaf-response in Diomxa — Leaf and stalk currents — Their opposite variations under stimulus — Similar leaf- and-stalk currents shown to exist in ordinary leaf of Ficus religiosa — Opposite- directioned currents in Citrus dectwtana — True explanation of these resting-currents and their variations— Electrical effect of section of petiole on Dioncea and Ficus religiosa — Fundamental experiment of Burdon Sanderson on lamina of Dionaa — Subsequent results — Experi- mental arrangement with symmetrical contacts — Parallel experiments on sheathing leaf of i^«j-fl! — Explanation of various results . . . 223 CHAPTER XIX THE LEAF CONSIDERED AS AN ELECTRIC ORGAN Electrical organs in fishes — Typical instances, Torpedo and Malepterurus — Vegetal analogues, leaf of Pterospermum and carpel of Dillenia indica or pitcher of Nepenthe — Electrical response to transmitted excitation — Response to direct excitation — Uni-directioned response to homo- dromous and heterodromous shocks — Definite-directioned response shown to be due to differential excitability — Response to equi-alternating electrical shocks — Rheotomic observations— Multiple excitations — Multiplication of terminal electromotive effect, by pile-like arrangement, in bulb of Uric lis lily .241 CHAPTER XX THE THEORY OF ELECTRICAL ORGANS Existing theories— Their inadequacy — The * blaze -current ' so called — Re- sponse uni-directioned, to shocks homodromous or heterodromous, characteristic of electric organs — Similar results with inorganic specimens — Uni-directioned response due to differential excitability— Electrical response of pulvinus of Mimosa to equi-alternating electric shocks — Re- sponse of petiole of Musa — Of plagiotropic stem of Cucurbita — Of Eel — The organ-current of electric fishes — Multiple responses of electrical organ — Multiple responses of Biofhyium 259 CONTENTS xix CHAPTER XXI DETERMINATION OF DIFFERENTIAL EXCITABILITY UNDER ELECTRICAL STIMULATION PAGE Advantage of electrical stimulation, in its flexibility — Drawbacks due to fluctuating factors of polar effects, and counter polarisation-current — Difficulties overcome by employment of equi-alternating electric shocks — Methods of the After-effect and Direct-effect — Experiment of ■ Von Fleischl on response of nerve — Complications arising from use of make and break shocks — Rotating reverser— Motor transformer — Re- sponse of Mtisa to equi-alternating shocks — Abolition of this response by chloroform — Response records of plagiotropic Cucurbita and Eel — IDiff"erential excitability of variegated leaves, demonstrated by electric response ............ 272 CHAPTER XXn RESPONSE OF ANIMAL AND VEGETAL SKINS Currents of rest and action — Currents in animal skin — Theories regarding these — Response of vegetal skin — Stimulation by Rotary Mechanical I Stimulator — Response of intact human skin— Isolated responses of upper and lower surfaces of specimens— Resultant response brought about by differential excitability of the two surfaces — Differences of excitability between two surfaces accounted for — Response of animal and vegetal skins not essentially different — General formula for all types of response of skin— Response of skin to different forms of stimulation gives similar results — Response to equi-alternating electric shocks: (i) Method of the After Effect ; (2) Method of Direct Effect — Response of grape skin — Similar response of frog's skin — Phasic variation of current of rest induced as result of successive stimulation in (a) grape skin ; [b) frog's skin ; {c) pulvinus of J/m^jd!— Phasic variation in autonomous me- chanical response of Desmodium gyratts — Autonomous variation of current of rest — True current of rest in skin from outer to inner — This may be reversed as an excitatory after-effect of preparation — Electrical response of skin of neck of tortoise — Electrical response of skin of tomato — Normal response and positive after-effect — Response of skin of gecko— Explanation of abnormal response 287 CHAPTER XXHI RESPONSE OF EPITHELIUM AND GLANDS Epidermal, epithelial, and secreting membranes in plant tissues— Natural [ resting-current from epidermal to epithelial or secretory surfaces — Current I of response from epithelial or secretory to epidermal surfaces — Response a 2 XX COMPARATIVE ELECTRO-PHYSIOLOGY oi Dillenia — Response of water-melon — Response of foot of snail — The so-called current of rest from glandular surface really due to injury — Misinterpretation arising from response by so-called ' positive variation ' — Natural current in intact foot of snail, and its variation on section — Response of intact human armpit — Response of intact human lip — Lingual response in man — Reversal of normal response under sub- minimal or super-maximal stimulation — Differential excitations of two surfaces under different intensities of stimulus, with consequent changes in direction of responsive currents, diagrammatically represented in characteristic curves — Records exhibiting responsive reversals . -312 CHAPTER XXIV RESPONSE OF DIGESTIVE ORGANS Consideration of the functional peculiarities of the digestive organ — Alter- nating phases of secretion and absorption — Relation between secretory and contractile responses. Illustrated by (a) preparation of Mimosa ; {h) glandular tentacle of Drosera — General occurrence of contractile re- sponse — True current of rest in digestive organs — Experiments on the pitcher of Nepenthe— Thx^a definite types of response under different con- ditions — Negative and positive electrical responses, concomitant with secretion and absorption — Multiple responses due to strong stimulation — Response in glandular leaf of Drosera — Normal negative response reversed to positive under continuous stimulation— Multiple response in Drosera — Response of fret's stomach to mechanical stimulation — Re- sponse of stomach of tortoise — Response of stomach of gecko— Multiple response of frog's stomach, showing three stages — negative, diphasic, and positive— Phasic variations 329 CHAPTER XXV ABSORPTION OF FOOD BY PLANT AND ASCENT OF SAP Parallelism between responsive reactions of root and digestive organ- Alter- nating phases of secretion and absorption — Association of absorptive process with ascent of sap— Electrical response of young and old roots — Different phasic reactions, as in pitcher of Nepenthe — Kes^nsQ to chemical stimulation— Different theories of ascent of sap— Physical versjis excitatory theories— Objections to excitatory theory— Assumption that wood dead unjustified— Demonstration of excitatory electrical re- sponse of sap-wood — Strasburger's experiments on effect of poisons on ascent of sap— Current inference unjustified 349 CONTENTS xxi CHAPTER XXVI THE EXCITATORY CHARACTER OF SUCTIONAL RESPONSE PAGE Propagation of excitatory wave in plant attended by progressive movement of water— Hydraulic response to stimulus— The Shoshungraph— Direct and photographic methods of record — Responsive variations of suction under physiological modifications induced by various agents — Effects of lower- ing and raising of temperature— Explanation of maintenance of suction, when root killed — Effect of poison influenced by tonic condition — Effect of anaesthetics on suctional response — Excitatory versus osmotic action — Stimulation by alternating induction-shocks — Terminal and sub-ter- minal modes of application — Three modes of obtaining response-records, namely (i) the unbalanced, (2) the balanced, (3) the over-balanced — Renewal of suction previously at standstill, by action of stimulus- -Re- ponsive enhancement of suction by stimulus — After-effect of stimulus — Diminution of latent period as after-effect of stimulus — Response under over-balance— Response under sub-terminal stimulation — Variation of response under seasonal changes ........ 365 CHAPTER XXVII RESPONSE TO STIMULUS OF LIGHT [eliotropic plant movements reducible to fundamental reaction of contrac- tion or expansion — Various mechanical effects of light in pulvinated and growing organs — Electrical response induced by light not specific, but concomitant to excitatory effects— Electrical response of plant to light not determined by presence or absence of chloroplasts — Effect of unilateral application of stimulus on transversely distal point — Positive response due to indirect effect and negative to transmission of true excitation — Mechanical response of leaf of Mimosa to light applied on upper half of pulvinus— Mechanical response consists of erection or positive movement, followed by fall or negative movement — Electrical response of leaf of Mhnosa to light applied on upper half of pulvinus ; induction in lower half of pulvinus of positivity followed by negativity — Longitudinal transmission of excitatory effect, with concomitant galvano- metric negativity -Direct effect of light and positive after effect- Circumstances which are effective in reversing normal response— Plants In slightly sub-tonic condition give positive followed by negative response ^Exemplified by (a) electrical and {b) growth response — Examples of positive response to light — Periodic variation of excitability — Multiple mechanical response under light— Direct and after-effect — Multiple electrical response under light, with phasic alterations of ( — + — -F ) or ( + — -I- — ) —After-effects ; unmasking of antagonistic elements, either ^lus or minus — Three types of after-effects , . , . . 392 XXll COMPARATIVE ELECTRO-PHYSIOLOGY CHAPTER XXVIII RESPONSE OF RETINA TO STIMULUS OF LIGHT •AGE Response of retina — Determination of true current of rest — Determination of differential excitabilities of optic nerve and cornea, and optic nerve and retina — The so-called positive variation of previous observers indicates the true excitatory negative— Retino-motor effects— Motile responses in nerve — Varying responsive effects under different conditions — Reversal of the normal response of light due to (i) depression of excitability below par ; (2) fatigue — The sequence of responsive phases during and after application of light — Demonstration of multiple responses in retina under light, as analogous to thosi in vegetable tissues — Three types of after-effect — Multiple after-excitations in human retina — Binocular Alternation of Vision — Demonstration of pulsatory response in human retina during exposure to light . . . • 415 CHAPTER XXIX GEO-ELECTRIC RESPONSE Theory of Hydrostatic Pressure and Theory of Statoliths — Question regarding active factor of curvature in geotropic response, whether contraction or expansion — Crucial experiment by local application of cold — Reasons for delay in initiation of true geotropic response— Geo-electric response of shoot —Due to active contraction of upper side, with concomitant gal- vanometric negativity — Geo-electric response of an organ physically restrained 434 CHAPTER XXX DETERMINATION OF VELOCITY OF TRANSMISSION OF EXCITATION IN PLANT TISSUES Transmission of excitation in plants not due to hydromechanical disturbance, but instance of transmission of protoplasmic changes — Difficulties in accurate determination of velocity of transmission — A perfect method — Diminution of conductivity by fatigue— Increased velocity of transmission with increasing stimulus— Effect of cold in diminishing conductivity — Effect of rise of temperature in enhancing conductivity— Excitatory concomitant of mechanical and electrical response — Electrical methods of determining velocity of transmission— Method of comparison of longi- tudinal and transverse conductivities — Tables of comparative velocities in animal and plant -Existence of two distinct nervous impulses, positive apd negative ..,,.,,.... 444 CONTENTS xxiii CHAPTER XXXI ON A NEW METHOD FOR THE QUANTITATIVE STIMULATION OF NERVE I'AGt Drawbacks to use of electrical stimulus in recording electrical response — Response to equi-alternating electrical shocks — Modification of response by decline of injury — Positive after-effect — Stimulation of nerve by thermal shocks— Enhancement of normal response after tetanisation — Untenability of theory of evolution of carbonic acid — Abnormal positive response converted into normal negative after tetanisation — Gradual transition from positive to negative, through intermediate diphasic — Effect of depression of tonicity on excitability and conductivity— Con- version of abnormal into normal response by increase of stimulus-intensity — Cyclic variation of response under molecular modification . . . 456 CHAPTER XXXn ELECTRICAL RESPONSE OF ISOLATED VEGETAL NERVE Specialised conducting tissues — Isolated vegetal nerve— Method of ob- taining electrical response in vegetal nerve — Similarity of responses of plant and animal nerve : {a) action of ether— {d) action of carbonic acid — (c) action of vapour of alcohol — {d) action of ammonia — {e) ex- hibition of three types of response, negative, diphasic and positive — (/) effects of tetanisation of normal and modified specimens — Effect of increasing stimulus on response of modified tissue .... 468 CHAPTER XXXIII THE CONDUCTIVITY BALANCE leceptivity, conductivity, and responsivity — Necessity for distinguishing these — Advantages of the Method of Balance — Simultaneous comparison of variations of receptivity, conductivity and responsivity — The Conductivity Balance — Effect of NaJZO^ on frog's nerve — Effect of CUSO4 — Effect, of chemical reagents on plant nerve — Effect of CaCl.^ on responsivity — Responsivity variation under KCl — Comparison of simultaneous effects of NaCl and NaBr on responsivity — Effects of NagCOa in different dilutions on conductivity — Demonstration of two different elements in conductivity, velocity and intensity — Conductivity versus responsivity — (a) effect of KI — {b) Effect of NaT — Effect of alcohol on receptivity, conductivity, and responsivity — Comparison of simultaneous effects of alcohol — (a) on receptivity versus conductivity- (3) on receptivity versus responsivity. ........... 479 XXIV COMPARATIVE ELECTRO-PHYSIOLOGY CHAPTER XXXIV EFFECT OF TEMPERATURE AND AFTER-EFFECTS OF STIMULUS ON CONDUCTIVITY PAGE Effect of temperature in inducing variations of conductivity : {a) by Method of Mechanical Response ; {d) by Method of Electric Balance— Effect of cold — Effect of rising temperature— The Thermal Cell— After-effect of stimulation on conductivity — The Avalanche Theory — Determination of after-effect of stimulus on conductivity by the Electrical Balance —After- effects of moderate stimulation — After-effect of excessive stimulation . 497 CHAPTER XXXV MECHANICAL RESPONSE OF NERVE Current assumption of non-motility of nerve— Shortcomings of galvano- melric modes of detecting excitation — Mechanical response to continuous electric shocks — Optical Kunchangraph — Effect of ammonia on the mechanical response of nerve — Effect of morphia — Action of alcohol — Of chloroform — Abnormal positive or expansive response converted into normal contractile through diphasic, after tetanisation — Similar effects in mechanical response of vegetal nerve — Mechanical response due to transmitted effects of stimulation — Determination of velocity of trans- mission — Indeterminateness of velocity in isolated nerve — Kunchan- graphic records on smoked glass— Oscillating recorder— Mechanical response of afferent nerve — Record of mechanical response of nerve due to transmitted stimulation, in gecko — Fatigue of conductivity — Conver- sion of normal contractile response into abnormal expansive, through diphasic, due to fatigue 507 CHAPTER XXXVI MULTIPLE RESPONSE OF NERVE Great sensitiveness of the high magnification Kunchangraph — Individual contractile twitches shown in tetanisation of nerve — Sudden enhance- ment of mechanical response of nerve on cessation of tetanisation — Secondary excitation — Multiple mechanical excitation of nerve by single strong stimulation — Multiple mechanical excitation of nerve by drying . 532 CHAPTER XXXVn RESPONSE BY VARIATION OF ELECTRICAL RESISTIVITY Variation of resistance in Dt'o/um, by * modification ' — Excitatory change, its various independent expressions — Characteristic difficulties of investi- CONTENTS XXV gation — Morograpbic record by variation of resistivity — Inversion of curves at death-point — Similarities between mechanical, electro-motive and resistivity curves of death — The true excitatory effect attended by diminution of resistance — Response of plant nerve by resistivity varia- tion — Independence of resistivity and mechanical variations — Responsive resistivity variation in frog's nerve, and its modification under anaesthetics 540 CHAPTER XXXVIII FUNCTIONS OF VEGETAL NERVE Feeble conducting power of cortical tissues — Heliotropic and geotropic effects dependent on response of cortical tissues only— Phenomenon of correlation — Excitability of tissue maintained in normal condition only under action of stimulus — Physiological activities of growth, ascent of sap, and motile sensibility, maintained by action of stimulus — Critical importance of energy of light — Leaf- venation a catchment-basin — Trans- mission of energy to remotest parts of plants — Plant thus a connected and organised entity . . . . . . . . . -551 CHAPTER XXXIX ELECTROTONUS « ctra-polar effects of electrotonic currents on vegetal nerve— Electrotonic variation of excitability — Bernstein's polarisation decrement — Hermann's polarisation increment— Investigation into the law of electrotonic varia- tion of conductivity — Investigation on variation of excitability— Con- ductivity enhanced when excitation travels from places lower to higher electric potential, and depressed in opposite direction — When feeble, anode enhances and kathode depresses excitability — AH electrotonic phenomena reducible to combined action of these factors — Explanation of apparent anomalies .......... 560 CHAPTER XL INADEQUACY OF PFLUGER'S LAW Reversal of Pfluger's Law under high E.M.F. — Similar reversals under ^^^ feeble E.M.F. — Investigation by responsive sensation — Experiments on ^B living wounds — Under moderate E. M. F. , intensity of sensation enhanced I^H at kathode, and depressed at anode — Under feeble E.M.F., sensation ^^H intensified at anode and depressed at kathode — Application of electrical XXVI COMPARATIVE ELECTRO-PHYSIOLOGY CHAPTER XLI THE MOLECULAR THEORY OF EXCITATION AND ITS TRANSMISSION PAGE Two opposite responsive manifestations, negative and positive — Such opposite responses induced by polar effects of currents of different signs — Arbitrary nature of term ' excitatory ' — Pro-excitatory and anti-excita- tory agents — Molecular distortion under magnetisation in magnetic sub- stances—Different forms of response under magnetic stimulation — Mechanical, magneto-metric, and electro-motive responses — Uniform magnetic responses — Response exhibiting periodic groupings — Ineffec- tive stimulus made effective by repetition — Response by resistivity- variation— Molecular model— Response of inorganic substance to electric radiation — Effect of rise of temperature in hastening period of recovery and diminishing amplitude of response — Sign of response reversed under •feeble stimulation — Conduction of magnetic excitation — The Magnetic Conductivity Balance — Effect of A-tonus and K-tonus, on excitability and conductivity — Conducting path fashioned by stimulus — Transmission of excitation temporarily blocked in iron wire, as in conducting nerve — Artificial nerve-and-muscle preparation 587 CHAPTER XUI MODIFICATION OF RESPONSE UNDER CYCLIC MOLECULAR VARIATION Anomalies of response — Explicable only from consideration of antecedent molecular changes— Continuous transformation from sub-tonic to hyper- tonic conditions — Two methods of inquiry, first by means of character- istic curves, second by progressive change of response — Abnormal re- sponse characteristic generally of A or sub-tonic state — Abnormal trans- formed into normal, after transitional b state — b state characterised by staircase response — Responses at c stage normal and uniform — At stages D and E responses undergo diminution and reversal — Responsive pecu- liarities seen during ascent of curve, repeated in reverse order during descent — All these peculiarities seen not only in living but also in in- organic substances, under different methods of observation — Elucidation of effect of drugs — Response modified by tonic condition and past history 615 CHAPTER XLlll CERTAIN PSYCHO-PHYSIOLOGICAL PHENOMENA — THE PHYSICAL BASIS OF SENSATION Indications of stimulatory changes in nerve : i , Electrical ; 2, Mechanical — Transmission in both directions — Stimulatory changes in motor and CONTENTS xxvii sensory nerves similar — Responsive molecular changes and the correlated tones of sensation — Two kinds of nervous impulse, and their character- istics — Different manifestations of the same nervous impulse determined by nature of indicator — Electrical, motile, and sensory responses, and their mutual relations — The brain as a perceiving apparatus — Weber- P'echner's Law — Elimination of psychic assumption from explanation of particular relation between stimulus and resultant sensation — Explana- tion of the factor of quality in sensation — Explanation of conversion from positive to negative tone of sensation after tetanisation —Various effects of progressive molecular change in nerve — Effects of attention and inhibi- tion — Polar variations of tonus, inducing acceleration and retardation . 644 CHAPTER XLIV DISSOCIATION OF COMPLEX SENSATION )nversion of pleasurable into painful sensation, and vice versa, by electro- tonus— The Sensimeter — Mechanical stimulation — Stimulation by ther- mal shocks — Chemical stimulation — Opposite effects of anode and kathode — Normal effects reversed under feeble E.M.P\ — Negative tone of sensation blocked by alcohol and ansesthetics — Separation of positive and negative sensations, by lag of one wave behind the other — Dissocia- tion of sensation by depression of conductivity — Abolition of the negative or painful element by block of conduction 666 CHAPTER XLV MEMORY 677 CHAPTER XLVI REVIEW OF RESPONSE OF ISOTROPIC ORGANS • • 687 CHAPTER XLVn REVIEW OF RESPONSE OF ANISOTROPIC ORGANS • 7^0 CHAPTER XLVni ^REVIEW OF RESPONSE OF NERVE AND RELATED PSYCHOLOGICAL PHENOMENA 7l8 CLASSIFIED LIST OF EXPERIMENTS 735 DEX . , 747 f ILLUSTRATIONS IS. i6. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. PAGE Series of Contractile Responses in Muscle ' i Response of Indiarubber ......... 2 Response of Selenium to the Stimulus of Light 3 Negative Response of Galena to Hertzian Radiation .... 3 Positive Response of Ag' to Electric Radiation 4 Electric Response in Metals . . . . . ' . . . 5 Uniform Electric Response in Tin ....... 6 Fatigue in the Electric Response of Metals ...... 7 Stimulating Action of NagCO^ on Electric Response of Platinum . 8 Abolition of Response in Metal by Oxalic Acid ..... 9 Response by Method of Relative Depression ..... 9 Arrangement for observing Simultaneous Mechanical and Electrical Responses 17 Simultaneous Mechanical and Electrical Responses in Biophytum . 19 Photographic Record of Electrical Response by Galvanometric Negativity of Pulvinus of Mimosa^ when leaf is physically restrained from falling ......... 20 Method of Transmitted Stimulation . 24 Excitation by Sudden Tension ........ 25 Excitatory Response to Tension and Compression .... 26 The Mechanical Tapper ......... 26 The Torsional Vibrator ......... 27 The Vibratory Stimulator 30 Complete Apparatus for Method of Block and Vibratory Stimulation . 31 Influence of Suddenness on the Efficiency of Stimulus . . . . 32 Spring Attachment for obtaining Vibration of Uniform Rapidity . 33 Additive Effect 34 Response Recorder 34 Photographic Record of Uniform Responses (Radish) . . . . 36 Stimulation by Thermal Shocks 38 Photographic Record of Uniform Response in Petiole of Fern to transmitted excitation ......... 39 Taps of increasing strength 1:2:3:4 producing increased response in leafstalk of turnip ......... 39 XXX COMPARATIVE ELECTRO-PHYSIOLOGY FIG. 30. Increased Response with Increasing Vibrational Stimuli (Cauliflower- stalk) 40 31. Responses to Increasing Stimulus obtained with Two Specimens of Stalk of Cauliflower 41 32. Genesis of Tetanus in Muscle 43 33. Photographic Record of Genesis of Tetanus in Mechanical Response of Plants (Style of Datura alba) 43 34. Fusion of Efi"ect of Rapidly Succeeding Stimuli . . - . . 43 35. Response of (a) quickly reacting Amaranth ; {b) of sluggish Colocasia 45 36. Arrangement for Instantaneous Stimulation . . . . . . 47 37. General Arrangement for Rheotomic Observation .... 47 38. Enlarged View of Balanced Keys 49 39. Curve showing Rise and Fall of Responsive E.M. Change, under moderate stimulation 51 40. Response Curve from Rheotomic Observation in Stem of Amaranth under strong stimulation 52 41. Artificial Hydraulic Response of il//w<7ja 55 42. Experimental Arrangement for obtaining Records on Smoked Drum of Responses given to Direct and Indirect Stimulation, by Leaf of Mimosa 56 57 58 60 43. Mechanical Responses of Leaf of Mimosa 44. Mechanical Response of Biophytum to Thermal Stimulation . 45. Record of Response of Mimosa Leaf, taken on a fast -moving drum 46. The Abnormal Positive preceding the Normal Negative in Mechanical and Electrical Responses in Biophytum 61 47. Photographic Record of Electrical Response of Petiole of Cauliflower. 62 48. Photographic Record of Electrical Responses of Potato-tuber . . 64 49. Photographic Record of Electrical Response of Petiole of Fern . . 66 50. Longitudinal Contraction and Retardation of Growth under Light in Hypocotyl of Sinapis nigra 73 51. Record of Growth in Crinum at Temperature of 34° C. and 35° C. . 74 52. Balanced Record of Variation of Growth in Flower bud of Crinum Lily under Diffuse Stimulation of Light 79 53. Diagrammatic Representation of the Tonic Level .... 82 54. Photographic Record of Abnormal Positive passing into Normal Negative Response in a Withered Specimen of Leaf-stalk of Cauliflower ........... S4 55. Photographic Record of Staircase Response in Vegetable Nerve . 91 56. Staircase Increase in Electrical Response of Petiole of Bryophyllum rendered sluggish by cooling 91 57. Photographic Record of Uniform Responses (Radish) ... 92 58. Photographic Record of Uniform Response in Petiole of Fern . . 92 59. Record showing Diminution of Response, when sufficient Time is not allowed for Full Recovery 93 60. Fatigue in Celery .......... 93 61. Fatigue in Leaf-stalk of Cauliflower 93 62. Photographic Record showing Fatigue in Tin Wire which had been continuously stimulated for Several Days . . . . . . 94 ILLUSTRATIONS xxxi FIG. 63. Effect of Over-strain in producing Fatigue 94 64. Rapid Fatigue under Continuous Stimulation in {a) Muscle ; {d) Leaf- stalk of Celery (Electrical Response) 96 65. Photographic Records of Normal Mechanical Response of Mimosa to Single Stimulus (upper figure), and to Continuous Stimulation (lower figure) .......... 96 66. Effect of Continuous Vibration (though 50°) in Carrot . . . . 97 67. Oscillatory Response of Arsenic acted on Continuously by Hertzian Radiation ........... 98 68. Alternate Fatigue {a) in Electrical Responses of Petiole of Cauli- flower ; {b) in Multiple Electric Responses of Peduncle of Biophytuni ; {c) in Multiple Mechanical Responses of Leaflet of Biophytuvi ; and {^d) in Autonomous Responses of Destnodiuni . . 98 69. Photographic Record of Periodic Fatigue in the Automatic Pulsation . of Desmodiu7n gyrans ... . . . . . . '99 70. Periodic Fatigue in Pulsation of Frog's Heart (Pembrey and Phillips) 99 71. Photographic Record of Periodic Fatigue under Continuous Stimula- tion in Contractile Response (Filament of Uriclis Lily) . . .100 72. Fatigue in the Contractile Response of Indiarubber . . . . loi 73. Reversed Response of Fatigued Nerve . . . . . .102 74. Preliminary Staircase, followed by Fatigue, in the Responses of Muscle (Brodie) 103 75. Preliminary Staircase, Increase, followed by Fatigue, in the Response of Galena to Hertzian Radiation 103 76. Photographic Record of Responses of Style of Datura alba in which ' , Qrowth had come to a Temporary Stop 104 77. Differential Contractile Response of Artificial Strip .... 108 78. Responses of Mimosa to Sunlight of not too long Duration . . . 109 79. Transverse Response of Pulvinus of /1/m(?ja . . . . .110 80. Diametric Method of Stimulation of an Anisotropic Organ . . . 1 1 1 81. The Thermal Variator 113 82. Responsive Current in Petiole of Mtisa from Concave to Convex Side 1 15 83. Parallelism of Natural Current in Pulvinus of Mimosa and Sheathing Petiole of iT/«j-rt 118 84. Effect of Variation of Temperature on Natural Current, \, , which in P,etiole of i^///ja flows from Convex to Concave Side . . .119 85. Photographic Record showing effect of Sudden, followed by steady Rise of Temperature on Natural Current, I, in J/wj-a . . . 120 86. Action of 7 per cent. Solution of Na^COj on Natural Current of Musa 122 87. Effect of CO2 on Natural Current of yT///^rt 12^ 88. Variation of the Transverse Natural and Responsive Currents in , VyAv'wiVA oi Mimosa . . . . . . . . . . 127 89. Photographic Record of Effect of Chloroform on Responses of Carrot . 13d 90. Photographic Record showing Action of Chloral Hydrate on the . Responses of Leaf-stalk of Cauliflower 131 91. Photographic Record showing Action of Formalin (Radish) . . . 132 92. Abolition of Response at both a and b Ends by the Action of NaOH 134 XXXll COMPARATIVE ELECTRO-PHYSIOLOGY FIG. I'AGR 93. Photographic Record showing the nearly complete Abolition of Response by strong KOH 135 94. Photographic Record showing the Stimulatory Action of Solution of Sugar 136 95. Photographic Record showing Continuous Action of 2 per cent. NslJCOs Solution 136 96. Photographic Record sfiowing the Depressing Action of 5 per cent. HCl Acid 138 97. Photographic Record showing Effect of i per cent. KHO . . . 138 98. Photographic Record of Effect of 5 per cent. KHO . . . .139 99. Striking-rods for stimulation of two ends of specimen and inducing phase-difference 143 100. Isolated and diphasic responses with increasing difference of phase . 144 1 01. Photographic Record showing Negative, Diphasic, and Positive Resultant Responses in Tin . . . . . . .146 102. Photographic Records of Response of ^rj/^-^^y/Zw/w . . . . 147 103. Photographic Record of Response of Petiole of Cauliflower by the Diametric Method . . . . , . . . .147 104. Distribution of Electrical Tension in Muscle-cylinder . . . . 150 105. Photographic Record showing Persistent Electrical After-Effect in Inorganic Substance under Strong Stimulation . . . • 151 106. Photographic Record exhibiting Persistent Galvanometric Negativity in Plant Tissue after Strong Stimulation . . . . . 152 107. Experimental Arrangement for determining Electrical Effect due to Section 154 108. Records showing increasing Persistent Galvanometric Negativity, according as injury is caused nearer to proximal contact . • . . 154 109. Curve showing the Electrical Distribution in Stem with one Sectioned End. 155 no. Electrical Distribution in Plant-cylinder with Opposite Ends Sectioned 156 111. Record of Responses in Plant (Leaf-stalk of Cauliflower) by Method of Negative Variation . . . . . . . -159 112. Response by Positive Variation of Resting Current . . . . 165 1 1 3. Distribution of Electric Potential in Lamina of Colocasia along a radial line from dead to living through intermediate stages . . .169 114. Straight Form Potentiometer . . . . • , . . 171 115. Distribution of Electric Potential in Petiole of Nymphea alba^ one end of which has been killed 172 116. Photographic Records of Responses of Vegetable Nerve, one end of which has been injured ......... 175 117. Typical Cases of Variation of Current of Rest and Action-Current. Specimen originally isotropic 175 118. Typical Cases of Variation of Current of Rest and Action-Current ; intermediate point naturally less or more excitable than either of terminal . . . . . . . . . . . 176 119. Typical Cases of Variation of Current of Rest and Action Current. Anisotropic organ, B end originally more excitable than a . .177 ILLUSTRATIONS xxxiii PI<^' PAGE 1 20. Photographic Record showing Effect of Rapid Cooling, by Ice-cold Water, on FuhaUons of 7)es7Hodtum gyrans . . . . . 181 121. Photographic Record of Pulsations oi Desjiiodium during Continuous Rise of Temperature from 30° C. to 39° C. . . . .182 122. Diminution of Response in Eucharis by Lowering of Temperature . 183 123. After-effect of Cold on Ivy, Holly, and Eucharis Lily . . .184 124. Photographic Record of Responses in Eucharis Lily during the Rise and Fall of Temperature . . . . . . . . 185 125. Diminished Amplitude of Response with Rising Temperature. (Stem oi Amarajith) . . . . , . , . . .186 126. Photographic Records of Autonomous Pulsations in Desmodium, showing Increase of Amplitude and Decrease of Frequency, with Lowering of Temperature . . . . . ... 188 127. Photographic Record showing Effect of Steam in abolishing Response 190 128. Record of Electric Responses oi Amaranth at various Temperatures . 195 129. Photographic Record of Thermo-mechanical Curve given by Coronal Filament of Passiflora . . . . . . . . . 198 1 30. Thermo-mechanical Curve of Two Different Specimens of Style of Datura alba, obtained from Flowers of the same Plant . .198 131. The Thermal Chamber 200 132. Photographic Record exhibiting Electric Spasm in the Petiole of Musa ........... 202 133. Photographic Record showing Electric Inversion at Death-point, 59'5°, in the Petiole of y^;//«ra«//4 . . . . . . 203 134. Record showing Inversion of Electric Curve and Simultaneous Reversal of Electric Response in Stem of Amaranth . . . 204 135. Multiple Mechanical Response of Biophytum^ due to a Single Strong Thermal Stimulus 208 136. Multiple Electro-tactile Response in Stem of Mimosa due to Single Strong Thermal Stimulus 209 137. Photographic Record of Multiple Electrical Response in Leaf of Biophytum ........... 209 138. Multiple Electrical Responses under Different Forms of Stimulus in Different Organs . . . . . . . . . 210 139. Photographic Record of Multiple Electrical Response to Single Thermal Shock in Frog's Stomach . . . . . . 210 140. Induction of Autonomous Response in Biophytum at Moderately High Temperature of 35° C. . . . . . . .7,11 141. Initiation of Multiple Response in Lateral Leaflet of Desmodium originally at Standstill . . . . . . . ..212 142. Photographic Record of Autonomous Mechanical Pulsation in Desmodium Leaflet . . . . . . . . .213 143. Spark-record of Single Pulsation in Leaflet of Z)^jw<7a?2Vz7/^«Va /«rf/Va . 256 164. Photographic Record of Normal Responses given by Pitcher of Nepenthe^ under Equi-alternating Electric Shocks . . .256 165. Responsive Currents in Lead Wire 264 166. Flat Strip of Lead, of which lower Surface is Brominated . . 265 167. Photographic Records of After-effect of Homodromous \ and Hetero- dromous \ Induction-shocks in prepared Strip of Lead . . . 266 168. Photographic Record of Responses to Equi-alternating Electric Shocks in Prepared Lead Strip 267 169. Response of Pulvinus of Mimosa to Equi-alternating Electric Shocks 268 170. Experimental Arrangement for Determination of Excitatory After- effect of Equi-alternating Electrical Shocks 276 171.. Method of Direct Effect of Excitation by Equi-alternating Shocks . 280 172. Excitation by Equi-alternating Shocks 281 173. Photographic Record of Response of Petiole of Musa to Equi-aller- nating Electric Shocks, before and after Application of Chloroform 284 ILLUSTRATIONS XXXV FIG. PAGE 174. Photographic Record of Responses of Plagiotropic Stem of Cucurhita to Equi-alternating Electric Shocks ...... 285 175. Electrical Responses of Eel to Equi-alternating Electrical Shocks . 285 176. Rotary Mechanical Stimulator 291 177. Diagram Representing Dififerent Levels of Excitability, Plus, Zero, and Minus . . - 296 178. Electrical Response of Grape-skin to Rotary Mechanical Stimulation 299 179. Electrical Response of Frog's Skin to Rotary Mechanical Stimulation 300 180. Photographic Record of Electrical Responses of Upper Surface of Intact Human Forefinger to Rotary Mechanical Stimulation . 301 181. Photographic Record of Electrical Responses of Grape-skin to Thermal Shocks 301 182. Photographic Record of Electrical Responses of Grape-skin to Stimu- lation by Equi-alternating Electrical Shocks .... 302 183. Photographic Record of Series of Electrical Responses of Frog's Skin to Equi-alternating Electrical Shocks 302 184. Photographic Record of Transverse Response of Pulvinus of Mimosa to Equi-alternating Electrical Shocks ...... 303 185. Continuous Photographic Record of Autonomous Pulsation of Des- jnodium gyrans from 6 P.M. to 6 A.M. . . . . . . -tp^ 186. Photographic Record of Electrical Responses in Skin of Neck of Tortoise to Stimulus of Equi-alternating Electrical Shocks . . 306 [87. Isolated Responses of Upper and Lower Surfaces of Skin of Tomato to Rotary Mechanical Stimulus 307 [88. Photographic Record of Series of Responses in Skin of Tomato under Equi-alternatin Electrical Shocks 308 189. A Single Response of Skin of Tomato to Equi-alternating Shock recorded on Faster Moving Drum 309 190. Photographic Record of Series of Normal Responses in Skin of Gecko . . . . . . . . . . .310 [91. Photographic Record of Abnormal Diphasic Responses in Skin of Gecko, converted to Normal, after Tetanisation . . . . 311 "192. Transverse Section of Tissue of Hollow Peduncle of Uriclis Lily . 313 193. Photographic Record of Responses of Water-melon to Equi-alter- knating Electric Shocks . . . . . . . -'SiS 94. Photographic Rec<^ d of Electrical Responses of Intact Human Armpit . . ........ 319 195. Experimental Arrangement for Response of Human Lip . . . 320 196. Photographic Record of Electrical Response of Intact Human Lip . 321 197. Possible Variations of Responsive Current, as between Two Surfaces A and B, shown by Means of Diagrammatic Representations of Characteristic Curves . . ...... 324 ^98. Photographic Record showing Reversal of Normal Response in Pulvinus of Mimosa due to Fatigue ...... 326 ^99. Photographic Record showing Reversal of Response in Carpel of Dillenia indica^ under Sub-minimal Stimulation . . . . 328 Pitcher of Nepenthe^ with lid removed ...... 335 Glandular Surface of a Portion of the Living Membrane of the Pitcher oi Nepenthe 336 xxxvi COMPARATIVE ELECTRO-PHYSIOLOGY FIG. PAGE 202. Transverse Section of Tissue of Pitcher of Nepenthe. . . . 336 203. Photographic Record of Series of Normal Negative Responses of Glan- dular Surface of Nepenthe in Fresh Condition to Equi-alternating Electric Shocks 338 204. Photographic Record of Responses of Pitcher in Intermediate Stage, having Attracted a Few Insects . . . . . . . 340 205. Photographic Record of Responses of Pitcher in Third Stage, the whole Glandular Surface thickly Coated with Insects . . . 340 206. Multiple Response of Pitcher of Nepenthe, in First or Fresh Stage, to Single Strong Thermal Shock . . . . . . • 34i 207. Multiple Response of Pitcher of Nepenthe^ in Third Stage, to Single Strong Thermal Shock 341 208. Photographic Record of Responses in Fresh Leaf of Drosera to Equi- alternating Electrical Shocks 342 209. Photographic Record of Multiple Response of Leaf of Drosera in Positive Phase . . 343 210. Photographic Record of Normal Negative Responses of Frog's . Stomach to Mechanical Stimulation . . . . . '345 211. Photographic Record of Normal Negative Responses of Stomach of Tortoise to Stimulus of Equi-alternating Electric Shocks . . . 345 212. Photographic Record of Normal Response in Stomach of Gecko to Equi-alternating Shocks, seen to be reversed after Tetanisation . 346 213. Photographic Record of Multiple Responses in Stomach of Frog to a Single Strong Thermal Shock 347 214. Photographic Record of Normal Negative Response of Young Root of Colocasia 353 215. Photographic Record of Positive Response in Older Root of Colocasia 354 216. Photographic Record of Electrical Response of Sap-wood . . . 362 217. Phot(^aphic Record showing Normal Responses of Living Wood to Vibrational Stimulus, and the Abolition of Response by a Toxic Dose of Copper Sulphate ........ 363 218. The Shoshungraph 369 219. Curve showing Normal Suction at 23° C, Increased Suction at 35° C, and the After-effect persisting on Return to Normal Tempera- ture 372 220. Action of Anaesthetics in Abolition of Suction 375 221. Effect of Strong KNOj Solution 376 222. Effect of Strong NaCl Solution 377 223. Record of Blank Experiment showing Absence of any Disturbance of Record from Induction-shocks as such 378 224. Terminal Mode of Application of Stimulus 380 225. Sub-terminal Mode of Application of Stimulus ..... 380 226. Renewal of Suction, Previously at Standstill, by Action of Stimulus . 383 227. Photographic Record of Effect of Stimulus in Enhancing Rate of Suction 383 228. Variation of Latent Period as After-effect of Stimulus . . . . 384 229. Photographic Record showing Variation of Latent Period as After- effect of Stimulus 385 230. Suctional Response under Over-balance 386 ILLUSTRATIONS xxxvii FIG. PAGE 231. Photographic Record of Effect of Stimulus on Over-balance . . 386 232. Photographic Record of Response to Continuous Sub-terminal Stimu- lation 388 233. Experimental Arrangement for Detection of Electrical Change induced at the Point transversely Distal to Point stimulated . . 397 234. Record of Response to Moderate Unilateral Stimulation under the Experimental Arrangement described 398 235. Record of Different Specimen under same Experimental Arrange- ment when Stimulus is first Moderate and then Increased . . 399 236. Mechanical Response of Pulvinus of Mitnosa to Continuous Action of Light from Above ......... 400 237. Electrical Response in the Lower Half of the Pulvinus of Mimosa due to Stinmlation of Distal Upper Half by Light . . . 401 238. Photographic Record of Series of Negative Responses of Petiole of Bryophyllum to Stimuli of Sunlight 402 239. Record of Responsive Growth-variation taken under condition of balance in slightly Sub -tonic Flower-bud of Crinum Lily under Diffuse Stimulation of Light 405 240. Photographic Record of Positive Response of the Petiole of Cauli- flower to Light .......... 406 241. Multiple Mechanical -Response of \.^'A.^tX. oi Biophytuni wrA&x the Continuous Action of Light . . ' . . . . . 407 242. Photographic Record of Multiple Electrical Response in Leaf of Bryophylhim under Continuous Action of Light . . . . 408 243. Diagrammatic Representation of Phasic Alternations, and After-effect in Type I. ......... . 409 244. Photographic Record of Phasic Alternations, showing Direct and After Effects of Light in Type I. , represented by Bryophyllum . 411 245. Diagrammatic Representation of Phasic Alternations, and After Effect in Type III 412 246. Photographic Record of Phasic Alternation, showing Direct and After Effects in Type III., represented by Petiole of Cauli- flower 413 247. Photographic Record of Pair of Responses obtained with a Second Specimen of Cauliflower, representative of Type III. . . .413 248. Experimental Arrangement for Determination of Differential Excita- bility of Optic Nerve and Cornea 417 249. Series of Photographic Records of Excitatory Responses in Frog's Eye to Equi-alternating Electric Shocks 417 250. Experimental Arrangement for Demonstration of Differential Excita- bility as between Retina and Optic Nerve 418 251. Series of Photographic Records of Excitatory Responses in Frog's Retina to Equi-alternating Electric Shocks . . . . , 419 252. Photographic Record of Multiple Response of Retina of Frog under Continuous Action of Light ....... 426 253. Response of petiole of Bryophylhim. Light was cut off on attain- ment of maximum positivity in the second of the multiple • responses . . • • 427 I xxxviii COMPARATIVE ELFXTRO-PHYSIOLOGY FIG. PAGE 254. Similar effect in response of retina of Ophiocephalus fish . . . 427 255. The same with another specimen. Light was here cut off after the first oscillation 427 256. Response of retina of Ophiocephalus when slightly fatigued . . 428 257. Response of frog's eye (Kiihne and Steiner) 428 258. After-effect of Light on Silver Bromide 429 259. Response of petiole of cauliflower. Light was here cut off on attain- ment of maximum negativity 430 260. Response of retina of Ophiocephalus fish when depressed . . . 430 261. Response of isolated retina of fish as observed by Kiihne and Steiner 430 262. Inclined Slits for Stereoscope and Composite Image formed in the Two Eyes 432 263. Composite Indecipherable Word, of which Components are Seen Clearly on Shutting the Eyes 432 264. Diagrammatic Representation of a Multicellular Organ laid Horizon- tally and Exposed to Geotropic Stimulus 435 265. Effect on Apogeotropic Movement of Temporary Application of Cold on Upper and Lower Surfaces respectively 436 266. Diagrammatic Representation of Experiment showing Curvature Induced by Unilateral Pressure Exerted by Particles . . . 437 267. Record of Responsive Curvature Induced in Bud of Crinum Lily by Unilateral Pressure of Particles 437 268. Record of Apogeotropic Response in Scape of Uriclis Lily . . 438 269. Photographic Record of Geo-electric Response in the Scape of Uriclis Lily laid horizontally 440 270. Experimental Arrangement for Subjecting Organ to Geotropic Stimulus, Mechanical Response being Restrained . . . 441 271. Geo-electric Response of the Physically Restrained Scape of Uriclis Lily 442 272. Diagrammatic Representation of Electrical Connections for Deter- mination of Velocities of Centrifugal and Centripetal Trans- missions 447 273. Experimental Arrangement for Comparing the Relative Conduc- tivities in Transverse and Longitudinal Directions . . . 454 274. Response of Frog's Nerve under Simultaneous Excitation of both Contacts, by Equi-alternating Electrical Shocks, one Contact being Injured .......... 457 275. Enhancement of Amplitude of Response, as After-effect of Thermal Tetanisation, in P'rog's Nerve 462 276. Conversion of Abnormal Positive into Normal Negative Response after Thermal Tetanisation 463 277. Gradual Transition from Abnormal Positive, through Diphasic, to Normal Negative Responses in Frog's Nerve .... 464 278. Abnormal Positive Response converted through Diphasic to Normal Negative under the increasingly Effective Intensity of Stimulus, brought alx)ut by Lessening the Distance between the Responding and Stimulated Points 466 279. Frond of Fern with Conducting Nerves exposed .... 469 ILLUSTRATIONS XXXIX FIG. * PAGH 280. Photographic Record of Effect of Ether on the Electrical Respons^pf Plant-nerve \ . 472 281. Photographic Record of Effect of CO.^ on Electrical Response of Plant-nerve 473 282. Photographic Record of Abolition of Response by Strong Application of Alcohol 473 283. Photographic Record of Effect of Ammonia on Ordinary Tissue of Petiole of Walnut ......... 474 284. Photographic Record of Effect of Similar Application of Ammonia on Plant-nerve 474 285. Photographic Record of Exhibition of Three Types of Response, Normal Negative, Diphasic, and Abnormal Positive, in Nerve of Fern under Different Conditions 475 286. Photographic Record of Effect of Tetanisation in Inducing Enhance- ment of Normal Negative Response in Nerve of Fern . . . 476 287. Photographic Record of Conversion of the Abnormal Di-phasic into Normal Negative, after Tetanisation, T, in Nerve of Fern . . 477 288. Photographic Record showing how the Abnormal Positive Response is converted through Diphasic into Normal Negative by the Increasing Effective Intensity of Stimulus, due to Lessening the Distance between the Responding and Stimulated Points . . 477 289. Diagrammatic Representation of the Conductivity Balance . . . 482 290. Photographic Record made during Preliminary Adjustment for Balance of Nerve of Fern ........ 482 291. Complete Apparatus of Conductivity Balance 484 292. Effect of Na2C03 Solution on Responsive Excitability of Frog's Nerve 485 293. Effect of CUSO4 on Frog's Nerve 485 294. Photographic Record showing Enhancement of Responsivity by Application of CaCl.^ 486 295. Photographic Record showing Depression of Responsive Excitability by Application of KCl 487 296. Photographic Record exhibiting Comparative Effects of NaCl and NaBr on Responsivity 487 297. Photographic Record of Effect of Dilute (-5 per cent.) Solution of Na-^COg on Variation of Conductivity 488 298. Photographic Record of Effect of Stronger Dose (2 per cent.) of Na^COg Solution on Conductivity 489 299. Responsivity versus Conductivity under KI . . . . 491 300. Responsivity versus Conductivity under Nal .... 492 301. Effect of Alcohol on the Responsivity of Frog's Nerve . . . 492 302. Photographic Record of Effect of Alcohol Vapour on Receptivity , 493 303. Photographic Record of Effect of Alcohol on Conductivity . . . 494 304. Photographic Record showing Effect of Alcohol on Responsivity . 494 305. Diagrammatic Representation of Experimental Arrangement for Demonstration of Receptivity versus Conductivity, or of Receptivity versus Responsivity 495 306. Receptivity versus Responsivity under Alcohol . . . 495 307. Photographic Record showing Effect of Cooling on Conductivity of Plant-nerve 499 xl COMPARATIVE ELECTRO-PHYSIOLOGY riG. PAGE 308. The Cork Chamber for Gradual Raising of the Temperature of one Arm of the Balance 500 309. Photographic Record Showing Effect of Rising Temperature on Conductivity 501 310. Experimental Arrangement for Studying After-effect of Stimulus on Conductivity and Excitability ....... 504 311. Photographic Record Showing Effect of Moderate Stimulation in Enhancing Conductivity and Excitability ..... 504 312. Photographic Record showing Effect of Excessive Stimulation in De- pressing Excitability and Conductivity . . . . . . 505 313. Record of Contractile Response in Frog's Nerve under Continuous Electric Tetanisation 510 314. Optical Kunchangraph for Mechanical Response of Nerve . . . 511 315. Dij^rammatic Representation of Arrangement for Obtaining Trans- mitted Effect of Stimulus . . . . . . . .512 316. Photographic Record of Effect of Ammonia on Mechanical Response of Frog's Nerve . . . . . . . . . . 516 317. Photographic Record showing Abolition of Mechanical Response of Frog's Nerve by Action of Solution of Morphia . . . .516 318. Photographic Record showing Preliminary Exaltation in Mechanical Response of Frog's Nerve after Application of Alcohol . . . 517 319. Photographic Record showing Effect of Chloroform on Mechanical Response of Frog's Nerve . . . . . . . .518 320. Photographic Record showing Abnormal Positive converted into Negative Response after Tetanisation 521 321. Photographic Records showing Gradual Disappearance of Positive Element in diphasic Mechanical Responses of Frog's Nerve and Plant-nerve 522 322. Photographic Record showing Staircase Effect in Mechanical Re- sponse of Frog's Nerve 524 323. Photographic Reproduction of Record of Mechanical Responses of PVog's Nerve and Plant-nerve obtained on Smoked Glass Surface of Oscillating Recorder 528 324. Record of Mechanical Responses to Electrical Stimulus obtained on Smoked Glass, and given by the Optic Nerve of Fish Ophiocephalus 529 325. Record, obtained on Smoked Glass, of Transmitted Effect of Stimu- lation on Nerve of Gecko ........ 530 326. Initiation of Multiple Response by Drying of Nerve . . . 539 327. Diagrammatic Representation of Experimental Arrangement for Re- cording Response by Resistivity Variation . . ... 544 328. Photographic Record of the Morographic Curve taken by Method of Resistivity Variation in Pistil of Hibiscus. Critical point of in- version at 6o-8° C 546 329. Photographic Record of the Morographic Curve taken by Method of Electromotive Variation in Petiole of Musa. Critical point of inversion at 59"6° C. . . . . . . . . . 546 330. Photographic Record of the Morographic Curve taken by Method of Mechanical Response in Filament of Passijlora. Critical point of inversion at 59-6 C 546 ILLUSTRATIONS xH FIG. 331. Response Records by Resistivity Variation, in the Nerve of Fern . 548 332. Effect of Chlorofornm seen in Modification of Resistivity Variation in Frog's Nerve . 550 333. Photographic Record of Effect of Tetanisation in Enhancing Mechani- cal Response of Plant-nerve ........ 554 334. Photographic Record showing Enhancement of Excitability under Action of Light in Nerve of Fern ...... 557 335. Distribution of Fibro-vascular Elements in Single Layer of Stem of Papaya 558 336. Extra-polar Kat-electrotonic Effect 560 337. Extra-polar An-electrotonic Effect ....... 560 338. Extra-polar Electrotonic Effects under an Acting E.M.F. which rises from -610 I -4 Volts 561 339. Diagram illustrating Bernstein's Decrement of Kat-electrotonic Current 562 340. Diagram illustrating Bernstein's Decrement of An-electrotonic Current 562 341. Diagram representing Hermann's Polarisation-increment under Tetanising Shocks, with reversed polarising Current . . . 563 342. Diagram representing Plermann's Polarisation-increment under Tetanising Shocks, with reversed polarising Current . . . 563 343. Experiment with Petiole of Fern demonstrating Variation of Con- ductivity by Polarising Current, Excitation travelling electrically Downhill ........... 565 344. Experiment with Petiole of P'ern demonstrating Variation of Con- ductivity by Polarising Current, Excitation travelling electrically Uphill 565 345. Photographic Records of Responses taken in last Experiment, when Excitation was transmitted with and against the Polarising Current. 566 346. Photographic Record of Modification of Conduction during Passage of Excitation from Anodic to Kathodic Region, under Increasing Intensity of Polarising E.M.F 567 347. Photographic Record showing Enhanced Conduction from Kathodic to Anodic Region 568 348. Experimental Arrangement to Exhibit the Enhancement of Excita- bility at Anode, when the Acting E.M.P\ is feeble . . . 569 349. Experimental Arrangement to Exhibit Depression of Excitability at Kathode, when the Acting E.M.P\ is feeble 569 350. Photographic Records of Response, illustrating the Enhancement of Excitability at Anode, and Depression at Kathode, under Feeble Acting E.M.F. in two Specimens of Nerve of Fern a and b . . 570 351. Experimental Arrangement demonstrating the Joint Effects of Variation of Conductivity and Excitability by Polarising Current . 572 352. Experimental Arrangement demonstrating the Joint Effects of Variation of Conductivity and Excitability by Polarising Current, when Current is Reversed . . . . . . . -572 353. Photographic Record of Response under the Arrangements given in Figs. 351, 352 in Nerve of Fern 572 354. Experimental Arrangements for Showing so-called Polarisation- increment by the Joint Effect of Increased Excitability at Anode and Enhanced Conduction of Excitation electrically Uphill . -574 C xlii COMPARATIVE ELECTRO-PHYSIOLOGY KIG. PAGE 355. Experimental Arrangements for Showing so-called Polarisation- increment by the Joint Effect of Increased Excitability at Anode and P^nhanced Conduction of Excitation electrically Uphill. Direction of Current in this is Reversed .... 356. Photographic Record of Responses in Nerve of Fern, under Anodic and Kathodic Action as described in Figs. 354 and 355. 357. Photographic Record of Similar Effects in Nerve of Frog . 358. Make-kathode and Break -anode Effects in Biophytum 359. Effect of Anode and Kathode on Responsive Sensation in Human Hand 360. Polar Effects of Currents due to Localised Application on Upper '\^2\{ oiVwWmw^oi Erythrina indica 361. Experimental Arrangement for Magnetometric Method of Record 362. Photographic Record of Uniform Magnetic Responses of Iron 363. Photographic Record of Periodic Groupings in Magnetic Responses 364. Photographic Record of Response and Recovery of Steel under Moderate and strong Magnetic Stimulus .... 365. Photographic Record showing Ineffective Stimulus made Effective by Repetition .......... 366. Molecular Model ......... 367. Method of Resistivity Variation ....... 368. Photographic Record of Response of Aluminium Powder in Sluggish Condition to Stimulus of Electric Radiation 3^9. Photographic Record Showing Uniform Response of Aluminium Powder to Uniform Stimulus of Electric Radiation . 370. Photographic Record of Response of Tungsten 371. Experimental Arrangement for obtaining Response in Iron by In- duction Current 372. Magnetic Conductivity Balance 373. Process of Balancing illustrated by Photographic Record of Responses. 374. Effect of K- and A-Tonus on Magnetic Conduction 375. Opposite Effects of K-Tonus when moderate and strong . 376. Effects of K- and A-Tonus on Magnetic Excitability . 377. Gradual Enhancement of Conductivity by the Action of Stimulus 378. Ciiaracteristic Curve of Iron under increasing Force of Magnetisation 379. Characteristic Conductivity Curve of Sensitive Metallic Particles be- longing to Negative Class, under increasing Electro-motive Force 380. Cyclic Curves of Magnetisation and'of Conductivity 381. Photographic Record of Magnetic Tetanisation of Steel, exhibiting Transient Enhancement of Response on Cessation . 382. Mechanical Response of Frog's Nerve to successive equal Stimuli, applied at Intervals of One Minute 383. Mechanical Response of Frog's Nerve, showing Conversion of Ab- normal Positive into Normal Negative Response after Tetanisation 384. Photographic Record showing Conversion of Abnormal ' Down ' Re- sponse in Tin to Normal * Up,' after Tetanisation . ILLUSTRATIONS xliii FIG. PAGE 385. Gradual Transformation from Abnormal to Normal Response in Platinum ........... 629 386. Normal Electro-motive Response in Tin, enhanced after Tetanisation 629 387. Photographic Record of Abnormal Response of Selenium Cell con- verted into Normal after Tetanisation . . . . . . 630 388. Photograpliic Record showing Moderate Normal Response of Selenium enhanced after Tetanisation 630 389. Photographic Record of Abnormal Response of Tungsten to Electric Radiation, converted after Tetanisation into Diphasic and Normal . 632 390. Moderate Normal Response of Aluminium, enhanced after Tetanis- ation ............ 632 391. Photographic Record of Enhancement of Magnetic Response after Tetanisation .......... 633 392. Vertical Series of Records showing Transformation of Abnormal into Normal Response after Tetanisation in Living and Inorganic alike in the a phase 633 393. Series showing how Tetanisation enhances Normal Response in the B Phase ........... 634 394. Photographic Record showing Responses corresponding with different parts of characteristic curve in frog's nerve . .... 635 J95. Photographic Record of Response of Tungsten showing Enhance- ment of Response after moderate Tetanisation, and Reversal of Response, due to Fatigue under stronger Tetanisation . . . 638 196. Series showing reversal of Normal Response by fatigue due to strong Tetanisation inducing the E phase ....... 639 197. Fatigue in Indiarubber giving rise to Diphasic and Reversed Re- sponses 642 198. Fatigue inducing Diphasic Variation and Reversal of Normal Response in Frog's Nerve 642 199. Abnormal Response of Muscle by Relaxation, followed by Normal Response of Contraction ........ 649 Hoo. Record of Response in Nerve of Gecko showing the Effect of Arith- metically increasing Stimulus 657 . Response of Nerve of Bull-frog to Stimuli I, 2, 3, . . . 12, increasing in Arithmetical Progression ........ 658 02. Response of Optic Nerve of Ophiocephalus to Arithmetically increasing Stimuli I, 2, 3. 4, 5, 6, 7 659 403. Mechanical Response of Nerve of Fern to Arithmetically increasing Stimulus ........... 659 404. Photographic Record of Magnetic Responses in Steel to Arith- metically increasing Stimulus 660 405. The Sensimeter .......... 670 406. Revival of latent Image in Metal 683 YHv^ iuZj^ COMPARATIVE ELECTRO-PHYSIOLOGY CHAPTER I THE MOLECULAR RESPONSIVENESS OF MATTER Response to stimulus by change of form — Permeability variation — Variation of solubility — Method of resistivity variation : (a) positive variation ; (d) negative variation — Sign of response changed under diflferent molecular modifications — Response of vegetable tissue by variation of electrical re- sistance — Response by electro-motive variation in inorganic substances — The method of block — Positive and negative responses— Similar responses in living tissues — Effects of fatigue, stimulants, and poisons on inorganic and organic responses — Method of relative depression, or negative variation, so called. In studying the properties of living tissues, we find one of their most important characteristics is found in the fact that they exhibit the state of excitation under the impact of ^!rv stimulus. On the cessation of stimulus, again, the excited fjj] tissue returns to its original condition. The excitatory change ^ thus undergone is fundamentally due to the derangement, or upset, of the molecules of the living tissue from their normal equilibrium, recovery being brought about by their restoration to that state. The excitatory condition is sometimes shown by change of form, as in the case . of the shortened length of excited ^HHHU^I^^^H muscle (fig. i). This might be ^HI||H|II&Ib^^^^H compared with the shortening of ^^^^^^^^^^^^^* Ai.i-j-j- uu J i-u Fig. I. Series of Contractile Stretched mdia-rubber under ther- Responses in Muscle mal stimulus (fig. 2). Now it is clear that the molecular change consequent on excitation must occasion various concomitant physical '% 2 COMPARATIVE ELECTRO-PHYSIOLOGY changes, and it should be theoretically possible to detect and measure this induced molecular change by recording such concomitant variations. Thus the stimulus of light, for example, may induce a mole- cular change which may in its turn induce, say, a variation x. in the permeability of the sub- stance to liquid. Bichromated gelatine becomes less perme- able under the action of light. The solubility of a substance Fig. 2. Response of India-rubber ^ay again undergo variation Thermal stimulus for i second at ^^^^^ external Stimulus-SUl- intervals of two mmutes. phur, for example, usually soluble in carbon disulphide, is rendered insoluble under the action of light. In order, then, to study the effect of a given stimulus with accuracy, we should be able to detect and measure the extent of the changes induced. The two effects which have just been referred to are not, as will be seen, highly susceptible of accurate measurement. But in the detection of molecular changes by electrical means, we have at our disposal methods for the measurement of such changes, the ease and delicacy of which leave nothing to be desired. Two such methods may be used — that of Resistivity and that of Electro-motive Variation. According to the method of resistivity variation, tTie sifbstance to be experimented on is placed in an electrical circuit, including a delicate galvanometer and a suitable electro-motive force, such as to cause a small deflection of the galvanometer. The impact of the stimulus on the sub- stance under examination now TnHuces "TrT it a molecular cKange by which its resistance is made to undergo a variation, wRlch in the case of certain substances may be an increase, Iortn that of others a diminution. On the cessation of external stimulus, the substance shows recovery, with a corresponding return to its original conductivity. Thus in the case of selenium, for instance, the conductivity is increased, or the .A flUiMWvlfc WfcW \^vii^ ^l\-Nl\>^^CLU4iTii*^ Oa i THE MOLECULAR RESPONSIVENESS OF MATTER I Lt^ j. resistance decreased, under the action of light. In fig. 3 is shown a number of responses to light, given on a series of separate exposures, each of one second's duration, the inter- vening periods allowed for recovery being of one minute each. Fig. 3. Response of Selenium to the Stimulus of Light (Resistivity variation method) (These responses were obtained by recording the increased eflection due to decreased resistance under the impact of ight, and the subsequent recovery. Such responses, by means o f decreased resistance, we may arbitrarily distinguish as negative. Similar responses are also given by a mass of metallic particles when acted upon by electric radiation.. In fig. 4 are seen several of these negative responses given by galena under the action of this stimulus. There are, on the other hand, some substances which ^ 1 ^ . , Fig. 4. Negative Response of Galena %\\^ positive responses ; that is to to Hertzian Radiation say, their resistance is increased, (Resistivity variation method) , ^ or conductivity decreased, under ^^" the action of stimulus. The deflection of the galvanometer under a constant electro-motive force now undergoes diminu- iv ^ , [tion during the impact of stimulus. Such positive responses ^^^S are obtained with potassium and sodiurn. That the sign ^\ '^ COMPARATIVE ELECTRO-PHYSIOLOGY of response does not depend on the electro-positivity or negativity of the substance is seen in the fact that while J highly electro-positive potassium gives positive response, ^^ the equally electro-negative dioxide of lead gives a response ^ of the same sign. Substances like magnesium, aluminium^i^^ and iron give negative response. v It is found, again, that the same substance, under different molecular conditions, will give responses of opposite signs. For example, a particular molecular variety of silver, Ag', gives positive (fig. 5), whereas ordinary silver gives nega- tive response. Again, while Ag' normally gives positive,\V^ yet the sign of this response is gradually reversed to nega^ tive, under the long-continued action of very strong stimulus.. By the employment of the same method of resistivity variation, I have been able to obtain excitatory response records from living tissues also. Details of these will be given in a subsequent chapter. The electric response, however, employed to obtain the excitatory reaction of living tissues, depends upon the electro-motive variation of the substance under stimulation. This electric reaction has been regarded as vitalistic in contradistinction to physical. But I have shown that similar responses are given by inorganic substances also. That is to say, the molecular excitability on which the phenomenon of response depends is not distinctive of animal tissues alone, but is common to all matter, both organic and inorganic. If, then, we desire to understand those funda- mental reactions which underlie the response of living tissues, it will be well to observe its occurrence in the much simpler case of the inorganic body.^ If we take an inorganic substance, say a piece of metal Fig. 5. Positive Response of Ag' to Electric Radiation ' For a Living. detailed account cf. Bose, Kespome in the Living and the Non- //• THE MOLECULAR RESI^ONSIVeNESS OF MATTER 5 wire, and if its molecular condition be the same throughout, it is obvious that its physical properties will likewise be uniform. Hence its electrical condition will also be the same at every point ; in other words, it will be iso-electric. ' But if a portion of this wire should now be made to undergo a molecular change, as, say, by hammering, the physical condition of this portion will be made different from that of the rest. There will, therefore, be an electrical difference, and the wire will no longer be iso-electric. This fact can be verified by making suitable connections between the molecularly strained and unstrained portions of the wire,- and a galvanometer, when a current will be found to flow! through the galvanometer, showing that a difference of electrical potential has been brought about by the induced (^) ic) irmru \ B Fig. 6. Electric Response in Metals (a) Method of block ; {b) Equal and opposite responses when the ends A and B are stimulated ; the dotted portions of the curves show recovery ; (f) Balancing effect, R, when both the ends are stimulated simultaneously. jquality of molecular conditions in different parts of the same wire. I shall now describe the method by which electrical responses to molecular disturbance may be obtained from inorganic substances. For this purpose, two different methods may be employed — first, the method of block, and, secondly, the method of relative depression. According to the first of these, the wire to be experimented on is held clamped at the middle, electrical connections being made with a galvanometer at two points, A and B, by means of two non-polarisable electrodes (fig. 6). We may now produce vXjA'^'^^^'^''''*"'^ K • 6 COMPARATIVE ELECTR0-PUVSlOLui.\ excitation of the A end of the wire, by infparting a torsions vibration, the molecular disturbance being prevented from reaching the B end by the intervening block. Using this method of experiment, I have obtained with different sub- stances two different types of response— namely, positive and negative. In the positive, the responsive current flow^ through the wire from the unexcited to the excited, <5ra towards the excited — that is to say, the excited point becomes galvanometrically positive. Responses of this kind are given b y tin, zinc, plat inum, and other metals. In fig. 7 IS seen a uniform series of such responses to uniform stimulus. The intensity of the response, moreover, does not appear to depend on the chemical activity of the substance. For the response of the chemi- cally inactive tin is much stronger tha n that of the active zin c. The very inactive platinum is also found to give a fairly strong respon.se, although the electrolytic j Fig. 7. Uniform Elec- tric Response in Tin contacts are made with pure water. The electro-motive response may also be obtained by other modes of molecular excitation. Thus, instead of torsional, we may use longitudinal vibration. A metallic rod of brass, A C, is clamped in the middle. A thin copper wire is led sideways from the clamp and connected with a piece of brass, B. A and B are connected with a galvanometer by means of noh-polarisable electrodes. If now the C end of the rod be rubbed with resined cloth, A may be thrown into longitudinal vibration, B being little affected by this. It is here interesting to observe the concomitance of the electrical response with the sonorous icsponse of the rod, and the dying of the electrical response with the waning of the musical note. The stronger the molecular vibration, the stronger the sound, and also the stronger the response. The direction of the responsive current in the metal is from the less excited B to the more excited A. We found under the method of conductivity variation THE MOLECULAR RESPONSIVENESS OF MATTER that when a substance is molecularly modified, the sign of its response tends to be reversed. Thus, as already said, ordinary silver gives positive, and modified silver, Ag', negative response. But the latter, under strong, and long- continued stimulation, has its response re-converted, as it were, to the normal positive. In the same way, under the method of electro-motive variation also, we find the normal positive response of, say, tin, or platinum, becoming con- verted by molecular modification into negative, to be again re- converted under continuous stimulation to the normal positive. There are, again, certain other substances, of which the normal response is negative. Thus a wire of brominated lead, for instance, when suitably prepared, is found to give an electro-motive response in which the current flows from the excited to the unexcited or away from the excited, the excited point becoming galvanometrically negative. These electro-motive responses of the inorganic have thus the same characteristics as those which have been observed in the case of animal tissues. Certain tissues, such as highly excitable muscle and nerve, give negative response — that is to say, the excited point omes galvanometrically gative. Other tissues, again, the skin for example, give positive response. The normal responses, moreover, are sometimes found to be reversed under molecular jjfV 'modification, and to be re- "JS* reversed to normal response ^ under long-continued stimu- Jfk lation. The electrical responses I of metals, again, are subject to an increase or decrease which is paralleled by the same phenomenon in the response of animal tissues under similar circumstances. That is to Fig. 8. Fatigue in the Electric Response of Metals / 8 COMPARATIVE ELECTRO-PHYSIOLOGY say, fatigue is found to depress the response of the inorganic as of the organic (fig. 8). As in the case of animal tissues, again, so also in that of metals, certain chemical substances act as excitants, enhancing the response (fig. 9), others as depressors and still a third class — such as oxalic acid — as poisons, abolishing response altogether (fig. 10). By taking advantage of the last of these facts, we arrive at a second means of obtaining response — that is to say, the method of relative depression. If both the contact points .uum ILL Before f After Fig. 9. Stimulating Action of Na^COj, on Electric Response of Platinum Records to the left exhibit response before, to the right after the application of reagent. A and B be equally excited — that is to say, subjected to diffuse stimulation — the responsive currents will be opposed, and there will be no resultant galvanometric effect. This was overcome, according to the method of block, by localising excitation at one point, say, A ; we might, however, neutralise altogether the counteracting excitatory effect at B by abolishing the excitability of that point, as, say, by the application of oxalic acid, A being left in its normal con- dition. If now the wire be subjected to diffuse stimulation THE MOLECULAR RESPONSIVENESS OF MATTER by vibrating it as a whole, a resultant response will occur. But by the application of oxalic acid to one contact, a resting or permanent current has been induced in the circuit. The responsive or action current originated under stimulation is now found to flow in a direction opposite to that of this" resting current — that is to say, it causes a negative variation of it (fig. ii). The method of response by the so-called negative variation, which is generally employed in studying re- sponsive phenomena in nimal tissues, is in reality, s will be seen later, a form f this method of relative Before j- After Fig. io. Abolition of Response in metal by Oxalic Acid lepression. Various means have been described, in the course of this :hapter, for the detection and record of that excitatory change ^hich is brought about by the upsetting of molecular equi- librium under stimulus, and the subsequent re- covery. The responsive ■■fchange may find expres- sion in different ways. This expression may, for instance, in the case IKDf living tissues, take the form of mechanical con- t raction, or of the el ec- t rical variation of ga l- vanometric negativit y. Or the opposite change, expressed mechanically as ex- p ansion, will be evidenced by galvanometric positlvity. But I Bit must be borne in mind that neither of these expressions Fig. 1 1. Response by Method of Relative Depression \ represents current of rest ; | represents the action current. h.M^ 10 COMPARATIVE ELECTRO-PHYSIOLOGY is consequent on the other It would be as incorrect to suppose that the electrical effect depended on the mechanical, as to assume that the mechanical was brought about by the electrical. The two are independent expressions of the same fundamental molecular change, brought about by the shock of stimulus. Again, those various responsive phenomena and their modi- fications which are the subject of our inquiry, are such as are induced by different external agencies. Under the influence of certain conditions, the responses of living matter undergo an abolition — the change which we associate with death. In inorganic matter also, we find a similar change of responsive- ness into irresponsiveness, to take place. The death-change In the case of living matter is thus not due to a change from the organic to the inorganic condition, but to some molecular :ransformation. And the nature of this very obscure trans- formation may one day be elucidated by a careful study of 'the changes which take place in inorganic matter, when it passes from responsivity to irresponsivity. The word * physiological ' is generally used to distinguish phenomena which are believed to be exclusively characteristic of the properties of living matter. Such phenomena, however, are found, as our power of investigation grows, to be increas- ingly capable of analysis into physico-chemical processes. In my own use of the term 'physiological,' therefore, it will be understood as a convenient expression for describing the response-phenomena of plant or animal tissues, but as in no sense opposed to the word ' physical.' We shall, in the following chapters, study excitatory effects in living tissues, and their variations under different conditions, using the methods of electrical response. These phenomena will be studied with special detail in the case of vegetable tissues, and it will be found that there is no responsive reaction exhibited by any one amongst the various types of animal tissues, which has not its exact corre- spondence in the vegetable. Those anomalies, further, which have been observed in the response of the animal, will be seen THE MOLECULAR RESPONSIVENESS OF MATTER II to be fully elucidated by the study of similar phenomena under the simpler conditions of the plant. And finally an attempt will be made to arrive at some generalisation which will show the continuity between the simplest form of response in the inorganic and the most complex which occur in the highest type of animal tissue. fi (/> I'Ai ... ^rtl Ci^Jf^-:, i^^ ^;;j}W\ THE ELECTRO-MOTIVE RESPONSE OF PLANTS 1 3 points of the midrib of the leaf On stimulation of the lamina, this current was found to undergo a diminution, or negative variation. But the current in the stalk, or petiole was found to undergo the opposite change — that is to say, a positive variation. Kunkel, in working with the pulvinus of Mimosa, found that on excitation a series of opposite or oscillatory electro-motive variations was induced. He also found electro-motive differences to be induced as the result f the flexure or injury of ordinary stems He believed all hese electrical phe'nomena to be consequent on hydrostatic isturbance or water-movement. By means of this ' migration of water' Kunkel attempted :o explain all the electrical phenomena in vegetable organs, he electro-motive difference between different parts of an irgan was due, according to him, to their different powers of absorption of water. The greater absorptiveness of one point, with its consequent greater movement of water, would render that point relatively positive. As against this, how- ver, Haake pointed out that electrical differences between different points were also to be found, even in submerged plants, like Valisnaria and Nitella, in which there could not possibly be any differences of absorptiveness. This difference, therefore, he suggested, must be ascribed to some vital ^ process, inasmuch as the P.D. is seen to undergo a change whenever the respiratory process is interfered with, as, say, by the substitution of hydrogen for oxygen. We shall study later in greater detail the conditions on hich this ' current of rest,' so called, actually depends (Chapter X.). But more important is the excitatory variation induced by stimulus. It has already been stated that Kunkel found oscillatory variations of the current to occur on stimulation of the leaf of Mimosa. These alternating variations were difficult to reconcile with his theory of the active, single-phased displacement of water, as he did not fail to see, and he suggested that the first negative swing ^■observed might be due to the disturbance of the diffusion- process through alterations of the protoplasm. xt \m.^ I 14 COMPARATIVE ELECTRO-PHYSIOLOGY Munk attempted to explain the complicated electrical effects which he observed in Dioncea by assuming the existence of two different kinds of electro-motive elements, affected in opposite ways, the maximum changes being initiated in one set earlier than in the other. In this way, he thought, it might be possible to explain the occurrence of positive and negative variations, holding that the upper parenchymatous layer of the leaf, and the upper midrib, went through the negative, and the under layer and the under midrib through the positive change, Burdon Sanderson, in his ' fundamental experiment ' on the lamina of Dioncea, had his led-off circuit connected with the upper and lower surfaces of one lobe, stimulus being applied at the other. In the experiments described in the ' Phil. Trans.' of 1882, he found that the upper or inner surface of the leaf become positive on excitation. This he regarded as the true excitatory change. The upper contact now, however, after a certain interval became negative, a change which Burdon Sanderson designated as the after-effect. This after- effect he ascribed to the electrical variations caused by that movement of water which had been observed by Kunkcl. But with regard to the preceding positive variation he says : .Wjp f^The excitatory disturbance which immediately follows V ^>\ '\^,<| excitation is an explosive molecular change, which by ^ * ^ jthe^'mode'^oFlts origin7the suddenness of its incidence, I and the rapidity of its propagation is distinguished from every other phenomena except the one with which I have identified it, namely, the corresponding process in the excitable tissues of animals. . . . The direction of the excitatory effect in the fundamental experiment is such as to indicate that in excitation excited cells become positive to unexcited, whereas in animal tissues excited parts always become negative to unexcited.' ^ In a subsequent series of experiments, however, given in *Phil. Trans.' for 1888 Burdon Sanderson finds the reaction ' Phil. Trans, vol. clxxiii. i»^ 11° THE ELECTRO-MOTIVE RESPONSE OF PLANTS 1 5 of leaves ' in their prime ' to be somewhat different. A strong negative phase was now observed on stimulation, but pre- ceded by a short-lived positive reaction. These results will be discussed in detail in a subsequent chapter, but it may be said here that from the records given by Burdon Sandersori it is difficult to know which of the various responsive phases are to be regarded as those of true excitation, and which as the results of some other cause. It will thus be seen that the results arrived at and the theories advanced by different observers in this field are some- what at variance with each other. This must have been due the difficulties met with in disentangling the fundamental , reaction from those various subsidiary effects which are apt to be found in combination with it. Chief among these difficulties was (i) the fact that positive and negative variations are generally measured in terms of an existing current of rest. But, as a matter of fact, these two apparently opposite responsive variations, positive and negative, are not always indicative of opposite reactions. For an identical excitatory reaction, added algebraically to the resting-current, might appear, according to circumstances, as either of the two. There was also (2) the difficulty of discriminating two opposed electrical effects, one of which was due, as I shall show, to true excitation, and the other to increase of internal energy brought about by mechanical movement of water. Under different conditions, it may be either the one or the other of these which becomes prominent. I shall hope to show that each of them is definite and dis- tinguishable from the other. With regard to the response of plants in general, then, it may be well to state here that the first fact to be demon- strated in the course of the present work is that such responsive phenomena as may be observed in the case of sensitive plants like Dioncea^ are not unique, but occur under similar circumstances even in ordinary plants, and are characteristic of all plant organs. I shall be able to show, moreover, that an explanation of these phenomena, much I [ l6 COMPARATIVE ELECTRO-PHYSIOLOGY f^ simpler than the theory of electro-motive molecules, is available. It will also be proved that the electrical response due to true excitation is quite distinct from that which is brought about by the hydrostatic disturbance, its sign being in fact opposite. This true excitatory electrical response, again, will be shown to be modified by all those conditions I which affect the physiological state of the tissue. And, lastly, it will be proved that there is no breach of continuity as between the electrical responses in plant and animal, for not only is the sign of response in both cases the same, but it is also true that every type of response and modification of response, which occurs in the animal tissue, is to be found under parallel circumstances in that of the plant also. In order to determine what is the electrical response characteristic of excitation, \we first select for experiment a sensitive plant, say Mimosa, because here, in the responsive fall of the leaf, we have a visible indication of the excitatory reaction. It is desirable at this point to say a few words re- garding the use of the terms ' excitatory ' and ' true excitation.' We are all familiar with the fact that, when muscle is excited by stimulus, it responds in a conspicuous manner by con- traction. This is universally accepted as the phenomenon of ) excitation, its electrical concomitant being galvanometric negativity, Having once applied the term * excitatory ' to this particular aspect of molecular response in living tissues, it is"*^ ; of course important that we should henceforth distinguish '^S carefully between it and its possible opposite, namely, ex- ^< pansion, with concomitant galvanometric positivity. Under '^ stimulation there is a contractioh of, and expulsion of water ^ from, the excited pulvinus, which brings about the depression | of the leaf. It is generally supposed that only the lower half ^ of the pulvinus is excitable. This, however, is an error, for both upper and lower halves are excitable, and contract under stimulation. If localised stimulus be applied on the upper side, that side contracts, and, by the concavity thus induced, the leaf is erected. But, though both halves-* are THE ELECTRO-MOTIVE RESPONSE OF PLANTS 17 sensitive, yet the excitability of the lower is, generally speaking, greater, and diffuse stimulation therefore causes greater contraction of that half Hence the resultant fall is due to the differential contraction of the two sides of the I organ. The excitatory reaction of the organ, then, consists of a contraction ; expulsion of water with consequent diminu- FiG. 12. Arrangement for observing Simultaneous Mechanical and Electrical Responses Leaf stimulated by electro-thermic stimulator. Mechanical record obtained by excursion of spot of light reflected from Optic Lever, which falls on the right of drum. Electric record obtained by excursion of galvano- meter spot of light adjusted to fall on the left side of the drum. tion of turgidity, or negative turgidity variation ; and fall of [the leaf. In order next to determine the electrical concomitant of [this reaction, we make suitable electrical connections by lon-polarisable electrodes, with a galvanometer. One of [these is made with the pulvinus, whose excitation is to be observed, and the other with a distant indifferent point. In .this way we can obtain the excitatory effect at the pulvinus, uncomplicated by that at the distal point. A spot of light, reflected from the galvanometer mirror, is thrown on the > :; 1 8 COMPARATIVE ELECTRO-PHYSIOLOGY recording drum. This spot of light, hitherto quiescent, shows, by its sudden deflection, the occurrence of the excitatory change. To show the concomitance of the mechanical and elec- trical responses, and in order to detect with certainty the exact moment of the initiation of the former, a magnifying arrangement is obtained by attaching the end of the leaf to an Optical Lever. The pull exerted by the falling leaf rotates the fulcrum-rod, carrying a light mirror. The spot of light from this mirror moves in a vertical direction, and that of the galvanometer horizontally or laterally. For the purpose of simultaneous record, it is necessary to have the two in one direction. The up and down movement of the spot from the Optic Mirror is therefore converted into lateral, by means of reflection from a second mirror, suitably inclined. Stimulation may be effected in the neighbourhood of the pulvinus by means of the electro-thermic stimulator, which has the advantage of producing no mechanical dis- ^ turbance. This consists of a V-shaped platinum wire, % suddenly heated by the momentary passage of an electric ^; current. On applying stimulus in this manner it is found >x that the two responses — mechanical and electrical — take ^ place at the same rnoment, the mechanical fall of the leaf JN being practically coincident with the induced electrical >Jn variation. As regards the sign of this electrical change, the ^ excited point is found to bec ome galvanometrically negative : that is to say, the electro-motive variation ihduced~Th the excited vegetable, is the same as that observed in animal tissues. I give below a series of these simultaneous records of mechanical and electrical response (fig. 13), obtained from Biophytum^ whose lateral leaflets give motile indications. It will be seen that the responsive fall of the leaflet, and the subsequent recovery, are synchronous with the electrical variation of galvanometric negativity, and its subsequent recovery. For convenience of inspection. I shall always, unless specially stated to the contrary, represent the normal THE ELECTRO-MOTIVE RESPONSE OF PLANTS 19 responses of mechanical depression and galvanometric nega- tivity by up-curves. The erectile movement and galvano- metric positivity will, conversely, be represented as down. A few words may be said here as regards the syn- chronism between the two forms of response. The excitatory molecular change takes place instantaneously, and the electro-motive variation is, as far as can be judged, strictly concomitant with it. This is shown by the rheotomic method of observation, described in Chapter IV. It will there be seen that a very considerable electro-motive change has already been induced, in a period so short as 'Oi of a second after the reception by the tissue of the stimulating shock. Fig. 13. Simultaneous Mechanical (m) and Electrical (e) Responses in Biophyiuin These responses are seen to take place at the same moment. In the electrical response, then, of highly excitable tissues there is practically no latent period. But if the same elec- trical variation be recorded by the galvanometer, there will be a lag in the response, owing to the inertia of the galva- nometer needle. Similarly, in the mechanical response, though the excitatory reaction is immediate, yet the motile response is delayed, by the antagonistic actions of the upper and lower halves of the pulvinus, the sluggishness of the tissue, and the mechanical inertia of the indicating leaf. The latent period of the mechanical response of a vigorous Mimosa, owing to all these causes, I find to be about twenty-four-hundredths of a second. But this may be still further prolonged when the tissue is in a state of depressed 20 COMPARATIVE ELECTRO-PHYSIOLOGY excitability. It will thus be seen that even when the funda- mental excitatory reaction is instantaneous, its outward expression, whether mechanical or electrical, may nevertheless appear to be subject to delay in consequence of the inertia of the particular indicator concerned. As regards these two forms of response, it should further be remembered that the mechanical and electrical responses are independent indications of the fundamental excitatory reaction, and that neither is dependent for its occurrence on the other. Thus, when the mechanical response is physically restrained, the electrical response takes place unimpeded. I shall here relate an experiment in illustration of this point. A leaf of Mimosa re- sponds to strong stimulus by complete collapse, and the recovery from this state is somewhat prolonged, taking from five to eighteen , ^ ^, minutes, according: to the Fig. 14. Photographic record of Elec- trical Response by Galvanometric season. In order tO obtain Negativity of I'ulvinus of Mimosa, qeries of resnonses with when leaf is physically restrained from ^ ^^^^^^ °' responses Wim falling. The first series in response to these recoveries, within a given uniform stimuli; the second t_i i.- t /^ j ^1 j. series to stimuli twice as strong. reasonable time, I find that it is necessary to apply moderate stimulus. There is then a moderate fall, without complete collapse, and recovery under such circumstances is found to take place within a minute or so. In order to show that electrical response takes place, even when the leaf is prevented from giving mechanical expression, I held the petiole in a clamp, and obtained the set of electrical responses seen in fig. 14. The first series of this record were taken in answer to uniform stimuli of a given intensity, and the * THE ELECTRO-MOTIVE RESPONSE OF PLANTS 21 second to stimuli twice as strong. We may here see how response is increased by increased intensity of stimulus. One peculiarity to be noticed in this figure is the trend of the base-line downwards, showing the increasing positivity of the pulvinus. In order to obtain a photographic record the experiment had to be carried out in a dark room, and under these circumstances the pulvinus undergoes an increase, or positive variation, of turgidity. And we shall see later that a positive turgidity variation is associated with galvano- metric positivity in the same way as the negative turgidity variation is found to be accompanied by galvanometric negativity. In consequence of the impression produced by the con- spicuous movements of the leaf of Mzmosa,it was assumed that only plants showing such movements were to be regarded as excitable. I have already shown elsewhere, however, that this test of lateral motile responses, as a sign of sensitiveness, is fallacious in the extreme. Such mechanical display is possible only when the two halves of an organ are unequally contractile, and there is consequently a greater expulsion of water from one. in response to stimulus, than from the other. If these conditions are not fulfilled, even the so-called * sensi- tive' Mimosa would appear to be insensitive. Thus, when we place a cut branch of Miviosa in water, the pulvini of the leaves, on account of vigorous suction at the cut end, are rendered over-turgid, and the leaves become highly erected. On now applying stimulation, no responsive fall is found to take place ; this is due to the difificulty encountered in the expulsion of water from the gorged tissue. An intact plant, again, which in the light has been found highly sensitive, will often be found insensitive after a short time spent in a dark room. It will then be difficult to believe that the plant is of the sensitive class, for the hardest blow will fail to evoke any mechanical response. And not only does the Mimosa cease to show responsive movement under these cir- cumstances, which may perhaps be regarded as exceptional ; byt under perfectly norrn^l condition? also, its sensitiveness 22 COMPARATIVE ELECTRO-PHYSIOLOGY varies so much that its motile response would seem at times almost to have disappeared. I have already pointed out that it is by the unequal excitabilities of the upper and lower halves of the pulvinus that that differential contrac- tion is induced which brings about the lateral response of the Mimosa leaf Now it is clear from this that if the differential excitability should be reduced or abolished, by any means whatsoever, there will then be a corresponding diminution or abolition of response. We shall see later that the excitability of a tissue depends upon its state of turgor, and in Mimosa, from internal causes, a periodic variation is induced in the relative turgescence of the two halves of the pulvinus. We might then expect, in consequence of this, to find a periodic variation of motile sensibility. And certainly, whatever may be the cause, a long course of observation will convince the inquirer of the occurrence of great varia- tions in the sensibility of Mimosa at different times of the day. Thus, I had six specimens of this plant growing in pots in the open, and I found, watching them in the month of August, that at eight o'clock in the morning the pulvini of the leaves of all these plants were sensitive in the highest degree. Half an hour later, however, this sensitiveness had so far waned that they would give hardly any motile indi- cation. It Ts, perhaps, worth while to remark, in connection with this, that a constant observer is able to judge, by a \ ipeculiar, though indescribable, attitude of the leaves, whether \or not this condition of insensitiveness has supervened. Thus the mechanical movements of the belauded sensitive plants, such as Mimosa, on which depended the arbitrary assumption that 'ordinary' plants were insensitive, rest on a basis which is itself extremely unreliable. For by this standard one identical plant ought to be classed as belonging to both the sensitive and insensitive groups, according to the time of day at which any particular observation is made. The fact that when the mechanical response of the leaf of Mimosa is physically restrained, excitatory electrical response takes place unimpeded, shows that we have a criterion by I THE ELECTRO-MOTIVE RESPONSE OF PLANTS 23 which to test the excitability of a plant, independently of any motile indication. On applying this test, I have found that not the so-called sensitive plants alone, but all plants and all organs of all plants, respond to stimulation. And from this I was led to the discovery that ordinary plants also, in spite of current misconceptions, exhibit motile response by mechanical contraction. The common error of regarding these plants as insensitive has arisen from the fact that in a radial organ, diffuse stimulation induces equal contractions on all sides. Hence those lateral movements, dependent on differential contraction, which are seen so conspicuously dis- played in Mimosa, cannot take place here. But that the organ as a whole undergoes a responsive contraction has been demonstrated by recording the consequent induced shortening of its length. Such longitudinal contraction is sometimes very considerable ; for instance, in the filamentous corona of Passiflora it may sometimes be as much as 20 per cent, of the original length. Having thus shown that all plants are excitable, I shall proceed to demonstrate the fact by means of electrical response. In studying the excitatory effect on ordinary plants, we must bear in mind that there are two different ways of stimulating a given point : that is to say, locally or directly, and by transmission of excitation from a distance. (Conducting tissues are capable of stimulation in either of these ways, but the feebly conducting must be subjected to local excitation, since the effect of stimulus applied at a- I distance cannot, in their case, reach the responding point. Organs containing fibro-vascular elements are fairly good conductors, and stimulus applied on them at one or two centimetres from the point to be stimulated will thus easil]^ reach it. It must, however, be remembered that stimulus becomes enfeebled by transmission through a long tract, its effect at a great distance being negligible. Pa renchymatous ^ '[ ^ tissues are bad conductors of excitation, and in order to ^'^*^ excite them, stimulus must therefore be applied directly. Turning first to the transmitted mode of stimulation. 24 COMPARATIVE ELECTRO-rilYSIOLOGY we take a petiole or stem, and making suitable electrical connections (fig. 15), apply stimulus, say by contact of hot r wire at the point marked x . After a short interval, necessary for the excitation to traverse the intervening distance, an . electrical response is obtained, of galvanometric negativity.^ ^ * It is thus seen that the electrical response of ' ordinary' is of V^ the same sign as that of ' sensitive ' plants, and that in both, ^ J again, it is like that of animal tissues. K^ I shall next proceed to demonstrate a very important ' proposition : namely, that all effective forms of stimulation induce an identical excitatory response of galvanometric negativity. Any sudden change of environmental conditions may constitute an efficient stimulus. Such are : sudden rise Fig. 15. Method of TransmiUed Stimulation Stimulus applied to the right at x . Excitation reaches right contact first, causing galvanometric negativity of the point. of temperature ; any variation of pressure, whether of tension or compression ; mechanical blows or torsional vibration ; any prick or cut ; the application of a chemical agent, such as acid; the application of light ; the incidence or variation of electrical currents ; and, lastly, the action of gravity. The stimulatory action of all these agents has already been demonstrated in my work on * Plant Response,' ^ the excita- tion induced being there shown to find expression in appropriate mechanical movements. In the present volume I shall deal more particularly with the electrical reactions which they induce. The effects of the stimulating action of ' Bose, Plant Response as a Means of Physiological Investigation^ 1906. THE ELECTRO-MOTIVE RESPONSE OF PLANTS 25 electrical currents, of light and of gravity, will be taken up in special chapters devoted to their consideration, while here I shall demonstrate the exci- tatory effects of the other forms of stimulus enumerated. We have already observed the responsive effect which results from the sudden application of heat, by means of a hot wire. The effects of various forms of mechanical stimulation may now be subjected to demonstration, and first we have to observe the effect of the stimulus resulting from sudden tension. The specimen is clamped securely in the middle (fig. 16), so that when a vertical pull is given to the upper half, that half alone is subjected to a sud- denly increased tension, the lower being left entirely unaffected. Under these circumstances, there is an electrical response, A becoming^ galvanometrically negative. A is next subjected to mechanical compressron7and for this purpose the piece of moistened cloth surrounding the specimen, and making the electrical connection at A, is placed between the two grooved halves of a cork. The enclosed plant tissue at A may now be made to undergo sudden compression, by squeezing the pieces of cork together. This gives rise to the same electrical response as before. This fact, that both tension a nd compression will give rise to similar excitatory responses of galvanometric negativity, may receive independent 'dernonstration by first making an electric connection at A with the upper side of the speci- men (fig. 17). When the tissue at A is now suddenly bent down, this upper side becomes convex : that is to say, it is subjected to tension, This gives rise to the excitatory Fig. 16. Excitation by Sudden Tension Plant securely clamped. When suddenly pulled, tension in- duces galvanometric nega- tivity of A. 26 COMPARATIVE ELECTRO-PHYSIOLOGY Fig. 17. Excitatory Response to Tension and Compression When E is connected with the upper point A a sudden bending down causes tension of A. When connection is made with a' the same flexure causes compression of a'. Both induce galvanometric negativity. response of negativity. The electrical connection at A is[ next removed to the lower side of the specimen, at a point A'. On now repeating the sudden flexure, a' undergoes com- pression instead of ten- sion. The result is a similar negative elec- trical response. Excitation may, again, be produced by means of a sudden blow at a point. This blow may be delivered by means of a spring-tapper (fig. 18), in which s is the spring proper and the attached rod R carries at its end the tapping head T. A projecting rod — the lifter L — passes through S R. It is provided with a screw-thread, by means of which its length, projecting downwards, is regulated. By means of this the height or intensity of the stroke may be varied. As one of the spokes of the cog-wheel C is rotated past L, the spring is lifted and released, and T delivers a sharp tap. The height of the lift, and therefore the intensity of the stroke, is measured by a graduated scale not shown in the figure. We can increase the intensity of this stroke through a wide range, first, by augmenting the projecting length of the spring by a sliding catch. We may give isolated single taps, or superpose a series in rapid succession according as the whjeel is rotated slowly or quickly. Fig. 18. The Mechanical Tapper THE ELECTRO-MOTIVE RESPONSE OF PLANTS V (a) (i) I Stimulation, again, may be effected by the prick of a needle or pin in the neighbourhood of A. Response to this also occurs by the normal galvanometric negativity. Successive pricks may thus give rise to successive responses. Or the specimen may be subjected to torsional vibration. It is here held in the middle by a clamp, and stimulus of torsional vibration is applied at one end. The stimulation of A makes that end gal- vanometrically negative, the direction of the current outside the circuit being towards, and in the tissue away from, A. Vibration of B induces responsive nega- tivity of B (fig. 19), the current of response being now reversed. In the cases just described, it will be noticed that stimulus is ap- plied directly. This method is, therefore, specially applicable when we wish to study the excitability of such tissues as are not good conductors of excitation, the method of transmitted stimulation being here, therefore, inapplicable. In order to observe the effect of chemical stimulation, the given agent — sulphuric or hydrochloric acid — is applied at X at a short distance from the proximal contact. The trans- mitted excitation is now again demonstrated by the induced galvanometric negativity of that contact. It will thus be seen that, whatever be the effective form of stimulus employed, / ^ ^ it gives rise to a definite and invariable electrical response Uft^ whose sign is always one of galvanometric negativity. t^tMJ h^ It was shown, then, in the course of this chapter that the excitatory change in ' sensitive ' plants is characterised by contraction, negative turgidity variation, mechanical depression of the leaf, and by the electrical response of Ctirrent of response when A is stimulated-^ Current of -response when B is stimulated-* Fig. 19. The Torsional Vibrator (a) The plant is clamped at c, between A and B. {b) Responses obtained by alternately stimulating the two ends. Stimula- tion of A produces upward response ; of B gives downward response. 28 COMPARATIVE ELECTRO-PHYSIOLOGY galvanometric negativity, all these effects being concomitant. It was further shown that electrical response is independent of the mechanical, being unimpeded in its occurrence when the leaf is physically restrained. The same electrical response of galvanometric negativity is also obtained from the tissues of the so called * ordinary ' plants. And these electrical responses of plant tissues, it was further noted, are identical in sign with the corresponding responses given by animal tissues. All forms of stimulus, moreover— mechanical, thermal, photic, chemical, and electrical — induce the same excitatory response of galvanometric negativity. L0 ^u^ CHAPTER III THE APPLICATION OF QUANTITATIVE STIMULUS AND RELATION BETWEEN STIMULUS AND RESPONSE Conditions of obtaining uniform response — Torsional vibration as a form of stimulus — Method of block — Effective intensity of stimulus dependent on period of vibration — Additive action of feeble stimuli — Response recorder — Uniform electric responses— List of suitable specimens — Effect of season on excitability — Stimulation by thermal shocks — Thermal stimulator — Second method of confining excitation to one contact — Increasing response to increas- ing stimulus— Effect of fatigue— Tetanus. A QUALITATIVE demonstration has been given in the last chapter of the induction of galvanometric negativity in plant tissues, in response to the excitation caused by various forms of stimulus. This galvanometric response is thus a sign or indication of the state of excitation ; and under normal con- ditions it will be of uniform extent, provided only that the stimuli are also uniform. Assuming this ideal condition to be secured, it is clear that the physiological modifications induced by various agents will be manifested by a corre- sponding modification of response. The conditions essential to such application of stimulus are, then, (i) that it should be capable of uniform repetition ; (2) that it should be capable of increase or decrease by definite amounts ; and (3) that it should be of such a nature as to cause no injury, by which the excitability of the tissue might be changed in some unknown degree. These conditions, on which the success of the electro-physiological investigation depends, are very difficult to meet. Chemical stimulation, for example, cannot be uniformly repeated. Electrical stimulation, again, which has the advantage of being easy to render quantitative, is open to the objection that by escape of current it may induce 30 COMPARATIVE ELECTRO-PHYSIOLOGY galvanometric disturbance. Indeed, as the response is electrical, it is obvious that if we are to obtain unimpugnable results, a non-electrical form of stimulus is almost a necessity. But it is only after providing against various sources of error that the electrical form of stimulation can be used with con- fidence. The stimulation caused by mechanical blows can be repeated, it is true, with uniform intensity. But the point struck is subjected to increasing injury, and its excitability thus undergoes an unknown variation. I have, however, been able to devise two different modes of stimulation, in which all these difficulties have been Fig. 20. The Vibratory Stimulator Plant P is securely held by a vice v. The two ends are clamped by holders C c'. By means of handles H h', torsional vibration may be imparted to either the end A or end b of the plant. The end view (d) shows how the amplitude of vibration is predetermined by means of movable stops, s s'. successfully overcome, rendering the results as perfect as possible. These are (i) torsional vibration, and (2) the application of thermal shocks. For the obtaining of perfect responses, it must be said here that there is still another condition to be fulfilled. If we wish to obtain the pure effect of stimulus at one contact, say A, special care must be taken that excitation does not reach the second contact, B ; for otherwise, unknown effects of interference will occur. This may, it is true, be obviated by means of the method of relative depression or method of negative variation, so called, to be described in a subsequent chapter. But the experi- mental mode which I am about to describe, in which a block THE APPLICATION OF QUANTITATIVE STIMULUS 3 1 is interposed between A and B, is much more perfect. According to this arrangement, the specimen is tightly clamped in the middle, by which device the excitation of either end is practically precluded from affecting the other. Stimulation is brought about by means of torsional vibration. The stem or petiole is fixed, at its middle, in a vice, V, the free ends being held in tubes, c c', each provided Fig. 21 Complete Apparatus for Method of Block and Vibratory Stimulation Amplitude of vibration which determines the intensity of stimulus is measured by the graduated circle seen to the right. Temperature is regulated by the electric heating coil R. For experiments on action of anoeslhetics, vapour of chloroform is blown in through the side tube. with three clamping jaws. A torsional vibration may now be imparted to the specimen at either end by means of the handles H and H' (fig. 20). The amplitude of vibration which determines the intensity of stimulus can be accurately measured by the graduated circle, and may be predeter- mined by means of the sliding stops S S/ The complete vibrational apparatus, by means of which various experi- mental investigations may be carried out, is given in fig. 21. 32 COMPARATIVE ELECTRO-PHYSIOLOGY Moistened cotton threads in connection with the non-polari- sable electrodes, E E, make secure electrical contacts with A and B. For experimenting on the effects of temperature, there is an electrical heating coil, R, inside the chamber. For the study of the effects of different gases, there are inlet and outlet tubes, which enable a stream of the required gas or vapour to be circulated through the chamber. If the A end of the specimen be now suddenly torsioned through a given number of degrees, a responsive electro- motive variation takes place, which after- wards subsides gradually. If next the torsioned end be suddenly brought back to the original position, a second electro- l I \ 1 motive response is obtained, similar to the V \ \ \ first. Hence, in the case of a to-and-fro CO h c d vibration, the responsive effects are addi- *" tive, and we have the further advantage Fig 22 Induence of ^hat the tissue at the end of the operation is Suddenness on the ^ Efficiency of Stimu- returned to its original physical condition. ^"^ In order that successive stimuli may are responses' to be equally effective, another factor besides vibrations of the t^g constancy of the amplitude of vibra- same amphtude, •' ^ 30°. In a the vi- tion has to be considered. It is to be bration was very ^ -^^ ^j^^ ^j^ ^ effectiveness of the slow i in <3 it was less slow ; it was stimulus in evoking response depends also rapid in 5.^" ^^^^ ^^ ^^e rapidity of the onset of the dis- turbance. In the application of vibratory stimulation to plants, I find the extent of response to depend to some degree on the quickness with which the vibration is effected. I give below records of responses to successive stimuli, induced by vibration through the same amplitude, which were delivered with increasing rapidity (fig. 22). It will be noticed that an increasing quickness of vibration increased the response, but that this reached a limit. If we wish, then, to maintain the effective intensity of stimulus constant, we must meet two conditions. First, the amplitude of vibration must be kept the same. This is done by means THE APPLICATION OF QUANTITATIVE STIMULUS 33 of the graduated circle and movable stops : and, second, the vibration period must be uniform. This last condition is effected by an arrangement shown in fig. 23. The torsion- head is kept tense by means of a stretched spiral spring, s, made of steel. From this torsion-head there projects an elastic brass piece, B. R is a striker which can be made to give a quick stroke to B, by the rotation of the handle. A quick to-and-fro vibration is thus produced, by the blow given to B, acting against the tension of the antagonistic spring S. The amplitude of the angular vibration is at the I Fig. 23. Spring Attachment for obtaining Vibration of Uniform Rapidity same time predetermined by means of the stops P and Q. The arrangements described are as used in ordinary work. But for certain experiments on differential excitability, a second striker, R', may be attached to the other end of the apparatus, and by this means the opposite contacts in con- nection with E and E' may be excited simultaneously. In order to obtain responses of great amplitude, it is now necessary to increase the amplitude of vibration. But this may give rise to fatigue. By way of avoiding this, therefore, it is still possible to obtain enlarged response by the additive effect of repeated feeble stimuli. In the electrical response of plants a sub-minimal ^tjmulvis, singly ineffective, is found 34 COMPARATIVE ELECTRO-PHYSIOLOGY to become effective by the summation of several. This is seen in fig. 24, where a single vibrational stimulus of 3°, alone ineffective, was found to evoke a large response when repeated with rapidity thirty times in succession. For the delivering of such equal and rapidly succeeding stimuli, I substitute for the single striker R an eight-spoked wheel, a complete rotation of which, by means of the handle, gives rise to a definite sum- mated effect : and a series of responses to such summated stimulations is found to be uniform. The galvanometer used for these experiments is a dead-beat instru- ment of D'Arsonval type. The sensitive- ness of this is such that a current of io~^ ampere causes a deflection of i mm. at a distance of i metre. For a quick and accurate method of obtaining records, I devised the following form of response-recorder. The curves are obtained directly, by tracing the excursion of the Fig. 24. Additive Effect {a) A single stimulus of 3° vibration pro- duced little or no effect, but the same stimulus when rapidly superposed thirty times pro- duced the large effect (d). (Leaf stalk of turnip.) ¥\G. 25. Response Recorder galvanometer spot of light on a revolving drum (fig. 25). This drum, on which is wrapped the paper for receiving the record, is driven by clockwork, Different speeds of THE APPLICATION OF QUANTITATIVE STIMULUS 35 revolution can be given to it by adjustment of the clock- governor, or by changing the size of the driving-wheel. The galvanometer spot is thrown down on the drum by the inclined mirror M. The galvanometer deflection takes place at right angles to the motion of the paper ; a stylographic pen attached to a carrier rests on the writing surface. The carrier slides over a rod parallel to the drum. As has been said before, the galvanometer deflection takes place parallel to the axis of the drum, and as long as the plant rests un- stimulated, the pen, remaining coincident with the stationary I galvanometer spot on the revolving paper, describes a straight line. If, on stimulation, we trace the resulting excursion of the spot of light, by moving the carrier which holds the pen, the rising portion of the response curve will be obtained. The galvanometer spot will then return more or less gradually to its original position, and that part of the curve which is traced during this process constitutes the recovery. The ordinate in these curves represents the electro-motive variation, and the abscissa the time. We can calibrate the value of the deflection by applying a small known E.M.F., say of 'i volt, to the circuit, and noting the deflection which results. This gives us the value of the ordinate. The value of the abscissa which represents time is determined by the distance through which the recording surface moves, in unit time. In this simple manner accurate records are obtained. It has the additional advantage of enabling the observer to see at once whether the specimen is suitable for the purpose of investigation. A large number of records might be taken by this means, in a comparatively short time. It is also easy to take the records photographically by wrapping a photographic film round the recording drum. I give in fig. 26 a series of responses taken from the root of radish {Raphanus sativus), in which the stimuli were applied at intervals of one minute. This shows how ex- tremely uniform the responses may be rendered, if proper I precautions are taken. It may here be onge more pointed P3 36 COiMPARATIVE ELECTRO-PHYSIOLOGY out, that for convenience of inspection, the records in this book have been so taken that the normal electrical responses of galvanometric negativity, unless specially stated to the contrary, are seen as up-curves, galvanometric positivity being represented by down-curves. These excitatory responses of Fig. 26. Photographic Record of Uniform Responses (Radish) galvanometric negativity are obtained with all plants, and with every organ of the plant. I give here a table containing a list of specimens which will be found on stimulation to give fairly large electro-motive effects, occasionally as high as •I volt. Organ Specimen Root Carrot {Daucus carota) Radish ( Raphanus sativus) Stem Geranium {Pelargonium) Vine ( Vitis vinifera) Amaranth (Amaranthus) Petiole . Horse Chestnut {Aiscuhis hippocastantnn) Turnip (Brassica napus) Cauliflower (Brassica oleracea) Celery {Apium graveolens) Eucharis lily [Eucharis amazonica) Arum lily {Ricardia africana) Peduncle. Fruit . . Egg-plant [Solanum melongena) THE APPLICATION OF QUANTITATIVE STIMULUS 3/ These responses, being physiological, vary in intensity with the condition of the specimen. The same plant which gives strong electrical response in spring or summer, may exhibit but feeble responsiveness in autumn or winter. Again, we shall see in a subsequent chapter that any agent which depresses physiological activity will also depress the electri- cal response ; and, lastly, when the specimen is killed, the normal response is abolished. I shall next describe a second and equally perfect method of stimulation, by means, namely, of thermal shocks. We have seen that a sudden thermal variation acts as an efficient stimulus. I have also shown in my * Plant Response ' that thermal radiation acts as a stimulating agent, in inducing excitatory contraction. Hence, if a tissue be surrounded by a platinum wire, through which an electrical heating-current can be sent, the enclosed tissue will be subjected to a sudden variation of temperature, and also to the thermal radiation proceeding from the heated wire. Now if in successive experiments the duration and intensity of the current flowing through the wire be maintained constant, the stimuli also will thereby be rendered constant. The thermal stimulator, as alread)?- said, surrounds the specimen, but is not in actual contact with it. This is to prevent any injury to the tissue, by scorching. The current is so adjusted as to make the platinum wire red-hot and this heating-current is closed for about half a second at a time. Should larger response be desired, it can be obtained by the summated effect of a number of such shocks, or the thermal stimulator may be put in direct contact with the tissue, if care be taken that the rise of temperature is not so great as to injure it. The difficulty of ensuring similarity of duration to each individual shock is overcome by the use of a balanced key actuated by a metronome (fig. 27). A second rod is attached at right angles to the vibrating rod of the metronome, and carries a bent piece of brass in the form of two prongs. During the course of each vibration these prongs dip into 38 COMPARATIVE ELECTRO-PHYSIOLOGY Fig. 27. Stimulation by Thermal Shocks two cups of mercury, thus closing the electrical circuit for a brief and definite time. When a second press-key, not shown in the figure, is open, the circuit is incomplete, and there is no thermal stimulation. The observer then presses this key, and counts, say, five strokes of the metronome, after which the press-key is again opened. In this way, the sum- mated effect is ob- tained, of five equal thermal shocks. This process is repeated as often as desired, at intervals of, say, one minute, by which time the tissue is generally found to have completely recovered from its ex- citatory electrical variation. In the case of the experimental arrangements of which the diagram is given in fig. 27, stimulation is confined to one contact of the responding circuit. The method by which excitation was here prevented from reaching the distal contact is important. I shall, in the course of the present work, show that the parenchymatous tissue of the lamina of a leaf or leaflet is a bad conductor of excitation. Hence if the second contact of the circuit be made with this tissue, the stimulus does not reach the distal point. It is true that a certain small proportion might conceivably be conducted through the attenuated fibro-vascular channel of the midrib. But even so remote a contingency is provided against by a transverse cut across the midrib on the hither side of the contact. The arrangements, then, being made in the manner de- scribed, the tissue may be subjected to the action of successive uniform stimuli. How regular the resulting responses may be rendered will be seen from fig. 28, in which is given a series of responses obtained from the petiole of a fern (fig. 28) THE APPLICATION OF QUANTITATIVE STIMULUS 39 under successive thermal shocks, imparted at intervals of one minute. We have hitherto studied the responses caused by uniform stimuli. We shall next observe the increase of responsive effects brought about by increase of stimulus. In ■tUw\^V. (^28. Photographic Record of Uniform Response in Petiole of Fern to transmitted excitation animal tissues it is found, speaking generally, that increasing stimuli induce increasing effects, but that this process has a limit ; and in plant tissues the same is found to be the case. In order to obtain effects of the simplest type, not compli- cated by any secondary phenomena, it is necessary to choose specimens which exhibit little fatigue. In the first of these the stimulus was ap- plied by means of the spring-tapper. The first stimulus was given by a fall of the striking-lever from the height h ; the second from 2h ; and the third from 3h. The response- curves (fig. 29) clearly show the in- crease of effect due to this increasing stimulus. In the second series, the stimulus applied was vibrational, and increased from 2-5° to 12-5° by steps of 2*5° at a time. Fig. 30 shows how the intensity of response tends under these conditions to approach a limit. The following table gives the absolute values of the responsive electro-motive variations. Fig. 29 Taps of increasing strength 1:2:3:4 producing in- creased response in leaf- stalk of turnip. 40 COMPARATIVE ELECTRO-PHYSIOLOGY Table showing the Increased Electro-motive Variation induced BY Increasing Stimulus. Angle of vibration Induced E.M.F. 2-5° •044 volt. 5° •075 >. 7;5° •090 ,, IO° •100 „ 12-5° •106 „ In such normal cases an increase of response is always induced with increasing stimulation. A diminution of response may, however, sometimes appear, with increasing Fig. 30. Increased Response with Increasing Vibrational Stimuli ( Cauliflower-stalk ) Vertical line = •! volt. Stimuli applied at intervals of three minutes. stimulus. But this is merely a secondary effect, due to fatigue. The following records (fig. 31) will show in what manner this may be brought about. They were taken with specimens of the petiole of cauliflower, in one of which (a) fatigue was absent, while in the other (b) it was present. In the first specimen the recovery from each stimulus was THE APPLICATION OF QUANTITATIVE STIMULUS 4I complete. Every response in this series starts, therefore, from a position of equihbrium, and the height of each single response increases with increasing stimulation. In the second case, however, the molecular derangement consequent on stimulation is not completely removed after any single Fig. 31. Responses to Increasing Stimulus obtained with Two Specimens of Stalk of Cauliflower In {a) recovery is complete, in (3) it is incomplete. Stimulus of the series. That the recovery is only partial is [seen in the gradual shifting of the base-line upwards. In the former case the base-line had been horizontal, represent- |ing a condition of complete equilibrium. Now, however, the ^base-line, or line of modified equilibrium, is tilted upwards. :Thus, even here, if we measure the heights of successive 42 COMPARATIVE ELECTRO-PHYSIOLOGY responses from the line of absolute equilibrium, they will be found to increase with increasing stimulus. Ordinarily, how- ever, no allowance is made for the shifting of the base-line, the responses being measured instead from the place of its previous recovery, or point of modified equilibrium. In this way these responses undergo an apparent diminution. I have occasionally observed another curious phenomenon in connection with the subject of response under increasing stimulus. During the gradual increase of the stimulus from a low value, there would at first be no response. But on reaching a certain critical value, a response would suddenly be evoked which was maximum — that is to say, would not be exceeded, even when the stimulus was further increased. We have here a parallel case to what is known in animal physiology as the 'all or none' principle. In the case of cardiac muscle, for example, there is a certain minimal intensity of stimulus which is effective in inducing response. But further increase of stimulation causes no concomitant increase of effect. When a tissue is subjected to rapidly succeeding stimuli, the excitatory effects are superposed upon each other. In muscle, for example, the contractile effect of the second stimulus is added to that of the first, before that has time to disappear. The result is a summation of effects more or less complete ; and these attain a maximum. With moderate frequency of stimulation, such a tetanic effect is incomplete, tending to become more and more complete, with the progressive increase of frequency (fig. 52). I have obtained results in every way similar to these, with the mechanical response of ordinary plants. In fig. 33 is given a photo- graphic record of tetanus, taken from the longitudinal motile responses of the style of Datura alba. Similar tetanic effects are also obtained in the electric response of plants, of which the records seen in fig. 34 form an example. The difificulties in the quantitative observation of electrical response have thus been overcome by the employment of two different methods of stimulation — namely, torsional vibration, and stimulation by thermal shocks. In the case of the THE APPLICATION OF QUANTITATIVE STIMULUS 43 former, the intensity of stimulation was seen to depend on the amplitude of vibration. In the latter, stimulus intensity was determined by that of the thermal variation, which again was regulated by the intensity and duration of the electrical heating-current. It was also seen to be important that the Fig. 32. Genesis of Tetanus in Muscle Record to left shows incomplete tetanus, with moderate frequency of stimulation. Record to right shows tetanus more complete, with greater frequency of stimulation (Brodie). excitation of one contact should be prevented from reaching the other, and this was provided against in two different ways. In the first of these, a physical block was interposed between the two contacts. In the second, the distal contact was made with the non-conducting tissue of a lateral leaf Fig. 33. Photographic Record of Genesis of Tetanus in Mechanical Response of Plants (Style of Datura alba) ^ 1\ .r- Wmk I-— 1 0^ ) \ ^ y J C "*^^.,nr..nnnr.J-^" HI ^^-^-^ e-J^-ff — ^r- And although this hydrostatic ^^ disturbance is transmitted very quickly, yet a certain time is con- sumed in the process of forcing water into the pulvinus, by which to bring about the erection of the leaf After the passage of the hydrostatic wave, there follows Fig. 42. Experimental Arrange- the wave of true excitation, passing ment for obtaining Records from cell to cell, and inducing the on Smoked Drum of Responses ... given to Direct and In- characteristic reaction, of negative 7^w''"'''^^^'°"' ^^ ^'^^ °^ turgidity - variation. And when Thermal stimulator at s produces this excitatory wave reaches the direct stimulation, and conse- pulvinus, the prcvious erectile quent fall of leaf. Moderate * 1 • 1 stimulation, at a distant point, movement should glVe place tO of e?ict?on!'' ^"^ '""^''''^ '^''^ excitatory depression, or fall of the leaf In fig. 42 are seen the arrangements for an experiment on Mimosa by which these inductions may be verified. Moderate thermal stimulus is applied at S, at a certain distance below the indicating leaf This latter is attached by a thread to a writing-lever, which traces the response-record on a smoked revolving drum. When the stimulus is applied at a point S very near the pulvinus, the response takes place by a negative turgidity- variation, with a concomitant fall of the leaf, seen in fig. 43 {a) as an up-curve. When a moderate stimulus is applied POSITIVE AND NEGATIVE TURGIDITY- VARIATIONS 57 at a greater distance S^^ the hydrostatic wave causing the positive turgidity-variation brings about an erectile twitch. This is followed by a responsive fall, when the true excitation reaches the organ (fig. 43, b). It has thus been shown that by separating the responding point from the point stimulated, or the receptive point, it is possible to discriminate two different effects which are both brought about by stimulus. It I^P is most important, moreover, to distinguish between these two factors : namely, the direct effect of stimulus causing contraction, and its indirect effect, causing expansion. We have seen that direct excitation and transmitted excitation both induce contrac- tion, negative turgidity-variation and fall of the leaf Unfor- ^Itunately, in animal physiology, § it has been customary to apply (the term indirect to that form of stimulation which is applied at a distance. And it has not been noticed that such stimulus is capable of inducing diametri- cally opposite results, according I as the true excitatory effect reaches, or does not reach, the responding organ. When the intervening tissue is highly conducting, the transmitted effect induces exactly the same 1 result as if stimulus were applied directly. But we shall see that when the intervening tissue is non-conducting or feebly conducting, true excitation is not transmitted, and the effect which makes its appearance at the responding point is then due to increase of hydrostatic tension, causing positive Fig. 43. Mechanical Responses of Leaf of Mimosa (rt) record of responsive fall when stimulus applied near the re- sponding organ ; {b) response when stimulus is applied on same side, but at greater dis- tance, s^^. A preliminary erectile response is here followed by the true excitatory depression. This is due to the indirect effect first transmitted being succeeded by the direct. Had the stimulus applied been feebler, or more distant, there would have been only the first, or indirect erectile effect. 58 COMPARATIVE ELECTRO-PHYSIOLOGY turgidity-variation, with the concomitant effect of expansion, and, in the case of Mimosa^ of erection of the leaf This latter effect of positive turgidity-variation and expansion, I shall therefore distinguish as the Indirect Effect of stimulus, in contradistinction to the term Indirect Stimu- lation, as it is generally used. The last-named, however, I shall myself always refer to under the title of TRANS- MITTED Stimulation. If the intervening tissue be of Fig. 44. Mechanical Response of Biophytum to Thermal Stimulation Stimulus was applied at some distance from the responding leaflet. And the preliminary erectile twitch is due to the prior arrival of the hydrostatic disturbance. Thick dot represents moment of application of stimulus. moderate conducting power, we shall, as in the case of the experiments on Mimosa^ obtain a preliminary erectile twitch, due to the indirect effect of stimulus, followed by a fall, in consequence of the transmission of the true excitatory effect. In fig. 44 is seen this twofold expression of the indirect and transmitted effects of stimulus, given by the leaflet of Biophytum. These two waves, then, of increased hydrostatic tension I POSITIVE AND NEGATIVE TURGIDITY-VARIATIONS 59 and of true excitation, induce, as we have now seen, opposite responsive reactions. But of these, that due to true excita- tion is, generally speaking, greatly predominant. Hence, when these two waves reach the responding organ in close succession, as is the case when the point of stimulation is very near, the excitatory effect masks the hydrostatic. In order, then, to separate them, we may employ various methods. First, in the case of highly conducting tissues, the stimulus must be applied at a sufficient distance to make the slow excitatory wave lag adequately behind the quickly travelling h5'drostatic wave. Or we may choose a direction of transmission of excitation which will be relatively slow. Thus I have found that transmission across a stem, for example, is very much slower than along its length. Hence, on applying moderate stimulus at S, (fig. 42) at a point on the stem diametrically opposite the pulvinus, of the given leaf, it is found that the excitation reaches the pulvinus only after a measurable interval, the hydrostatic effect inducing erectile response much earlier. Thus in a given experiment, whose record was taken on a fast-moving drum (fig. 45), the erectile response took place "6 second after the application of stimu- lus, whereas the true excitatory fall did not occur till 3 '45 seconds had elapsed — that is to say, 285 seconds later. It is to be borne in mind that a certain interval of time passes, even after the arrival of the respective waves, before the tur- gidity-variation is able to give rise to the motile indication. Let us next examine the results at the responding tissue of the indirect effect of stimulus. The distant receptive point contracts on stimulation, and sends to the responding organ a wave of increased hydrostatic tension. This, as we have seen, forces water in, and expands the tissue. Work is thus done on the tissue which increases its store of energy. In this the indirect is unlike the typical direct effect of stimulus. For the latter causes the impulsive fall of the leaf, which represents work done by the tissue, and an expenditure of energy. We must, therefore, recognise two distinct respon- sive effects, according as the work done is positive — done on 6o COMPARATIVE ELECTRO-PHYSIOLOGY the tissue — or negative : that is to say, done by the tissue. The outward manifestations of these two processes are respectively expansion and contraction. The positive, as we shall see, is not the result of hydrostatic disturbance as such, but is the effect of energy transmitted hydraulically. The indirect effect of stimulus, then, gives rise to positive turgid ity-variation, and increases the internal or potential Fig. 45. Record of Response of Mimosa Leaf, taken on a fast- moving drum Stimulus applied at moment a, on point of stem diametrically opposite to responding leaf. Hydro-positive erectile response occurs at b, '6 second after application of stimulus. True excitatory response of fall takes place at r, 3*45 seconds after application of stimulus. Time-marks represent fifths of a second. energy of the organ. A positive turgidity-variation is thus concomitant with an increase of internal energy and a negative turgidity-variation with the reverse. In observing the mechanical response, we saw that the expression of positive turgidity-variation, due to the indirect effect of stimulus, consisted of an erection, and was therefore of opposite sign to that of the negative turgidity-variation, caused by true excitation, and expressing itself in a fall, of POSITIVE AND NEGATIVE TURGIDITY-VARIATIONS 6 1 the leaf. We shall now see whether a similar difference exists between the electrical expressions of the positive and negative turgidity-variations. In carrying out this experiment, I took a specimen of Biophytum and applied stimulus at a distance from the par- ticular leaflet whose responses were to be observed, arranging, at the same time, for a simultaneous record of the mechanical and electrical responses. It will be seen from fig. 46 that the preliminary erectile twitch, due to the positive turgidity- variation, has, as its concomitant, galvanometric positivity. And this is followed in both records by its opposite : namely, the contractile fall and the galvanometric negativity of true excitation. It will thus be seen that the increase of internal energy, with its positive turgidity-variation, has, as its electrical expression, galvanometric positivity. Besides this, the mere physical movement of water in the tissue gives rise to a certain electrical varia- tion of positivity, and this can still be detected, even after the tissue is killed. The question of how to discriminate what proportion of the electro- positivity was due to this mere water - movement, and what to the increase of turgidity, associated with the increase of in- ternal energy, I at first found it very difficult to decide. But I ultimately succeeded in doing this by bringing a plant to a condition just short of death, and thus abolishing its true excitatory reaction. In this condition, the responsive Fig. 46. The Abnormal Positive preceding the Normal Negative in Mechanical and Electrical Responses in Biophytum X represents the moment of application of stimulus. The upper is the mechanical and the lower the electrical record. The records downward indicate erection of the leaf or galvanometric positivity. 62 COMPARATIVE ELECTRO-PHYSIOLOGY indication was found to be one of considerable electro- positivity. On finally killing the plant, however, the positive change due to water-movement was found to represent so insignificant a proportion of the whole as to be negligible. In order to exhibit the electrical expression of the excitatory and hydro-positive effects of stimulation, in ordinary plants, I took a petiole of cauliflower and made one connection, the proximal, with a point on it, and the other with an indifferent point on the surface of the lamina. In order to obtain the unmistakable hydro- static effect, the petiole was sud- denly squeezed, at a distance of 6 cm. from the proximal contact, and this gave rise, as will be seen (fig. 47, a), to a positive response, represented downwards. This was repeated once more, and the same effect observed. I next applied thermal stimulus at a dis- tance of 4 cm. from the respond- ing point. In this case hydro- static and excitatory disturbances reached the contact, the hydro- ^'^itcuicai'lTspT^^^^^^ shortly followed by the of Cauliflower excitatory, as in the case of the fl, hydro-positive ; b^ di-phasic ; f, excitatory negative responses. experiment on Mimosa, This is seen in the record, as a di- phasic response, the hydrostatic positive being followed by the excitatory negative (fig. 47, b). Reference has already been made to the observation of Burdon Sanderson, that in the lamina of the Dioncea leaf the immediate response was one of galvanometric positivity. Mis- taking this for the true excitatory effect, he concluded that the response of the plant was of opposite sign to that of the t^OSlTlVE AND NEGATIVE TURGtDITY- VARIATIONS 63 animal. From the experiment just described, however, it will be seen that the effect observed by him was in reality due, not to true excitation, but to the hydrostatic disturbance, or indirect effect of stimulus. In the next record (fig. 47, c) we see the effect of stimulus applied nearer : that is to say, at a distance of 2 cm. from the proximal contact. Owing to the propinquity of the point of stimulation, the two disturbances are not now sufficiently separated, and the excitatory negative reaction completely masks the hydrostatic positive effect. It is thus seen that, as has been said, one method of exhibiting these two effects separately is to apply stimulus at a point so distant from the proximal contact that there is an interval between the arrival of the two waves of hydro- static and excitatory disturbance respectively. It is obvious, then, that if the tissue under experiment be a good con- ductor of excitation, we must place the point of stimulation at a long distance from the first electrode, in order that the effect of excitation may lag sufficiently behind the hydrostatic wave. Similarly, in a bad conductor of excita- tion, it will be the indirect effect alone which will reach the proximal contact, unless the stimulus applied be very near, and very strong. In order to distinguish these two opposite effects from each other, I shall in future refer to that hydrostatic effect which causes expansion and galvanometric positivity as * the hydro-positive effect,' by way of differentiating it from 'the true excitatory effect,' of negative turgidity- variation and galvanometric negativity. It has already been said that tissues which exhibit ajiigh degree of conduction are characterised by more or less of protoplasmic continuity. Hence, fibro-vascular elements are relatively good, and parenchymatous tissues bad, conductors of excitation. The cells of the potato tuber for this reason exhibit very little power of transmitting excitation. When, therefore, in experimenting with this j77| 64 COMPARATIVE ELECTRO-PHYSIOLOGY tissue, stimulation was caused by application of a hot wire so near as i cm. to the proximal contact, it was the hydro- positive effect alone which reached it, giving rise to posi- tive response. It was only, indeed, by applying the stimulus very near, at a distance of 3 mm., that the true excitatory response of galvanometric nega- tivity was in this case obtained (fig. 48). From what has been said, it will be seen that when a given point is excited by transmitted stimulation, two antagonistic elec- trical effects are induced — one of positivity, due to hydro-positive action, and the other of negativity, due to true excitation. When the stimulator is near to, or co- incides with, the responding point, the tissue is subjected to rapidly succeeding positive and negative turgidity-variations, and the elec- trical indication of the latter being the more intense, it masks the former, and the resulting response is determined by an algebraical summation of the two. In a vigorous specimen, whose excit- ability is great, the excitatory gal- vanometric negativity masks the positivity. The resultant electrical response in general is expressed by the formula Ne — P„, where N^ is the galvano- metric negativity due to the true excitatory effect, and P„ the positivity due to the hydro-positive effect, which immediately precedes it. From this, it is clear that, as regards the resultant galvanometric response of vegetable tissues under stimulation, there may occur the two typical cases displayed in the following table : Fig. 48. Photographic Record of Electrical Responses of Potato- tuber a, Positive response to stimulus applied at distance ; b. Negative response to stimulus applied near. POSITIVE AND NEGATIVE TURGIDITY-VARIATIONS 65 TvpiCAL Cases of Resultant Response Conditions Constituent Factors Resultant Response Excitability great Excitability diminished or nearly abolished N. >P„ N, - positivity. These two factors of external stimulus and internal energy are thus seen to be antagonistic in their general expressions. Ih But while stimulus from outside, impinging on an excit- able area, thus caused an expenditure of energy at that area, by inducing excitatory response there, yet it was also found that, by its indirect effect, it brought about an increase of energy in neighbouring tissues. By the sudden contraction, and expulsion of water from the excited area, energy was transmitted hydraulically, and the consequent positive tur- gidity-variation caused an erectile response of a neighbour- ing motile organ. In the simple case in which the point of receptivity was at a distance from that of response, it was seen that these two effects of stimulus, direct and indirect, ere easily discriminated from one another. But as the •p It I 70 COMPARATIVE ELECTRO-PHYSIOLOGY stimulator was brought nearer and nearer, the two effects became superposed, and one was masked by the other. Even in this case, however, on careful examination, it is possible to infer the results due to the action of the internal factor. Thus, we may suppose stimulus to be applied directly on the pulvinus of Mimosa^ bringing about a responsive fall of the leaf The expelled water from the excited pulvinus will now be forced into the neighbouring tissues, making them over- turgid, and raising their energy above par. On the cessation of stimulus, the water that has been forced away will flow back from the region of heightened, to that of lowered tension, and re-establish the normal turgidity of the pulvinus, which had undergone a negative variation. The erectile recovery of the leaf is thus seen to be, not a merely passive process, but an effect dependent on the internal energy of the plant. This is also shown by the fact that in autumn and winter, when the internal energy is low, the period of recovery is very long, being sometimes as much as eighteen minutes ; whereas in summer, on the other hand, with the increased internal energy of the plant, it takes place nearly three times as quickly. I have elsewhere shown that it is this internal energy which is vaguely referred to as the tonic condition of the tissue, and that it consists of the sum total of energy derived from external stimuli previously absorbed and held latent by the plant. The different forms of stimulus may be very various. We have for instance tonicity, as imparted by light, or phototonus ; by favourable temperature, thermotonus ; by electrical current, electrotonus ; by internal hydrostatic pressure, hydroionus ; or by the presence of favour- able chemical substances, chemotonus. We have seen that, as a general rule, external stimulus and internal energy find responsive expressions of opposite sign, the former inducing a negative, and the latter, a positive turgidity-variation. It is, nevertheless, important to demon- strate the extensive applicability of this law, by many different results and different modes of manifestation. And such a EXTERNAL STIMULUS AND INTERNAL ENERGY yi demonstration would undoubtedly become still more con- vincing, if we should succeed in discovering some mode of response in which the antagonistic effects of internal energy and external stimulus found opposite expressions. Such an example, of a very striking character, I have in my work on ' Plant Response ' shown to be found in growth- response. But the same opposition, between the effects of external stimulus and internal energy, I shall now proceed to demonstrate not only by means of growth-response, but also through mechanical and electrical responses. In the case of growth, the responsive expression of the growing organ, under increased turgidity, consists of an expansive elongation. If the organ be growing at a uniform rate, an increase of internal energy will enhance that rate. But when external stimulus acts directly on the growing organ, the normal rate of growth is retarded during the action of stimulus. Thus it will be seen that though the mechanical response of a motile organ, and the movement of a growing organ, appear so different, yet these two expressions are not fundamentally distinct. For while in one, the application of direct stimulus, causing nega- tive turgidity-variation and contraction, induces depression of the leaf, in the other, the same negative turgidity-variation and contraction under external stimulus causes a depression of the rate of growth. And, on the other hand, indirect effect of stimulus, or increase of internal energy, inducing positive turgidity-variation, brings about in the one case the erection of the leaf, in the other an enhancement of the normal rate of growth. This parallelism is displayed in detail in the table given below. It is thus understood that the indication of response to external stimulus is the depression of the motile leaf, or depression of the rate of growth, while the effect of increased internal energy is the erection of the motile leaf, or enhance- ment of the rate of growth. We shall first deal with the responsive expression to that positive turgidity-variation which is due to the increase of internal energy. One mode of increasing the internal 72 COMPARATIVE ELECTRO-PHYSIOLOGY Tabular Statement showing Comparative Effects of External Stimulus and Internal Energy on Pulvinated and Growing Organs Mechanical response Growth response Effect of normal turgidity : Normal horizontal position of leaf. Effect of norfnal turgidity : Uniform rate of growth. Local action of external stiimilus : Local action of external stimulus : Contraction ; Contraction ; Diminution of turgidity ; and concomitant depression of leaf. Diminution of turgidity ; and concomitant depression of rate Action of internal energy exhibited by (a) Recovery; Re-establishment of turgidity and of growth. Action of internal energy exhibited by {a) Recovery : Re-establishment of turgidity and gradual return of leaf to normal gradual return of organ to normal horizontal position. [b) Increased hydrostatic pressure : Erection of leaf. rate of growth. [b) Increased hydrostatic pressure : Increased rate of growth. ' energy of a plant is by a moderate rise of temperature. And this finds expression in the case of a motile organ, by the erection of the leaf. Thus, when Mimosa is raised in tem- perature, all its leaves become highly erect. A diminution of energy, on the other hand, by cooling, brings about a depression of the leaves. In the same way, the rate of growth is exalted by rise of temperature. Thus in a growing flower of Crinum lily the normal rate of growth at 30° C. was -0040 mm. per minute, and this was exalted to •on 3 mm. per minute, or nearly three times, when the temperature was raised to 35*5° C. Lowering of temperature, on the other hand, greatly de- presses the rate, and may even, if it proceed far enough, cause arrest of growth. As regards external stimulus, on the contrary, we have seen that its effect on the motile organ is one of depression. The following record (fig. 50) shows that it has a similar influence on the rate of growth. The first part of the curve shows the normal rate of elongation. But after the applica- tion of stimulus of light, growth is not only retarded, but there is an actual shortening of the organ. On the cessation of stimulus, the normal rate of growth is gradually re- established. EXTERNAL STIMULUS AND INTERNAL ENERGY 73 I shall next give examples in which the opposite effects of external stimulus and internal energy are exhibited in growth response, motile response,' and electrical response. To take first the response of growth : we have seen that a steady rise of temperature brings about an increase of internal energy, while a sudden variation of temperature acts as a stimulus. Thus, if we effect a sudden augmenta- tion of temperature, this will act on the organ, during the period of variation, as a stimulus; but afterwards, when the temperature itself, or its rate of rise, has become Fig. 50. Longitudinal Contraction and Retardation of Growih under Light in Hypocotyl of Sinapis nigra The first part of the curve shows the normal rate of growth. Arrow ( \ ) indicates moment of application of diffuse light, which is seen not only to retard growth, but also to induce a marked contraction. The second arrow indicates moment of withdrawal of light, and dotted portion of the curve shows recovery. steady, the condition will act by increasing the internal energy of the organ. These opposite results are seen to be strikingly illustrated by growth response in the case of the following record (fig. 51), The normal rate of growth at 34° C. was here -015 mm. per two minutes. By a sudden application of heat, raising the temperature of the chamber ultimately by i°C., a respon- sive contraction was caused, as is seen in the record. But, on the attainment of a steady augmented temperature of 35° C, an increased rate of growth, which now amounted to K24 mm. per two minutes, was observed, owing to the 74 COMPARATIVE ELECTRO- PHYSIOLOGY Or the same difference may be demonstrated by means of mechanical response. A Mimosa is placed in a small chamber and subjected to a sudden rise of temperature. In consequence of this there is a preliminary excitatory depression, followed, on the attainment of a steady rise, by gradually increasing erectile re- sponse, which carries the leaf above its original level. These opposed motile effects can be shown, moreoyer, even in the case of ordinary plants. We take a spiral tendril of Passiflora. In this, the outer or convex surface is more ex- citable than the inner or con- cave, and external stimulus, causing greater contraction of this more excitable outer side, induces a movement of un- curling. This movement corre- sponds to the excitatory fall of Mimosa. The response by increase of internal energy is, however, the opposite of this, and consists of a movement of curling. When the tendril is The rate of growth became con- placed in a vessel of water, of slant when the temperature be- i • i .1 ^ ^ l came peniianent at 35° C. ^hich the temperature can be varied at will, a sudden rise of temperature causes a preliminary excitatory movement of uncurling, followed by a movement of curling, when the higher temperature has become steady. The electrical expressions of external stimulus and in- ternal energy are similarly opposed. In fig. 85 (Chapter X.) will be seen a record showing that sudden variation of temperature, acting as an external stimulus, induced a responsive galvanometfic negativity, whereas steady rise of Fig. 51. Record of Growth in Crinwti at Temperature of 34° C. and 35° C. The dotted line represents the variable period of temperature change. Note the contractile twitch and transient highly ac- celerated growth which follows. EXTERNAL STIMULUS AND INTERNAL ENERGY 75 temperature had the opposite effect — that, namely, of inducing galvanometric positivity. It is thus seen that while the characteristic effect of external stimulus on an excitable tissue is to cause a nega- tive turgid ity-variation, that of increased internal energy is to induce a positive turgidity-variation. The former of these, or negative turgidity-variation, finds electrical expres- sion in galvanometric negativity ; in a motile organ by the fall of the leaf, and in a growing organ by retardation of the rate of growth. The latter, or increase of internal energy, on the other hand, is expressed electrically by galvanometric positivity ; mechanically, by erection of the leaf ; and in a growing organ by an acceleration of the normal rate of growth. We thus arrive at the following laws of response of isotropic organs : 1. Effective stimulation induces contraction and galvano- metric negativity. 2. Increase of internal energy induces expansion and galvanometric positivity. CHAPTER VII ABSORPTION AND EMISSION OF ENERGY IN RESPONSE Sign of response determined by latent energy of tissue, and by intensity of external stimulus — Sub-tonic, normal and hyper-tonic conditions — The critical level — Outward manifestation of response possible only when critical level is exceeded— Three typical cases : response greater than stimulus ; response equal to stimulus ; and response less than stimulus — Investigation by growth-response — Instance of sum of work, internal and external, performed Iby stimulus constant — Positive response of tissues characterised by feeble protoplasmic activity or sub-tonicity — Enhancement of normal excitability of sub-tonic tissue by absorption of stimulus. We shall find, in this and succeeding chapters, that the nature and intensity of response are determined not merely by the intensity of stimulus, but also by the molecular con- dition of the responding substance. The excitatory mani- ^tation is dependent upon the occurrence of a particular direction^d molecular distortion. Hence, if by the action of the stimuli of the environment, an incipient distortion in this direction has already been induce^ in the tissue, the incidence of even moderate stimulus will then prove sufficient to precipitate visible excitatory manifestation. A tissue in this condition is said to be highly excijable or fully tonic. If, on the other hand, the tonic condition be less favourable, long- continued stimulation will be necessary to evoke the excita- tory effect. Here, during the first part of application, stimulus will appear to be ineffective. As a matter of fact, however, it is at work to give such a predisposition to the molecules that the action of subsequent stimulus shall be rendered effective. In the simple case in which the tonic condition is favour- able, a given stimulus will induce a given responsive expres- ABSORPTION AND EMISSION OF ENERGY IN RESPONSE 77 sion. If further, the tissue, on recovery, return to its original condition, then a second similar stimulus will induce the same responsive expression as the first. These responses will thus be uniform. But if, owing to the after-effect of stimulus, the condition of the tissue itself be changed, the responses will be found to exhibit either staircase increase or fatigue decline, according to the particular molecular con- siderations involved, which will be described in detail in the 1^^ following chapter. I^P It will be interesting here, however, to take an extreme case of a tissue in the opposite condition — namely, of great sub-tonicity. It is clear that since the excitability of the tissue is here feeble, there will be little or no outward manifestation of excitatory response. The incident stimulus will thus be absorbed, and entirely held latent. And the 1^ increase of internal energy thus brought about may find B expression mechanically by expansion, or electrically by galvanometric positivity. 1^ In the case which we have selected, the excitability of the Btissue is too low for the ordinary excitatory expression to occur. Hence the incident stimulus will be entirely absorbed, and there will be a gain of energy without any loss. The whole responsive expression will in this case consist, not of con- traction but of expansion. In other words, it will be exactly opposite to that which is usually consequent upon external stimulus. In order to demonstrate this, I took a seedling of Taniarindus indicus which had been cut off from its supply of external energy, and was consequently sub-tonic. Owing to the insufficiency of internal energy, its growth had, in fact, come to a standstill. On now subjecting this seedling to thermal stimulation, the absorbed stimulus raised the internal energy and found expression in growth expansion. We have seen that the effect of external stimulus on a growing organ in normal tonic condition was the retardation or arrest of growth. But here, in a tissue which is sub-tonic, we find t" that the effect is exactly opposite. 78 COMPARATIVE ELECTRO-PHYSIOLOGY J which sub-tonicity is manifested by arrest of growth, we may select a specimen in which, while the tissue is not fully tonic, there is still, nevertheless, a feeble rate of growth. In such a case we may expect the income from the absorption of stimulus to prove greater than the expenditure in the form of true excitatory response. Hence, if we subject such a tissue to the constant action of an external stimulus, we shall in the first stage obtain the predominant effect of the internal factor, with its positive turgidity-variation and enhanced rate of growth. But by the continued action of this accumulating income, the tonic condition of the tissue will be raised to the normal, with a concomitant increase of excitability. It will now therefore be the excitatory component which becomes pre- dominant, resulting in the negative turgidity-variation, con- traction, and retardation of growth. In order to detect these variations of the normal rate of growth, under the action of stimulus, it is necessary to have at our disposal some very delicate means of record. This need I have, however, been able to meet, by devising the Balanced Crescograph, more fully described in my book on * Plant Response.' Here, the uniform rate of elongation of a growing organ causes a rotation of the recording Optic Lever. The spot of light from this lever falls upon a second mirror, which is subject to a compensating movement. When the balance is exact, the spot of light, reflected from the two mirrors, remains quiescent. When, however, the normal rate of growth, under the action of any agent, undergoes varia- tion, the balance is upset. Thus, when growth is accelerated, there is a movement of the recording spot of light in one direction, say up, and when retarded, a movement in the opposite, say down. In order to study the effect of external stimulus on a tissue in a slightly sub-tonic condition, I took a flower-bud of Crinum lily, and first obtained a balanced record, seen as a horizontal line (fig. 52). Stimulus of light was now applied, ftnd it will bQ §eQn th^t after a short latent period the ABSORPTION AND EMISSION OF ENERGY IN RESPONSE 79 absorbed stimulus induced a positive turgidity-variation, with enhanced rate of growth, as seen by the up-curve. But by this absorption of the stimulus itself, the tonic condition of the specimen was raised, with consequent increase of ex- citability, and the response became normal. That is to say, it now consisted of contraction and retardation of growth as seen by the downward curve. The external stimulus was now cut off and the (dotted portion of the curve shows the after- effect. The after-effect is thus not a mere re- covery, but an enhanced IKrate of growth, due to the increased energy which remains latent. It was only when this was ex- hausted that the normal rate of growth was re- established, as seen in |^:the horizontal part of the Fig. 52. Balanced Record of Variation of Growth in Flower-bud of Crinum Lily under Diffuse Stimulation of Light Continuous lines represent the effect during application of light, the dotted line on withdrawal of light. The plant was originally in a sub-tonic condition, and application of light at x , after short latent period, induces preliminary ac- cderation of growth. After this follows the normal retardation. On withdrawal of light, in the dotted portion of the curve is seen the after-effect, followed by return to the normal rate of growth. A second and long-continued application of light induces retardation, followed by oscillatory response. curve. And as the tissue was now in full tonic [^■Condition the renewed ^^■application of stimulus Hof light did not again ■ induce a preliminary en- chancement of the rate of growth, but the normal contraction and retarda- l^ption. Its long-continued application gave rise to the further phenomenon of multiple response, a subject which will be I fully dealt with in a future chapter. Now owing to this fact that the response of growth gives us by means of the enhancement or depression of its rate, fefifects which correspond to the positive and negative, wc ar^ So COMPARATIVE ELECTRO-PHYSIOLOGY (i) That when the tonic condition or the excitabih'ty of the tissue is low, the predominant effect will be the positive. This has been shown during the course of the present chapter in the case of a very sub-tonic tissue of Tamarindus indicus^ where the positive or growth-expansion effect was initiated by the action of stimulus. (2) That when the sub-tonicity of a tissue is not very great, incident stimulus will at first give the positive effect of an enhanced rate of growth. But with the absorption of the stimulus itself, the tonic condition of the tissue will be raised, and we shall then obtain the true excitatory reaction of con- traction and retardation of growth. Thus, in this intermediate case, the positive response will be seen to pass into normal negative. (3) And, lastly, that when the tonic condition is already high, the excitatory negative response will predominate and we shall obtain normal contractile response. Both this and the previous intermediate cases are illustrated by the experi- ment described on Crinum lily. It was there seen that the first effect of incident light was positive, the tissue being sub-tonic ; subsequently, the tonic condition being raised, this response was converted into the excitatory negative. And on renewed application of stimulus thereafter the immediate response continued to be negative. The fact that by means of growth-response, it is possible to obtain indications of the external and internal work per- ; formed by absorbed stimulus, enables us to demonstrate a proposition of great importance, that, namely, under certain [conditions, the sum of the work done, internally and externally, by a given stimulus, is constant. This will be the case where there is little or no dissipation of energy in the course of transformation. In considering the question of the relative proportions of the incident stimulus utilised for in- ternal and external work respectively, we find it clear, from considerations already adduced, that the lower the tonic con- dition the greater will be the proportion of stimulus held latent for the performance of internal work. The nearer is ABSORPTION AND EMISSION OF ENERGY IN RESPONSE 8 1 the tonic condition, on the other hand, to the critical level, the greater will be the excitatory overflow, and the smaller the latent component. The internal and external factors will thus be complementary to each other. On subjecting this inference to experimental demon- stration by means of growth-response, I fully succeeded in verifying it. According to this method of growth-response, it will be remembered, the true excitatory effect is measured by retardation of the normal rate of growth, the internal factor of increased latent energy being represented, on the other hand, by a corresponding enhancement of the rate of growth. This being understood, it was found that in a particular specimen of growing tissue, whose tonic condition was somewhat low, the external and internal effects caused by a given stimulus were in the proportion of 32 to 13-5. When the tonic condition of the specimen was raised, how- Ufever. and the same stimulus was applied, the external effect was found to be enhanced to 38 at the expense of the internal, which was now found to be lowered to 85. The sum of the work done, both internally and externally, is seen to be in both these cases approximately the same, being in the former experiment 45*5, and in the latter 46-5. We have seen that, of the two antagonistic factors of response, the positive will predominate if the excitability of the tissue be in any way diminished. Such a loss of excitability may occur in either of two ways: (i) by the sub-tonicity of the tissue itself; (2) by the depression con- sequent on fatigue. Under either of these conditions then we may expect to obtain the exhibition of the positive effect. The exhibition of the positive effect under fatigue will be described in the course of the next chapter. We shall here consider instances in addition to those already given, of the occurrence of the positive effect in a tissue which is sub-tonic. We have to bear in mind that the work which incident stimulus is called upon to perform is two-fold, both internal and external, and that there is a certain critical excitatory level, above which only is the normal responsive expression G 82 COMPAkATIVE ELECTRO-PHYSIOLOGY NT ST possible. The actual potential or excitatory level of a tissue depends on its tonic condition and the intensity of the incident stimulus. Now this existing potential of the tissue may be anything within a wide range, s T, when sub-tonic, N when n'ormal, or H T when hyper- tonic or above the ordinary normal degree (fig. 53). Since C it is necessary that the incident stimulus should cause the critical level C to be slightly exceeded, if there is to be an excitatory overflow, we can see that the intensity of the stimulus re- quisite to evoke response will be greater in proportion as the tonicity of the tissue itself is low. Thus when the tissue is extremely sub-tonic, a stimulus of or- dinary intensity could never avail to raise the energy of the system above the critical point, and the response must then therefore be positive. Under these circumstances it will only be by the impact of excessively strong sti- mulus, or by the cumulative action of a series of moderate stimuli, that the critical point can be reached and passed, and the normal negative response evoked. Thus the intensity of the minimally-effective stimulus in evoking normal response will afford us a measure of the tonicity of the tissue. If the latter be high, then the feeblest stimulus will precipitate outward response, and indeed, if excessive, response will occur on little or no provocation, and such movements we call ' autonomous.' It must be remem- bered, however, that it was by the previous absorption of stimuli that the tissue was brought to this point of unstable equilibrium at which the added impact of an infinitesimal stimulus causes it to bubble over, as it were, into apparently spontaneous activity. The predominant expression of the highly tonic tissue Fig. 53. Diagrammatic Representation of the Tonic Level N, normal ; s t, sub-tonic ; H T, hyper-tonic ; and c, the critical level. ABSORPTION AND EMISSION OF ENERGY IN RESPONSE 83 being thus negative, we must go to the other extreme of great sub-tonicity if we are to be successful in demonstrating the occurrence of the unmixed positive response. This considera- tion leads us to expect that positive response will be evoked o n mode rate stimulation from tissues that are either not highly tonic or protoplasmically defective. I shall show in CKapter^XXII. that in cells of epidermis, where the proto- plasmic contents have been reduced to a minimum, response to moderate stimulus tends in general to be positive. Even highly excitable tissues like nerve, as will be shown later IB when cuFofT^from their supply of energy, often become so sub-tonic as to give positive response. I shall liere show I how ordinary tissues exhibit this effect, when the tonic con- Bdition is allowed to fall to such an extent as to render the tissue extremely sub-tonic. For this purpose I took a cut specimen of petiole of cauliflower, and kept it without water for a couple of days. By this process the specimen became IK somewhat withered. I next proceeded to take records of its electrical responses under increasing stimuli. The intensity of these stimuli rose from i to 10 units. It will be seen from the record (fig. 54) that each stimulus up to 9 evoked positive response, and that it was the strong stimulus of 110 which gave rise to the normal response of negativity. This constitutes the first instance of a phenomenon which I shall show later to be of very extended occurrence — the induction, namely, of one effect under moderate, and its opposite under very feeble stimulation. It is not so easy to demonstrate this fact with a highly excitable, as with a some- what sub-tonic tissue, where the critical intensity of stimulus for the evoking of normal response need not be impracticably low. A point to be taken into account here is the after-effect IH of sub-minimal stimulus in enhancing subsequent normal ? excitability. Thus it is found in taking the record of responses to a succession of feeble stimuli, that though they tare at first abnormal positive, they are afterwards converted into normal negative. That it is the after-effect of the previous stimulation which thus enhances previous excitability G 2 84 COMPARATIVE ELECTRO-PHYSIOLOGY may also be demonstrated by subjecting the tissue to con- tinuous stimulation or tetanisation, when the abnormal positive is found to pass into normal negative. From the experiments that have been described, it would appear that the several kinds of response characteristic of various tissues are relatively rather than absolutely different. The true excitatory reaction of an excitable tissue, is one of galvanometric negativity. Any diminution of the ex- citability — whether by lowering of tonic condition or other Fig. 54. Photographic Record of Abnormal Positive passing into Normal Nega- tive Response in a Withered Specimen of Leaf-stalk of Cauliflower Stimulus was gradually increased from i to 10, by means of spring-tapper. When the stimulus intensity was 10, the response became reversed into normal negative. (Parts of 8 and 9 are out of the plate.) This record is to be read from right to left. Down-records stand for positive, and up-curves for negative responses. causes — will bring about a decrease of this negativity, which may culminate in actual positivity. Thus negative is not separated from positive response by any break of continuity ; but we are able, on the contrary, to trace a gradual transition from one to the other. Moreover, in every response we have the two antagonistic elements, positive and negative, either actually or potentially present. The form taken by the resultant response is entirely determined by the question of what proportion of the stimulus impinging upon the tissue becomes latent ; and this in its turn depends upon the tonic ABSORPTION AND EMISSION OF ENERGY IN RESPONSE 85 condition of the tissue. When the absorbed stimulus is wholly retained, response is positive, but by this absorption the tonicity of the tissue and its excitability are both raised. When the tonic condition of the tissue, on the other hand, is already high, and its excitability great, a large proportion of the energy finds outward expression, and we obtain the normal negative response. Between these two extremes, we may observe many effects of interference, due to the play of the two antagonistic elements. If, then, the time-relations be not coincident, variations will be induced which will find expression in different types, diphasic response, positive followed by negative, and vice versa. The question considered in the course of the present chapter has been that of the energy received and given out by the tissue, and the molecular work, positive and negative, performed during these processes. Such work, however, is itself the result of molecular distortions brought about by stimulus, and the question of the amplitude of response, as related to the degree of distortion, will be discussed in the following chapter. CHAPTER VIII VARIOUS TYPES OF RESPONSE Chemical theory of response — Insufficiency of the theory of assimilation and dis- similation — Similar responsive effects seen in inorganic matter — Modifying in- fluence of molecular condition on response — Five molecular stages, A, B, c, D, E — Staircase effect, uniform response, fatigue — No sharp line of demarcation between physical and chemical phenomena — Volta-chemical effect and by- products — Phasic alternation — Alternating fatigue — Rapid fatigue under con- tinuous stimulation— In sub-tonic tissue summated effect of latent components raises tonicity and excitability — Response not always disproportionately greater than stimulus — Instances of stimulus partially held latent : staircase and ad- ditive effects, multiple response, renewed growth — Bifurcated responsive ex- pression. According to current theories, living matter is maintained in a state of equilibrium by the two opposed chemical pro- cesses of assimilation and dissimilation. It is supposed that stimulus causes a down or dissimilatory change, which is again compensated during recovery by the building-up or assimilative change. In the case of uniform responses, again, these two processes are regarded as balancing each other. On this theory, when the down change is the greater of the two, the potential energy of the system falls below par ; for the building-up process cannot then sufficiently repair the chemical depreciation caused by it. Hence occurs dimi- nution of response, or fatigue, which is supposed to be further accentuated by the accumulation of deleterious fatigue-stuffs. The disappearance of fatigue after a period of rest is ex- plained by the renovating action of the blood-supply, which is also regarded as the means of carrying away the fatigue- stuffs. A serious objection to these explanations lies, however, in the fact, that even excised and bloodless muscles exhibit recovery from fatigue after a period of rest. In isolated VARIOUS TYPES OF RESPONSE 87 vegetable tissues, again, where there is no active circulation of renovating material, the same effect, and its removal after a period of rest, are observed. Thus the difficulties en- countered in explaining fatigue, on purely chemical con- siderations, are great enough ; but still greater are those difficulties which arise when we come to deal with the stair- case effect— typically shown in cardiac muscle — in which IK successive responses to uniform stimuli exhibit a gradual ■™ enhancement of amplitude. The results obtained here are in direct opposition to the theory described ; for in this particular case we have to assume that the same stimulus which is usually supposed to cause a chemical breakdown, I has become efficient to induce an effect exactly the reverse. B Of the two antagonistic elements in the electrical response, moreover, it is the positive which is supposed to be associated with the assimilative, and the negative with the dissimilative change. If this supposition were correct, however, it would IB be natural to expect that the positive response would be manifested predominantly in vigorously growing tissues, in j^ which assimilation must be at its greatest Fatigued tissues I Bon the other hand, in which dissimilatory changes are sup- posed to be predominant, should manifest negativity as their characteristic response ; moribund tissues, in contrast with the actively growing, might also be expected to exhibit respon- sive negativity. In actual fact, however, the very reverse is ^ the case. For in vigorous tissues, normal response is by galvanometric negativity,; and it is the over-fatigued or «/ moribund which characteristically exhibit the positive re- * sponse. It would be difficult again to conceive of assimilation and dissimilation in the case of inorganic matter. Yet even in inorganic matter we find reproduced all the various types met with in the response of living tissues : namely, uniform response^ the staircase effect, anjd fatjg^ue. Response being r eally due to molecular upset from a co ndition of equiUbrium, we can see how different forms of responsive expression will occur, according to the various molecular conditions of the I 88 COMPARATIVE ELECTRO-PHYSIOLOGY substance at the time being. One of the most important factors, then, in determining the character of response is the molecular condition of the substance itself The numerous anomalies hitherto encountered in our interpretation of responsive phenomena are all traceable to our failure to take this factor of molecular condition into account. For a full exposition of the modifying influence which it exercises on response, however, though I shall here state some of the principal conclusions which I have arrived at, the reader is referred to Chapter XLII. From the fact, that every type of response is to be obtained from inorganic matter, where chemical assimilation and dissimilation are obviously out of the question, it is clear that the fundamental phenomenon must be dependent on physical or molecular, and not on such hypothetical chemical changes. It must, however, be remembered that though re- sponse phenomena and their modifications are undoubtedly in the first place physical or molecular, yet in the borderland 1 between physics and chemistry there is no sharp line of demarcation. For example, yellow phosphorus becomes converted, under the stimulus of light, into the red, or allotropic, variety. This molecular change, however, is also attended by a concomitant change in the chemical activity, phosphorus in its allotropic condition being less active than ^ in the yellow. Under certain circumstances, further, it is possible to have a secondary series of chemical events follow- ^ ing upon a condition of unequal molecular strain. A homo- >^ eneous livi n g tissue, when un stimulated, is iso-electric./'^ en stimulated, however, an electro-motive difference is nduced, as between the stimulated and unstimulated parts of .^ he tissue. The result is an electrical current attended )y electro-chemical changes. As a consequence of such ^ rolta-chemical action, when prolonged, by-products (fatigue jtuffs ?) may be accumulated, and these may have a de- )ressing effect on the activity of the tissue. Hence, just as, after very prolonged activity of a voltaic combination, it is necessary to renew the active element and change the VARIOUS TYPES OF RESPONSE 89 electrolyte, surcharged with by-products, so after sustained activity of a living tissue, the process of renewal, or renova- tion, will be necessary. It is thus seen how upon the funda- mental molecular derangement, a chain of very various chemical events may follow, as its after-effect. And it is only by going in this way to the very root of the pheno- Imenon that we can avoid the many contradictions with which we are confronted by the chemical theory. In studying various response phenomena, our conclusions are necessarily based upon the observation of the amplitude ||kDf responses. It is therefore important at this point to draw attention to the danger of hasty inferences. On finding, for instance, that the amplitude of response in a given case is diminished, we are apt to infer that the responding tissue has undergone depreciation. But this is not invariably the case. In the entire process of response, while stimulus induces molecular upset, we must remember that there is also an internal factor, which brings about molecular restitu- tion. Now, if this force of restitution be in any way enhanced, it is easy to see that the responsive distortion of the mole-'^ cules will find itself opposed, with consequent diminution of IBpimplitude. We shall thus often find that a rise of tem- perature, by enhancing the force of recovery, actually causes a diminution of response. That this is not due, however, to any depreciation of the tissue is seen from the facflTiat tfi£ I^same rise of temperature enhances another excitatory pro- "perly of the tissue— namely, the speed of its conduction. I shall now give a brief account of the modifying influence IHtexercised on response by the molecular condition. It will be IHfehown, in the Chapter (XL 1 1) on the Modification of Response IBunder Cyclic Molecular Variation, that a given response is iHnot determined merely by the nature of the responding substance, but also by the amount of the energy which it possesses. Starting from the lowest condition of sub-tonicity, a substance undergoes progressive molecular transformation by the action of the impinging stimulus itself Five stages may be roughly distinguished in this transformation. In the I 90 COMPARATIVE ELECTRO-PHYSIOLOGY first, or A, stage of extreme sub-tonicity, we have absorption without excitatory response. By this absorption the sub- stance passes into the next, or B, stage, which is the stage of transition, where response is converted from the abnormal to normal. Above this stage the rate of molecular transforma- tion is very rapid. From the residual after-effect of stimulus, the substance now passes from the stage B to the stage C, which is a condition of more or less stability. Further stimulation carries the substance to stages D and E. Here the molecular distortion from the normal equilibrium is very great. Stimulation applied in this condition has little further effect in inducing response. That is to say, excit- ability is here reduced to a minimum. In this extremely distorted position, moreover, the substance has a strong tendency to revert to the position of normal equilibrium. In the A condition of extreme sub-tonicity, since there is absorption without excitation, the response which we obtain is abnormal positive. Intense or long-continued stimulation carries the substance into the B stage, with its normal negative response often preceded by diphasic. An example of this has already been given in fig. 54, obtained from the sub-tonic petiole of cauliflower. We shall meet, however, with numerous other examples in a great variety of tissues. Arriving at the B stage, the substance is still somewhat sub-tonic, and the rate of molecular transformation here is rapid. From the after-effect of stimulus the mole- cules of the somewhat inert substance become incipiently distorted in the same direction as that of normal response. A proportion of the incident stimulus is thus utilised in inducing a favourable molecular disposition. A repetition of the original stimulus will now give rise to a greater excitatory reaction than before. Thus at the B stage we obtain a stair- case increase of response. This fact — that by the after-effect of previous stimulation the molecules may be incipiently dis- torted in a direction favourable to excitatory response — finds illustration in stimuli individually ineffective being made effective by repetition. The result here is evidently made VARIOUS TYPES OF RESPONSE 91 I conspicuous by the summation of the after effects of all the preceding stimuli with the direct effects of their successors. The staircase effect is seen in the two accompanying records. In fig. 55 is given a photographic record of the staircase increase in the electrical response of a vegetable nerve ^ in somewhat sub-tonic condition. In fig. 56 we have a second example of this effect, seen in the electrical response of the petiole of Bryophyllum, rendered artificially sub-tonic by cooling. We next arrive at the C stage, which is, as has been said, one of more or less stability. Expenditure is here, for a certain length of time, balanced by income. The molecular condition of the tissue being thus constant, the responses are uniform. I give below records of such uniform re- sponses to uniform stimuli, ex- IG. 55. Photographic Re- cord of Staircase Response in Vegetable Nerve Fig. 56. Staircase Increase in Electrical Response of Petiole of Bj-yophylluin, rendered sluggish by cooling t Hhibited by different tissues. In fig. 57 are seen uniform electrical respon.ses to uniform mechanical stimuli, given l^by the root of radish. Fig. 58 shows uniform electrical l^responses to uniform thermal stimuli, given by the petiole of fern. tThe C is succeeded by stages D and E, representing L condition of over-strain. In fig. 59, a, are shown uni- brm responses to uniform stimuli, applied at intervals ' An account of the discovery of certain vegetable tissues, with the function of nerves, will be found in Chapter XXXII. 92 COMPARATIVE ELECTRO-PHYSIOLOGY of one minute. An inspection of the record shows that there is in such cases a complete recovery, at the end of which the molecular condition is the same as before stimula- tion. Hence, successive responses are exactly similar to each other. The stimulation-rhythm was now chang^ed, to intervals Fig. 57. Photographic Record of Uniform Responses (Radish) of half a minute instead of one, while the stimuli were main- tained at the same intensity as before. It will be noticed (fig. 59, ^) that these responses are now of much smaller .LLli^'L Fig. 58. Photographic Record of Uniform Response in Petiole of Fern amplitude, in spite of the equality of stimulus. An inspec- tion of the figure also shows that, when greater frequency of stimulation was introduced, the tissue had not had time to effect complete recovery from previous strain. The mole- cular swing towards equilibrium had not yet abated, when VARIOUS TYPES OF RESPONSE 93 the new stimulus with its opposing impulse was received. There is thus a diminution of height in the resultant (a) (S) (c) Fig. 59. Record showing Diminution of Response, when sufficient Time is not allowed for Full Recovery In {a) stimuli were applied at intervals of one minute ; in {d) the intervals were reduced to half a minute ; .this caused a diminution of response. In {c) the original rhythm is restored, and the response is found to be enhanced (Radish). y u u Fig. 60. Fatigue in Celery Vibration of 30° at in- tervals of half a minute. ■L response. The original rhythm of one minute was now restored, and the succeeding records (fig. 59, c) at once show increased response. Residual strain is thus seen to e one of the principal reasons of reduced response or fatigue. This is also shown in a record which I have obtained with a petiole of celery (fig. 60). It will be noticed there that, owing to imperfect molecular recovery, during the time allowed for rest, the heights of succeeding responses undergo Stimulus : 30° a continuous diminution. Fig. 61 gives a photographic record of fatigue in the petiole of cauliflower, and fig. 62 of fatigue in inorganic response. tit is evident that residual strain, other things being equal, Fig. 61. P'atigue in Leaf-stalk of Cauliflower vibration at interval one minute. IS 94 COMPARATIVE ELECTRO-PHYSIOLOGY seen in fig. 63, where the first set of these responses, A, is for an intensity of mechanical stimulation of 45° vibration, and the second set, 13, of augmented amplitude, for an intensity of 90° vibration. On reverting, in c, to the first stimulus- intensity of 45°, the re- sponses are seen to undergo a great diminution, as com- pared with the first set, A. This change is due to the over-strain of the previous excessive stimulation. But we should expect that the effect of such over-strain would disappear with time, and the responses regain their former height, after a period of rest. In order to verify this, therefore, I re- newed stimulation (at the intensity of 45°) fifteen minutes after C. It will be seen from the record D how far fatigue had been removed in this interval. Fig. 62. Photographic Record showing Fatigue in Tin Wire which had been continuously stimulated for several Days A 90 ■ ^J M A u B 4-5 4-5 Fig. 63. Effect of Over-strain in producing Fatigue Successive stimuli applied at intervals of one minute. The intensity ot stimulus in C is the same as that of A, but response is feebler owing to previous over-stimulation. Fatigue is to a great extent removed after fifteen minutes' rest, and the responses in D are stronger than those in C. The vertical line between arrows represents -05 volt. (Turnip leaf-stalk.) One peculiarity that will be noticed in these curves is that, owing to the presence of comparatively little strain, the first response of each set is relatively large. The succeeding VARJOUS TYPES OF RESPONSE 95 responses are approximately equal, where the residual strains are similar. The first response in fig. 6^, A, shows this, because there had been long previous rest. The first of B shows it, because we are there passing for the first time to an increased intensity of stimulus. The first of C does not show it, because of the strong residual strain from the preceding excessive stimulation. And the first of D, again, does show it, because the strain has now been removed, by the interval of fifteen minutes' rest. Ih Of the antagonistic elements of positivity and negativity which are present in response, we have seen that the positive becomes predominant when the excitability of the tissue is in any way depressed. And since a tissue under fatigue has |H|ts excitability lowered, it follows that in this condition it may be expected to exhibit a tendency towards positive I response : that is to say, expansion in the case of mechanical ^kmd galvanometric positivity in the case of electrical, response. Thus, when a tissue is subjected to continuous stimulation, tthe first effect will be the maximum negative response, contraction and galvanometric negativity. But on the setting- in of fatigue, the positive effect will predominate, inducing a fatigue-reversal of the response. In cases where such fatigue is very great, as, for instance, in certain muscles, IBthe top of the tetanic curve undergoes rapid decline (fig. 64, a). ' The normal contraction now exhibits a reversal, or relaxation. In the sensitive plant, Mimosa, similarly, continuous stimula- tion by electrical shocks gives rise to results which are essen- tially the same. It will be noticed that after the responsive fall of the leaf it returns to its former erect position, in spite of the fact that stimulus is still being continued. Here also, ■■as in the corresponding case of muscle, we have the usual sequence, of (i) normal contraction and (2) fatigue relaxation I— (fig. 65). I^ In electrical response, also, under continuous stimulation, the normal galvanometric negativity, owing to the increasing positive effect, undergoes decline or abolition. This is seen in fig. 64, d, which exhibits the decline of electrical response I 96 COMPARATIVE ELECTRO-PHYSIOLOGY under continuous stimulation in the petiole of celery. The fatigue in the mechanical response of muscle under similar conditions is given in a for the purpose of comparison. The effect of rest in inducing molecular recovery, and hence in the removal of fatigue, is illustrated in the following set of photographic records (fig. 66). The first of these shows the curve of electrical response, obtained with a fresh plant. It will be seen that under a continuous stimulation of two minutes the response first attains a large amplitude, after which it declines, in a fatigue-reversal. Another two minutes were now Fig. 64. Rapid Fatigue under Con- tinuous Stimulation in (a) Muscle ; (<^) Leaf-stalk of Celery (Electrical Response) Fig. 65. Photographic Records of Normal Mechanical Response of Mimosa to Single Stimulus (upper figure), and to Continuous Stimu- lation (lower figure) In the latter case the leaf is erected in spite of continuous stimulation. allowed for recovery, and we observe that a partial recovery takes place. Stimulation was now repeated throughout the succeeding two minutes, to be followed once more by two minutes' rest. The response in this case is seen to be decidedly smaller than at first. The same effects are seen in the third response. A period of rest of five minutes was next given, and the curve subsequently obtained under the VARIOUS TYPES OF RESPONSE 97 same two minutes' stimulation as before shows greater response than the preceding, owing to the partial removal of residual strain. There is one aspect of the subject of fatigue-reversal which now demands our attention. We have seen that under continuous stimulation, a maximum contraction is induced, which is attended by the depression of the leaf of Mimosa. I P'iG. 66. Effect of Continuous Vibration (through 50°) in Carrot In the first three records, two minutes' stimulation is followed by two minutes' recovery. The last record was taken after the specimen had a rest of five minutes. The response, owing to removal of fatigue by rest, is stronger. ,^with r l^lhcory I This is followed, however, by a reversal — namely, expansion, with re-erection of the leaf According to the chemical of a.ssimilation and dissimilation, the fatigue-effect is assumed to be due to an explosive dissimilatory change, with consequent run-down of energy. In the case of Mimosa^ however, it is difficult to understand how, by a mere run- down of energy and consequent passivity of the tissue, an active movement of erection — involving the performance of work in lifting the weight of the leaf — could be brought H 98 COMPARATIVE ELECTRO-PHYSIOLOGY about. Now we have seen that the diminution of normal response may be brought about by the augmentation of the internal factor, tending to enhance the force of restitution, and the necessary augmentation of the internal factor may be the result of an increase of internal energy. Thus while the plant is the recipient of a continuous income, its responsive expression is alternately one of emis- sion and absorption of energy. Thus negative and positive succeed each other or vice versa. Such a phasic alternation is widely present, as we shall see, in the response, not only of various living tissues, but also of inorganic substances. The follow- ing record (fig. 6"j) exhibits this oscillatory response in arsenic under the continuous stimulation of electric radiation. In the case of Mimosa^ under continuous stimulation, we obtain Fig. 67. Oscillatory Re- sponse of Arsenic acted on Continuously by Hertzian Radiation Taken by method of con- ductivity variation. * \ A f \N vv ". Fig. 68. Alternate Fatigue (a) in Electrical Responses of Petiole of Cauliflower ; {b) in Multiple Electric Responses of Peduncle of Biophytum ; (r) in Multiple Mechanical Responses of Leaflet of Bio- phytum ; and {d) in Autonomous Responses of Desmodium a single alternation, and a certain period must then elapse, before the response can be repeated. In other cases, how- ever, continuous stimulation may give rise to two, or three, or a large number of similar alternations. VARIOUS TYPES OF RESPONSE 99 In connection with this subject of phasic alternation I may describe a certain curious phenomenon, which 1 have often noticed ; I refer to the periodic waxing and waning of both mechanical and electrical responses. The simplest example of this will be a case in which the responses are Ualternately large and small. But others are to be found in which the groupings are more complex. In fig. 68 a is seen such a simple alternation, in the electrical response of the petiole of cauliflower, under successive uniform stimuli. In ^^^^^^^^^ d, c, and d are shown similar alter- I^^^^^^^Hh nations in multiple and autonomous ML ^^^^B^Hll responses. I give also a photographic H ^■l^Hl^HII record (fig. 69) of a similar alterna- I ^H^^U^^hI ^^^" ^" ^^^ automatic pulsations of H liBllHIiH ^^^ leaflet of Desmodium gyrans. Fig imilar alternations are sometimes observed in the beating ,^of frog's heart (fig. 70). ly In the following record of mechanical response (fig. 71), taken from the style of Datura alba, we find that fatigue, as already understood, would not explain the phenomenon observed. For here, under the continuous action of stimulus, without any intervening period of rest for the so-called * assimilatory ' recuperation, we see that a second response occurs. I shall later give other instances in which pulsating responses, with their alternating negative and positive phases, Kre given, under the action of continuous stimulation. We ass here imperceptibly from the ordinary phenomenon of Fig. 69. Pholograpliic Record of Periodic Fatigue in the Auto- matic Pulsation of Des- inodium gyrans m 1 ■ . WWWWi . . 1 . 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Fig. 70. Periodic Fatigue in Pulsation of Frog's Heart (Pembrey and Phillips) lOO COMPARATIVE ELECTRO-PHYSIOLOGY individual response to individual stimulus, into that of multiple response, either to continuous, or to a single strong stimulation. The excess of energy derived from impinging stimulus is in the latter case held latent in the tissue to find subsequent expression in phasic alternations of negative and positive variations in series (cf. Chapter XVII). There can be no doubt that these effects of periodic alternation of phase are due to two antagonistic reactions, becoming effectively predominant by turns. Thus the con- tinuous impact of stimulus on a tissue may first give rise to the negative phase of response. But by the continuous absorption of incident stimulus, the internal energy is in- creased, with its opposite reaction of positivity. Hence, the negativity will be gradually diminished, anH" the positive phase^ become predominant. The existence of these two antagonistic factors will be understood, from an inspec- FiG. 71. Photographic Record of ^ion of the top of a tetanic Periodic Fatigue under Continuous curve. Here, the more Or Stimulation in Contractile Response , , . i ,. (Filament of ^r/r/w Lily) less horizontal Ime repre- sents a state of balance between the two opposite forces of excitatory response by contraction, with galvanometric negativity and recovery or expansion, with galvanometric positivity. When this state of balance is disturbed, by a sudden cessation of the hitherto continuously acting stimulus, a brief overshooting of the response in the negative direction is sometimes seen, followed by recovery. We shall meet with examples of this in, among others, the responses of retina and certain vegetable tissues under light. Such facts it has been suggested afford a demonstration of the two antagonistic processes of assimilation and dissimilation, characteristic of living tissues. But that they are really to be accounted for VARIOUS TYPES OF RESPONSE lOI 1^' from molecular considerations will be seen from the fact that effects exactly similar are met with in the response of inorganic matter (cf figs. 258 and 383.) In the case of responses exhibiting fatigue from over- strain, we have a diminution of normal response, which may ultimately culminate in reversal. We may imagine a spiral spring, undergoing increasing compression from a gradually augmenting force. The responsive compression will at first be considerable. But this will soon reach a limit, beyond hich added force will seem to have but little power to induce further responsive distortion. In a somewhat similar way, we may visualise the condition of the responding molecule at he stage D or E. Here, molecular distortion has almost ached its limit. It follows that added stimulus can induce Fig. 72. Fatigue in the Contractile Response of India-rubber Note the periodic alternation and the reversal at the end. little further distortion. But the maximally distorted mole- cule has now a great tendency to revert to the position of equilibrium, and the shock of stimulus, instead of inducing excitatory acti on, induces the reverse. That this is to be ex- plained by molecular rather than chemical considerations, is seen in the following record (fig. 72) of the contractile ■response of india-rubber to thermal stimulation. This fcepresents the last part of a long series of responses, whose amplitude was already undergoing a progressive decline. Further symptoms of growing fatigue are seen in the periodic alternations of amplitude, and in the final reversal of response 1^0 one of expansion. I shall later give another record in which the normal negative response is seen reversed to positive through an intermediate diphasic. I02 COMPARATIVE ELECTRO-PHYSIOLOGY The fact that the normal response of Hving tissues may be reversed under fatigue, I am here able to show by an experiment of an unexpected character. It is usually supposed that fatigue is typical of such tissues as muscle, and absent from nerve. But I shall show with regard to all the various types of response, that there is none of these which is distinctive of any one tissue. The difference is one of degree and not of kind. The same intensity and duration of stimulus which is efficient to cause fatigue in muscle will not be enough to do so in the case of nerve. But even nerve will display fatigue when ex- cessively stimulated. In the record given in fig. 73, a particular nerve of frog had been previously fatigued, by over- stimulation, and on now taking individual responses to individual stimuli, it was found that they had become reversed to positive. Thus a particular type of response is the result of a particular condition of the responding substance, and there is none which is exclusively characteristic of any one tissue. Were it otherwise, ordinary muscle, in which the explosive molecular change is supposed to be so predominant, should typically show only the fatigue, and never the staircase effect. But the following record (fig. 74) shows that this is not the case. For at first it exhibits a characteristic staircase effect ; the responses are then for a time uniform ; and lastly, we see fatigue, in a manner exactly corresponding to the theoretical considera- tions which we have anticipated in stages B, C, and D. The staircase response is thus not peculiar to cardiac muscle, but is to be seen, under appropriate conditions, in skeletal muscle, in nerve, and even in inorganic substances. In fig- 75 is given a series of responses of Galena to Hertzian Fig. 73. Reversed Re- sponse of Fatigued Nerve VARIOUS TYPES OF RESPONSE 103 radiation, which in its various phases of staircase^, uniform and fatig-ue-decline, is parallel to that just seen in muscle, 'rhe "phasic change, due to molecular transformation, which I have already pointed out under continuous stimulation, is seen in both these records in the shifting of the base-line. In fig. 64 a under continuous stimulation, we see the mechanical response of muscle passing from a condition of growing contraction into one of relaxation. In the record of individual- responses given in fig. 74, the same is seen to take place. A similar phenomenon is ob- • served in the mechanical re- sponse of Mimosa (fig. 65). When the mode of record, however, is electro-motive, in- m Fig. 74. Preliminary Staircase, followed by Fatigue, in the Responses of Muscle (Brodie) Fig. 75. Preliminary Staircase, In- crease, followed by Fatigue, in the Response of Galena to Hertzian Radiation (Resistivity variation method) stead of mechanical, the increasing galvanometric negativity which corresponds to increasing contraction, is found gradually to give place to positivity (fig. 64 U). And finally, when the mode of record is by resistivity-variation, we find, by the shifting of the-base line in fig. 75, that the residual negative variation of resistance at first waxes and then wanes. I: Instances have been given, in which a portion of the in- cident stimulus has been seen to be held latent to do internal work. And from this it is clear that the current assumption that response must always be larger than stimulus is quite un-^ 104 COMPARATIVE ELECTRO-PHYSIOLOGY tenable. There are cases, again, in which a large portion of the incident stimulus is held latent for a time, to find subsequent manifestation externally. This I have been able to demon- strate by the discovery of multiple response in plants. Thus while a single moderate stimulus in such cases evokes a single response, a single strong stimulus is found to give^ repeated or multiple responses. This I have shown7 not only in mechanical, but also in electrical response, and the latter subject will be taken up in detail in a subsequent chapter. And, lastly, it follows from what has been said, that incident stimulus need not always cause depreciation of the energy of the tissue, but that, on the contrary, it may actually raise it above par. I shall now describe an example in which incident stimulus was seen to find bifurcated expression. In fig. ^6 is given a photo- graphic record of contractile responses in the style of Datura alba^ in which growth had previously been in a state Fig. 76. Photographic Record of Responses of Style of Datura alba in which Growth had come to a Temporary Stop The up curve shows contraction. As long as the base-line is horizontal, growth is seen to be at standstill. Renewal of growth at sixth re- sponse, after which growth-elon- gation is shown by the trend of the base-line downwards. of standstill. The first five responses of this series are seen to be uniform. A portion of the stimulus applied must, however, from the first have been absorbed and held latent in the organ, thus increasing that internal energy, or tonic condition, on which growth depends. For at the sixth response we find that growth recommences, and the stimulus now finds bifurcated expression, in maintaining response and in renewing growth, as seen in the trend downwards of the VARIOUS TYPES OF RESPONSE I05 thitherto horizontal base-line. This bifurcation causes the first contractile response of the now growing organ — sixth of the series — to be smaller than usual. But, as a favourable tonic condition is gradually established by the absorption of energy and the molecular mobility of the responding organ is increased, the contractile response becomes larger, and growth goes on at a certain steady rate. This constitutes an instance in which stimulus, so far from lowering the energy of the responding system, has actually raised it I above par. It would thus appear that while the theory of assimilation and dissimilation is insufficient for the explanation of the various characteristics of response, the difficulties there en- ■^■countered are, on the contrary, satisfactorily explained, on taking full account of the influence on response of the molecular condition of the responding substance. From the chemical hypothesis of an explosive molecular change, with its attendant dissimilation and run-down of energy, it would follow that previous stimulation should always induce a depression of the subsequent responses. Instead of this, however, it is found that previous stimulation sometimes exalts, and at other times depresses, the subsequent re- sponses. This apparent anomaly we have seen to be ex- plained by the consideration of molecular transformation. From the sluggish condition A, we have seen tissues trans- formed, by the impact of moderate stimulus, to condition B, with its greater excitability. It is only when the molecular condition has been brought to D or E, that the responses undergo a diminution or reversal. The molecular condition, then, undergoes a continuous ransformation, in consequence of the action of stimulus, from the extreme of sub-tonicity A to the overstrained molecular conditions D and E. In the A stage, there is no true ex- citatory expression, response to stimulus being here by the abnormal positive variation. The substance is next trans- formed into stage B, where response exhibits a staircase character. In the next stage C, the respon.ses are uniform. I ■ I06 COMPARATIVE ELECTRO-PHYSIOLOGY Under over-stimulation, the stages D and E are reached, characterised by diminished amplitude of response, or actual reversal into positive. There are thus two conditions under which we obtain abnormal positive responses. One of these is that of sub-tonicity, and the other, the reversal due to fatigue. There is, again, no tissue which is exclusively characterised by any specific type of response. All these — staircase, uniform, and fatigue — will occur in muscle, nerve, plant, and even inorganic matter, under certain definite and ap- propriate conditions. In a future chapter, we shall study in detail the characteristic molecular curve, from which light will be thrown on the internal molecular condition of the tissue, and the influence of that condition on response. I CHAPTER IX (DETECTION OF PHYSIOLOGICAL ANISOTROPY BY ELECTRIC RESPONSE Anomalies in mechanical and electrical response — Resultant response determined by differential excitability — Responsive current from the more to the less excitable — Laws of response in anisotropic organ — Demonstration by means of mechanical stimulation — Vibrational stimulus — Stimu'ation by pressure — Quantitative stimulation by thermal shocks. It has been customary, as we know, to ascribe the varied movements of plant-organs under external stimulus, to the presence of different specific sensibilities ; and, indeed, it IBwouId seem at first sight impossible to reduce such highly complex and apparently unrelated phenomena, to the terms of a single fundamental reaction, common to all alike. There is no denying, for instance, that certain plant-organs, when a cted on by light, bend towards it, and others away . I have elsewhere shown, ^ however, that all these diverse movements are clearly traceable to one fundamental excitatory reaction, and that the different effects observed are due merely to the differential excitabilities of various parts of the structure ; and that the resultant movement is in all cases brought a bout by the great er contraction of the^rnore excited side^ 1^ Passing next to the electrical response of living tissues, « animal and vegetable, we encounter many anomalies. Not only will one tissue give positive, and another negative response, but we find also that the same tissue will give sometimes one and sometimes the other. These apparent inconsistencies are often due, as we shall find, to the differential excitability of anisotropic structures— a factor in the problem which has not hitherto been recognised. An investigation on ' Bose, Plant Response. I ■ I08 COMPARATIVE ELECTRO-PHYSIOLOGY this subject, then, demands that we first discover some means of determining the relative cxcitabilities of different parts of a tissue. ^ As the simplest example of an anisotropic structure, we may take a compound strip of ebonite and stretched india- rubber, glued firmly together throughout their length. Of these, the india-rubber is the more contractile, and when the strip as a whole is subjected to periodic thermal stimulation, response takes place by the greater contraction induced in the india-rubber. Ifthe strip be held, with the india-rubber below, response will be by the induced concavity of the (lower side. In fig. 77 is shown a series of these responses of the compound strip, taken on a smoked surface by means of a recording lever. In anisotropic motile organs, such as the pulvinus of Mimosa, response takes place by differential contraction, the more excitable side being that which under diffuse stimulation becomes concave. If we apply very moderate stimulus ^'"cJntracmf '^^Re! ^^^^^^^ °" ^^^ "PP^^ ^^^^ ^^ ^^^ pulvinus, sponse of Artificial we shall find that, by the excitatory con- "^ traction of this half, the leaf is raised. A similar contractile effect, though of greater intensity, is induced when the lower half of the pulvinus is stimulated locally, the leaf in this case undergoing a depression. When both upper and lower halves, then, are excited simultaneously, the resulting fall of the leaf shows that the contraction of the lower half must in this case be the greater, or, in other woids, that this half is the more excitable of the two. This experi- ment may be carried out very easily by using the stimulus of light. Fig. 78 gives the results observed {a), showing the up movement consequent on stimulation of the upper half; {b) that caused by equal stimulation of the lower half ; and {c) the resultant fall when the two are excited simultaneously. In the case of mechanical response, then, we find it true that response is by the greater contraction of the more excitable. DETECTION OF PHYSIOLOGICAL ANISOTROPY 1 09 I We shall next observe what is the electrical mode of response for a tissue which is anisotropic, or unequally ex- citable on two sides. For this purpose we may again take the pulvinus of Mimosa, and make electrical connections at two diametrically opposite points on the upper and lower halves of the pulvinus respectively. It is to be remembered that electrical response takes place on excitation, whether the leaf be free to move, or physically restrained. We may, herefore, hold it in a fixed position ; and indeed this is advisable, in order to avoid that shifting of the electrical contacts which might 1)ossibly take place if t were allowed to fall. The two contacts ire made with two ine straws filled with Kaolin paste, moistened in normal saline. On l^piow applying a series of thermal stirnuli, on the petiole, near the pulvinus, I obtained the responses given in fig. 79. It will be seen that the responsive current flows in the tissue from the rela- tively more excited lower, to the less excited upper, half of the organ. IK We thus arrive at a comprehensive law of the mechanical '"^and electrical response of anisotropic organs : Diffuse stimulation induces greater contraction and galvanometric negativity of the more excitable side. The laws of electric response in the anisotropic organ may then be detailed as follows : — 1. On simultaneous excitation of two points, A and B, the responsive current flows in the tissue from the more to the less excited. Fig. 78. Responses of Mimosa to Sunlight of not too long Duration {a) Light acting on pulvinus from above ; {h) light acting on pulvinus from below ; (f ) light acting simultaneously from above and below. Dotted line represents recovery on cessation of light. no COMPARATIVE ELECTRO-PHYSIOLOGY 2. Conversely, if under simultaneous excitation the responsive current be from B to A, then B is the more excitable of these two points. These form only an instance of the general law that the responsive current always flows from the more to the less excited. For when a point, B, is excited locally — this point, that is to say, being the more excited — the responsive current is found to flow away from it to a neutral or in- different point, A, for which any distant point will serve, provided the tissue be non-conducting. Should it be con- ducting, the neutrality of A is maintained by interposing a Fig. 79. Transverse Response of Pulvinus of Mimosa The petiole is securely held to prevent movement, and diametric electric contacts made in the upper and lower surfaces of pulvinus. Re- sponsive current is from lower to upper surface. block. Should the stimulus, however, not be local, but diffuse, a resultant response may still be obtained by injuring or killing the point A, and thus diminishing or abolishing its excitability. On stimulation, the point B is now necessarily the more excited, and the responsive current is still away from B, towards A. And finally, owing to physiological anisotropy, B may be naturally more excitable than A, and on stimulation the responsive current will then be found to flow from the more excited B to the less excited A. The comparison of the excitabilities of the two points A and B, therefore, reduces itself to the application of similar DETECTION OF PHYSIOLOGICAL ANISOTROPY II I stimuli to the two points simultaneously, and then ascertaining the direction of the responsive current. For this purpose we might employ any form of stimulus, and it is extremely interesting to find that, however diverse the stimuli, the results obtained by them are always identical. And here we have not merely a means of qualitative demonstration, but in some cases one of quanti- tative also. If we take an erect stem of Cucurbita^ it being radial and isotropic, all its flanks will be found equally excitable. Hence, if two diametrically opposite contacts are made, there Fig. 8o. Diametric Method of Stimulation of an Anisotropic Organ Diametrically opposite contacts are made at a and B, and tissue subjected to vibrational stimulus. will, on diffuse stimulation, be no resultant response. But when such a stem becomes recumbent, the upper side, being now constantly exposed to light, becomes fatigued by over- stimulation, with consequent diminution of its excitability. This is true only when the stimulus has been excessive and long contiaued ; for we have seen moderate stimulus may sometimes enhance the excitability. By the unilateral action of light, then, the organ has been converted from radial into anisotropic, the lower side being that which we shall expect to find the more excitable. On mounting such a stem in the vibratory apparatus (fig. 80), and making diametrically opposite contacts on the two anisotropic surfaces, we find that on applying vibration 112 COMPARATIVE ELECTRO- PHYSIOLOGY both sides are subjected to similar stimulus simultaneously ; and the responsive current is now found to flow across the tissue, from the lower to the upper side. The lower is thus, as we expected, the more excitable. Since we can by means of vibration apply measured stimuli, it will be seen that we have here a quantitative method of investigation. Moreover, as the stimulus is applied directly, it is applicable not only to conducting but also to non-conducting tissues. If we next take a radial stem or petiole of Cucurbita, and slit it longitudinally, we obtain, in either of the halves, a specimen having an inner and an outer surface. As one of these has been exposed to light and the other protected from it, we should expect to find, on examination, that there has been an induction of physiological anisotropy. As such a specimen is not very well adapted for vibrational stimula- tion, we may use that of pressure. Two moistened rags, in connection with non-polarisable electrodes, pass through two pieces of cork, adjusted on the two surfaces — outer and inner — at diametrically opposite points. When the inter- posed tissue is now subjected to sudden pressure its two surfaces are excited simultaneously, and the responsive current is found to flow from the inner concave to the outer convex surface, proving that the former was the more excitable. We might again use the chemical form of stimulation, and the results obtained by this method will be described in the course of the next chapter. But these forms of stimulus — by pressure, or by chemical means — are not capable of exact measurement. For quantitative observations, then, it is necessary to employ some other form of stimulus, and the electrical offers us in this respect many advantages. There are, however, in this case many possible disturbing influences to be considered, all of which must be carefully eliminated before the method can be used without misgiving. How this may be done will be shown in a future chapter. For the present I shall describe another method of stimulation which I have been able to bring to great perfection, by which DETECTION OF PHYSIOLOGICAL ANISOTROPY 113 two points of an anisotropic organ may be simultaneously- excited, under a series of stimuli of uniform or increasing intensity. This mode of excitation, by thermal shocks, will be found in every way satisfactory and convenient. The Thermal Variator, by which stimulation is effected, consists of a spiral of german-silver wire, the diameter of the spiral being about 3 cm. The electrical circuit, through which the heating- (Current is sent, is closed periodically for a definite length of Fig. 81. The Thermal Variator The anisotropic tissue-petiole of Musa is held in ebonite clip, c. e, e', electrodes connected with opposite sides. Specimen after adjustment pushed inside heating-spiral, T, by slide, s. Spiral heated periodically by closure of electric circuit by metronome, M. »' I ime, by means of a metronome (fig. 81). The thermal variation within the coil can be controlled by a suitable adjustment of the battery-power, or by the duration of losure, or both. The experimental tissue is held in an ebonite clip, C, fixed on a slide, S, on the same stand as the heating-spiral. This slide is pulled out for the purpose of adjustment. Square or circular pieces of wetted muslin make contacts with equal areas on two opposite sides of the experimental tissue, these pieces of cloth being connected with non-polarisable electrodes, E and E)' After the adjust- I 114 COMPARATIVE ELECTRO-PHYSIOLOGY ment is made, the slide is pushed in, till the tissue is well in the centre of the coil. When the circuit is completed, for a brief period, both the sides A and B are subjected to the same sudden variation of temperature, which, as we know, acts as a stimulus. As the two contacts are thus in practice raised to the same temperature, there will be no thermo-electrical disturbance. The responsive current, therefore, will be determined by any difference of excitability which may exist as between A and B. The spiral also gives out heat-radia- tion, which acts as a contributory stimulus. That it is the thermal variation, and not the temperature, which acts as the efficient external stimulus, is seen from the fact that when the tissue is subjected to the higher temperature con- tinuously, the galvanometric deflection obtained is opposite in direction to that induced by tlie thermal shock. This is because the absorption of heat, as such, increases the internal energy, and thus induces an electrical effect opposite to that caused by external stimulation. As experimental tissue, we may use the sheathing petiole of Musa. The required piece is cut and mounted in the apparatus, the concave surface being taken, say, as B, and the convex as A. I have mentioned Musa as suitable for this purpose, because I find it, when fresh, to show practically no sign of fatigue in its responses. There are many other sheathing petioles, which would doubtless answer the same purpose more or less perfectly. In obtaining records with this .specimen, it is found that the responsive current flows across the petiole, from the inner concave surface B to the outer convex surface A, showing that it is the inside which is more excitable. Uniform stimuli of short duration were applied at intervals of one minute, and the responses obtained are seen to be fairly uniform (fig. 82). The specimen was next subjected to the anaesthetic action of chloroform. This, it will be seen, in- duced a very great depression of the response. It has thus been shown that just as the greater contrac- tion and concavity of a motile organ enables us to discrimi- DETECTION OF PHYSIOLOGICAL ANISOTROPY II 5 nate which side of two is the more excitable, so here also the more excitable side is that which, on diffuse stimulation, exhibits galvanometric negativity relatively to the other. From this it becomes possible to determine the relative excitabilities of any anisotropic organ, even though it be non-motile, and therefore incapable of exhibiting any con- spicuous mechanical response. The difficulty of applying equal and quantitative stimulus on two sides simultaneously Fk;. 82. Responsive Current in Petiole of Musa from Concave to Convex Side First series, normal ; after application of chloroform subsequent depression. has now been overcome by vibrational stimulation, and by the perfection of the method of thermal shocks. Thus a definite resultant response has been shown to be determined by the differential excitabilities of two parts of an experi- mental tissue. And that from this consideration it becomes further possible to resolve many of the remaining anomalies of electrical response will be fully demonstrated in a sub- sequent chapter. I 2 CHAPTER X THE NATURAL CURRENT AND ITS VARIATIONS Natural current in anisotropic organ from the less to the more excitable — External stimulus induces responsive current in opposite direction — Increase of internal energy induces positive, and decrease negative, variation of natural current — Effect on natural current of variation of temperature — Effect of sudden variation — Variation of natural current by chemical agents, referred to physiological reaction — Agents which render tissue excitable, induce the positive, and those which cause excitation, the negative variation — Action of hydrochloric acid— Action of Na./JOg — Effect modified by strength of dose — Effect of CO2 and of alcohol vapour — Natural current and its variations — Extreme unreliability of negative variation so-called as test of excitatory reaction— Reversal of natural current by excessive cold or by stimulation — Reversal of normal response under sub-tonicity or fatigue. We have seen that when the pulvinus of Mimosa is excited by an external stimulus, there is a relatively greater expulsion of water from the more excitable lower half, with a con- comitant greater contraction. Conversely, the lower half of the pulvinus is capable of absorbing more water, and of expanding to a greater extent, than the upper. Increased internal energy, in contrast to the action of external stimulus, has the effect of causing a greater expansion of the lower half of the pulvinus, and thus raising the leaf. This we saw exemplified when the plant was subjected to a gradually rising temperature, so as to increase its internal energy, its leaves being thereby made to show increased erection (p. ^2), Hence the more excitable tissue in the pulvinus of Mimosa is characterised, both by greater power of absorption and by greater emission of energy, according to circumstances. In this we see a close analogy to the action of inorganic bodies, in which also we find the greatest power of emission to be associated with a correspondingly great power of absorption of energy. 11 THE NATURAL CURRENT AND ITS VARIATIONS II7 We have thus seen that in order to maintain a high state of excitability, absorption of energy is necessary. On excitation, emission of energy occurs. In this latter case, of emission, we observe a concomitant galvanometric negativity of the more excited lower side. Since to have maintained its excitability the opposite process of absorption would have been necessary, it follows that the more excitable lower side must under normal conditions be galvanometrically positive. This is found to be the case. For when the leaf is in an excitable condition, there is an electro-motive difference between the upper and lower halves of the pulvinus, in con- sequence of which a current flows across the tissue, from the less excitable upper to the more excitable lower half, which is thus galvanometrically positive, in relation to the upper. ,''"e have here, then, an additional instance of the opposite effects of internal energy and external stimulus. Internal energy, maintaining a greater excitability of the lower half of the pulvinus, induces in it a relative galvanometric positivity. External stimulus, on the other hand, gives rise to precisely the opposite effect — namely, the relative galvanometric negativity of the lower half. Under typical conditions, then, we may expect the more excitable point to be galvano- metrically positive ; and the more excited to be galvano- metrically negative. Turning next to non-motile tissues, we find the same conclusions to hold good. We saw that in the case of the sheathing petiole of Musa^ the concave was more excitable than the convex side. The concave is thus normally positive to the convex side, and the natural current flows across the tissue from the convex to the concave. While the natural current flows from the more excitable to the less excitable, external stimulus gives rise to a responsive current in the opposite direction, from the more excited to the less excited, constituting a negative variation of the current of rest. Let us next consider what would be the effect of an increase of internal energy on the natural current. Since the action of internal energy is opposite to that of external stimulus, we should expect it to induce a positive variation of i8 COMPARATIVE ELECTRO-PHYSIOLOGY the natural current. Diminution of internal energy on the other hand might be expected to cause a negative variation. These effects are diagrammatically represented in fig. 83, which also exhibits the parallelism between the electric responses of motile pulvinus and non-motile anisotropic organ. We next proceed to subject the question of the effect of increased or diminished internal energy on the natural current to experimental verification. As regards the increase of internal energy, we have already seen that this can be secured by a gradually rising temperature, its diminution being, con- trariwise, secured by a falling temperature. In the case of Mimosa^ we • saw that the former in- duced an erection of the leaves, and the latter a gradual depression. In order, then, to observe the effect of increase or de- crease of internal energy on the existing current of rest, we have only to sub- ject the specimen to gradual thermal ascent or descent, and record the consequent variation of current. The specimen of Musa is placed in a chamber and two diametrically opposite contacts are made, with the internal and external surfaces, and led off to the galvanometer. To take first the effect of cooling : a stream of ice-cold water is sent through a hollow tube in the chamber : this gradually lowers the temperature, say from 30° C. to 27° C. It will be seen from left-hand curve of fig. 84 that this has the effect of diminishing the natural current of rest in the tissue, as represented by the dotted arrow ;. When the chamber is Fig. 83. Parallelism of Natural Current in Pulvinus of Mimosa and Sheathing Petiole of Musa Upper and less excitable surface of former corresponds with outer or convex sur- face of latter. The natural current, N, is in both from the less to the more ex- citable. In both excitatory current, e, is in opposite direction, i.e. from the more to the less excitable. In Musa increase of internal energy ( + i ) in- duces a positive, and diminution of internal energy ( — t ) a negative, varia- tion of the natural current. THE NATURAL CURRENT AND ITS VARIATIONS II9 te IRfJ allowed to return to the temperature of the room, this diminution of current is annulled. To study the effect, on the other hand, of a rising tempera- ture, the chamber is gradually heated, by means of the electric heating-coil already described. In thus raising the temperature, it is found (fig. 84, right hand curve) that the natural resting-current undergoes an increase. On cooling gain to the original temperature, this in- crease is annulled. That these effects are ue to induced elec- tro-motive variations, and not to any changes of resistance, is demonstrated from the fact that the effect Bflescribed takes place ^Reven when the original " E. M. F. is exactly balanced by a poten- fttiometer. ^m It was specially ■^tated that these ob- ^fcervations with regard to the effect of varying temperatures apply only to steady varia- tions. In the case of » thermal ascent, we iiave seen that a steady rise brings about an increase of the existing current. But since sudden variation of tempera- ture acts as a stimulus, we shall, in the preliminary stage, obtain an excitatory reaction, which will cause a transient diminution of the current of rest. This will be followed, when the rise of temperature is steady, by an increase of ^he current of rest. I give here (fig. 85) a photographic Fig. 84. Effect ot Variation of Temperature on Natural Current, |, which in Petiole of Musa flows from Convex to Concave Side Effect of cooling from 30° C. to 27° C, seen on left, induces negative variation of natural current. Restoration to original value on return to surrounding temperature. Warm- ing induces positive variation (see record to right). In this and subsequent figures in the present chapter 4' indicates the direction of the natural current of rest. I 120 COMPARATIVE ELECTRO-PHYSIOLOGY \ ( record of these contrasted effects. In the first part of the curve we observe a sudden movement of the record upwards corresponding to the sudden rise of temperature. This is so great as to carry the curve out of the photographic field. We have here, then, a sudden excitatory diminution of the natural current. In the next stage, while the temperature is steadily ascending, we find a reversal of the curve, and the natural current is enhanced above the normal. On now allowing the chamber to cool down to the original temperature of the room, the natural current was found to return more or less to its normal value. We shall next study the effect of chemical agents on the natural current. The mode of procedure is to apply the given agent on both the contacts at the same time. If the substance be liquid, it can be applied by a pipette. If it be gaseous, the specimen is placed in a chamber through which the gas or vapour is allowed to stream. In observing the effects of various agents we obtain results which are at first sight very perplexing. For example, certain substances will be found to induce a diminution of the natural current, and others an increase. The effect, moreover, is found to be modi- fied by the strength of the dose. Thus an agent which, in a given strength, will cause a diminution of the natural current, may often be found to cause an increase, when sufficiently diluted. This inquiry is of great importance, since it is directly connected with many equally obscure problems in medical practice, where the effect of a drug Fig. 85. Photographic Record showing effect of Sudden, followed by steady Rise of Tem- perature on Natural Cur- rent, >;, in Alusa During sudden variation of temperature an excitatory negative variation of natural current takes place, as shown by first up curve ; when rise of temperature becomes steady there is a positive variation, as shown by the down curve ; on re- turn to surrounding tem- perature, the normal cur- rent is restored to its original value. THE NATURAL CURRENT AND ITS VARIATIONS 121 is well known to be modified by the amount of the dose. Much light appears to be thrown on this subject, when we consider the electrical reactions of the chemical agents as due to their physiological action. If a drop of hydrochloric acid be applied to the pulvinus of Mimosa^ the leaf falls, showing that the more excitable side has undergone a greater iBexcitatory contraction. We have also seen that when a drop of this acid is applied on any tissue in the neighbourhood of, ,na but not directly touching, an electrical contact, it induces an IJpexcitatory galvanometric negativity. If now we apply it in solution, say, of lo per cent, on the two diametrically opposite i contacts of Musa, we shall expect that the greater excitatory reaction induced on the concave side will give rise on that |ide to a relative galvanometric negativity, resulting in a negative variation of current of rest. On the application of - — the reagent this is found to be the case, the responsive Ufcurrent flowing in a direction opposite to that of rest : that is to say, it flows from the more excited concave to the less excited convex. It is by considering chemical agents from the point of ll^iew of their physiological reaction, that we are able to " explain their diversity of effects, according to the strength of I the dose and the duration of application. We have seen that HR^hile a strong stimulus induces the excitatory effect of negativity, a feeble stimulus will bring about the opposite, or positivity. This abnormal positive response, however, by the continued action of moderately feeble stimulus becomes converted into normal negative. Now a chemical substance which in a certain strength acts as an efficient excitatory agent, may, when sufficiently diluted, act as a feeble stimulus, inducing a positive response. If the same agent again were applied in a slightly greater concentration, its immediate effect might be positive, to be succeeded under continued application by the normal negative. tWe might thus expect, using a strong solution of a given 122 COMPARATIVE ELECTRO-f HYSlOLOGY of rest ; using a dilute solution, to obtain a positive varia- tion ; and, lastly, applying a dose of intermediate strength, to discover the very interesting case in which the reagent would give rise immediately to a positive variation, and after a longer or shorter continuance of its action, to a reversed, or negative variation, of the current of rest. These inductions are found fully verified in the experiment which I am now about to describe. Fig. 86. Action of 7 per cent. Solution of NajCOa on Natural Current of Mttsa Preliminary positive variation represented by down curve followed by reversal, 50 seconds after application. Fig. 87. Effect of CO, on Natural Current of Miisa Preliminary positive seen to be succeeded by negative variation 5 minutes after application. Applying a strong solution of sodium carbonate — 10 per cent, or above — at the electrical contacts on Musa, the result is a negative variation of the natural current. If now a dilute solution of I per cent, be applied on a similar specimen, we obtain a response by positive variation. And if, lastly, we use a 7 per cent, solution, we obtain, as will be seen from the record (fig. 86) the preliminary positive, succeeded, under the continued action of the agent, by reversal to the negative, variation. We pass next to the question of the effect of gases. In fig. 87 is given a record of the action of carbonic acid on THE NATURAL CURRENT AND ITS VARIATIONS 1 23 the natural current in Musa. It will be seen here that in the first stage there was an enhancement, or positive variation, of the existing current. In a later stage, however, this is followed by a reversal, the resting-current now undergoing a diminution. We have here an effect parallel to that of the intermediate dose of sodium carbonate. Vapour of alcohol I also exerts an effect very similar ; that is to say, it induces a preliminary exaltation, followed by a depression of the natural current. In connection with this subject, of the changes induced in the natural electro-motive difference between the two surfaces, by the action of a chemical reagent, it is well to distinguish between the effects of two different factors : namely, the electrical variation caused by the chemical sub- I stance as such, and that brought about by the excitatory reaction. Let us suppose both the electrical contacts to be tmade on iso-electrical surfaces, with normal saline solution ; there will then be no difference of potential, as between the two. But this state of things will be disturbed, by the appli- cation of another chemical solution, say acid, on either one of the two contacts. The resulting disturbance may be dis- Itinguished as due to heterogeneity of chemical application. But if the same chemical agent be applied at both the , contacts, no such chemical heterogeneity will ensue. If, then, any electro-motive difference be induced, it must be due primarily to some induced physiological change. The contact which has been rendered more excitable will become increas- ingly positive ; that which is more excited, on the other hand, will become increasingly negative. That the induced electro- motive variation under such circumstances is indicative of a variation of excitability or excitation, was seen in the fact that the same chemical agent — for example, Na2C03— caused I a positive variation when dilute, a negative when strong, and positive followed by negative under the continued action of an intermediate dose. This conclusion — that the variation of the existing current, by the simultaneous application --—--— — 124 COMPARATIVE ELECTRO-PHYSIOLOGY to a physiological reaction, the positive variation being a sign of relatively increased, and the negative of decreased, excitability — will be verified by an independent mode of in- quiry, to be described in the following chapter. It follows from the experimental demonstration which has just been given that the phenomena of the natural current and its variations may be summarised in general as follows : 1. Under normal conditions, the current of rest flows in the tissue from the less to the more excitable. In other words, the more excitable is galvanometrically positive to the less excitable. 2. Increase of internal energy induces an increase or positive variation of the existing current ; and diminution of internal energy induces a negative variation. 3. External stimulus induces a negative variation of the true or natural current of rest. The natural current and its variations under normal con- ditions have now been studied. We shall next proceed to trace out those conditions under which abnormal results may occur. Excessive cooling, by diminishing internal energy, may thus reverse the normal current, which reversal may become more or less persistent. It has been shown, moreover, that in an anisotropic organ, external stimulus gives rise to a current opposite in direction to the natural current. By this excita- tory reaction. the more excitable side, hitherto positive, is rendered negative, and if the excitatory reaction be great, it may remain for a considerable period in this reversed condition of galvanometric negativity. We have seen that under normal conditions, the direction of the natural or true current of rest is from the less to the more excitable, and that external stimulus causes a responsive current in the opposite direction, which thus constitutes a negative variation of the current of rest. This state of things we shall distinguish as the primary condition. It frequently happens, however, in consequence of previous stimulation, with its after-effect, that extremely varied effects, appearing at first very anomalous, occur, with regard to the I THE NATURAL CURRENT AND ITS VARIATIONS 125 direction, not only of the current of rest, but also of the current of response. We shall be able to obtain a clear understanding of these effects, on subjecting the underlying phenomena to close analysis. As a concrete example, we may take for our investigation the various effects to be observed in the pulvinus of Mimosa. In this anisotropic organ, the directions of the resultant current of rest and the current of response are determined, as we have seen, by the differential excitability and differential excitation of the two sides of the organ. We shall fix our attention, however, for the sake of simplicity, on the changes which occur in the more effective lower half. I shall here succinctly describe the various post-primary phases in the changing effect, culminating in the onset of fatigue from over- stimulation. In the primary condition, as we have seen, the lower half of the pulvinus is positive to the upper, the direction of the resting-current being down i. On stimula- tion, the lower half becomes negative and the response current is up t. Response thus takes place by a negative variation of the resting-current. We may now suppose the pulvinus to be in a state of slight excitation, its molecular condition at or about the B stage. This existing state of moderate excitation will annul the previous positivity, and the current of rest will now be zero. At this stage, however, as we have seen, the excit- ability of a tissue is enhanced. Hence, on stimulation, the lower half of the pulvinus will exhibit responsive negativity, and the direction of the response current will be up t. We . are, however, unable to describe this variation in terms of the current of rest, since that, as we have already seen, is zero. A condition of still stronger excitation, bringing the tissue to the C condition, will induce galvanometric negativity of the lower half. The so-called current of rest is thus upwards t. But as the excitability of the lower half is still relatively greater than that of the upper, it follows that external stimulus will bring about a responsive current whose direction is upwards t, This normal excitatory response, 126 COMPARATIVE ELECTRO-PHYSIOLOGY then, in this particular case, appears as a positive variation of the resting-current. And, finally, we may imagine the pulvinus to have been strongly excited, so as to be in the condition D or E. The resting-current will in this case be upwards t. But the ex- citability of the lower half, owing to fatigue, has now become depressed, a condition which, as we have seen, tends to give rise to the abnormal positive response, the responsive current being thus downwards i. As the modified current of rest is upwards, this abnormal current of response will appear as a negative variation of it. All these cases are conveniently tabulated as follows : Tabular Statement of the Relative Directions of the Current OF Rest and the Responsive Current under Various Conditions. Condition. Current of rest. Current of re- sponse. Variation of current of rest Primary condition Feebly excited . Moderately excited Strongly excited and fatigued . i O t t t t t Negative variation Positive variation Negative variation The various conditions mentioned may be induced accidentally in the responding organ, or may be brought about by the excitatory effect of experimental prepara- tion. 1 give here the records of certain experiments performed on Mmiosa, in which some of these changes were seen to occur as the result of stimulation (fig. 88). The normal natural current is seen to be from above to below, as represented by the dotted arrow. The first strong stimulus, applied at the moment represented by the thick dot, gives rise to a responsive current whose direction is from below to above. Owing to the strong intensity of the stimulation, there is here a slight indication of multiple response. As an after-effect of stimulus, we observe that the normal resting- current has undergone a reversal, the lower surface, which was formerly positive, having now become relatively negative. A second stimulus now gave rise to a response THE NATURAL CURRENT AND ITS VARIATIONS 12/ similar to the first. But after this, owing to the greater fatigue with loss of excitability induced in the lower half of the pulvinus, the succeeding responses are seen to be reversed, the responsive current being henceforth from above to below. From the table given on the previous page, it will be seen that hardly could any standard have been devised for the study of excitatory reaction, so likely to be prolific of confusion as this, of the so called variation of the resting- current. For in the first three cases displayed, we l^ftsee one identical excitatory effect, appearing now as a negative, again as doubtful, and a third time as a positive variation of the current of rest. In the fourth case, again, it is actually the l^feibnormal response which appears as the normal nega- Itive variation ! But while these responses appear to be so various, the underlying reaction is nevertheless con- stant. The direction of the l^kresponsive current is always r from the more to the less Fig. 88. Variation of the Transverse Natural and Responsive Currents in Pulvinus of Mimosa Natural current ^ which is normally down, reversed in consequence of strong external stimulus. The first two responses are normal, i.e. current being from below to above. Strong stimulus is here seen to induce mul- tiple responses. After the second response on account of the greater fatigue induced in the lower half of the pulvinus, the direction of the responsive current is seen to be reversed. Thick dots represent moment of application of stimulus. : excited. IB It has thus been shown in the course of the present chapter that under normal I conditions the current of rest flows in the tissue from the less to the more excitable ; that increase of internal energy causes a positive variation of the current of rest ; while its diminution gives rise to a negative variation ; that reagents which increase excitabilit}^ induce a positive, and [■those which cause excitation a negative, variation of the IBresting-current ; and, finally, that external stimulus induces 128 COMPARATIVE ELECTRO PHYSIOLOGY the normal reactions, however, under abnormal conditions, they may be reversed. Thus, excessive cooling or strong external stimulation may reverse the normal current of rest. There are, moreover, two different conditions, those, namely, of sub-tonicity {cf. p. io6) and fatigue, which may be effective in bringing about a reversal of the normal direction of the responsive current. In this way, by means of induced varia- tions of the resting-current and of the responsive current, many very varied effects become possible. J I CHAPTER XI VARIATIONS OF EXCITABILITY UNDER CHEMICAL REAGENTS Induced variation of excitability studied by two methods : (i) direct (2) trans- mitted stimulation — Effect of chloroform — Effect of chloral — Effect of formalin — Advantage of the Method of Block over that of negative variation— Effect of KHO — Response unaffected by variation of resistance — Stimulating action of solution of sugar — Of sodium carbonate— Effect of doses — Effect of hydro- chloric acid— Di-phasic response on application of potash — Conversion of normal negative into abnormal positive response by abolition of true excitability. tT has been said in a previous chapter that the electrical esponse is a true physiological response. This is demon- strated by the fact that, while a vigorous specimen gives strong electrical response of galvanometric negativity, the same specimen, when killed, whether by heat or by poison, ceases to respond. This particular electrical response is thus seen to be a concomitant of physiological efficiency. It follows that, whatever diminishes physiological activity "^XW, pari passu, modify the amplitude of the response. But in cases in which the death of the tissue is brought about by steam or by poison, it is the last stage only, namely the abolition of response, that can be observed. It is also impor- IB|ant, however, to be able to trace the growth of physiological changes through the concomitant modification of response. In this way it is possible not only to study the gradual onset of death, as induced by a poison, but also the action of other chemical agents, some of which might be of such a nature as to induce exaltation, others depression, and still others^ like the narcotics, a temporary aboHtioi) of the electrical response, I30 COMPARATIVE ELECTRO-PHYSIOLOGY An essential condition of this investigation is first to obtain a uniform series of responses. Having once done this, those subsequent changes in the response which are due to the appli- cation of a given reagent can be demonstrated in an unmis- takable manner. I have already explained in Chapter III. that this may be done by either of two different methods : namely, those of direct and of transmitted stimulation. In the first of these we employ vibrational stimulus, using the Method Before | After Fig. 89. Photographic Record of EflFect of Chloroform on Responses of Carrot Stimuli of 25° torsional vibration at intervals of one minute. of Block. In the second, the stimulus of thermal shocks is used, the excitation of the proximal contact being due to transmitted stimulation. We shall investigate the effect of chemical reagents by both these methods. And first I shall give results obtained by the employment of the Method of Block, the tissue being subject to direct stimulation. In cases where the effect of gaseous reagents, like chloroform, is to be studied, the vapour is blown into the plant-chamber (see fig. 21). In EXCITABILITY UNDER CHEMICAL REAGENTS I3I cases of liquid reagents, they are applied on the points of contact A and B, and in their close neighbourhood. The experiment is carried out by first obtaining a series of normal responses to uniform stimuli, applied at regular intervals of time, say one minute, the record being taken the while on a photographic plate. Then, without interrupting this procedure, the given agent — say, vapour of chloroform — is applied, by being blown into the chamber. It will be seen from fig. 84 how rapidly chloroform induces depression of response, and how the effect grows with time. If the speci- men be subjected for a short time only to the anaesthetic, the depressing action proves transient, passing off on the reintroduction of fresh air. But too strong or too pro- longed an application induces a permanent abolition of Response. ■ I give below (figs. 90, 91), two sets of records, one of which shows the effect of chloral and the other formalin. It .lUU^ \\ \ ^ ^ I Before ^ After Fig. 90. Photographic Record showing Action of Chloral Hydrate on the Responses of Leaf- stalk of Cauliflower Torsional vibration of 25° at intervals of one minute. 'hese reagents were applied as solutions on the tissue at le two leading contacts and adjacent surfaces. Both re seen to induce a rapid decline of the response. In the I K 2 132 COMPARATIVE ELECTRO- PHYSIOLOGY normal responses, shown in fig. 91, is seen a very interesting instance of alternating fatigue. In order to bring out clearly the main phenomena, I have postponed till now the consideration of a point of some difficulty. To determine the influence of a reagent in modifying the excitability of a tissue, we rely upon its effect in exalting or depressing the responsive E.M. Variation, and we read this effect by means of changes induced in the galvanometric deflection. Now as long as the resistance of the circuit remains constant, an increase or decrease of galvanometric deflection will accurately indicate a heightened or depressed E.M. Variation, due to augmented or lowered .\\\\\s.VV.W^ Before f After Fig. 91. Photographic Record showing Action of Formalin (Radish) excitability, induced by the reagent in the tissue. But by the introduction of the chemical reagent the resistance of the tissue may undergo a change, and, owing to this cause, modification of response, as read by the galvanometer, may be induced without any E.M. Variation. The observed variation of response may thus be partly owing to some unknown change of resistance, as well as to that of the E.M. Variation. This difficulty may, however, be obviated by interposing a very large and constant resistance in the external circuit. The variation in the tissue then becomes negligible, the galvanometric deflections being now proportional to the electro- motive variation. An actual experiment will make EXCITABILITY UNDER CHEMICAL REAGENTS 1 33 this point clear. Taking a carrot as a specimen, I found its resistance //?/i' the resistance of the non-polarisable electrodes to be 20,000 ohms. The application of a chemical reagent reduced this to 19,000 ohms. The resistance of the galva- nometer used was 1,000 ohms, and the high constant external resistance interposed was i million ohms. The variation of resistance induced in the circuit by the application of the reagent was thus 1,000 in 1,020,000, or less than one part in a thousand. In studying the variation of excitability in animal tissues, the method of negative variation is employed. But I may here draw attention to the advantage which is afforded by the employment of the Method of Block instead. For, l^kin the method of negative variation, one contact being injured, the chemical reagents act on injured and uninjured (unequally. It thus happens that by this unequal action the resting difference of potential is indefinitely altered. But the intensity of response in this method of injury may to a certain extent be dependent on the resting difference. It is thus seen that, when this method is employed, a factor is Introduced which may give rise to complications. According to the Block Method, however, the two contacts ire made with uninjured surfaces, and the effect of the eagents on both is similar. Thus no advantage is given to cither contact over the other. The changes now detected in the response are therefore due to no adventitious circum- stance, but to the reagent itself. If further proof be desired |^k>f the effect ascribed to the action of the reagent, we can now obtain it by the alternate stimulation of the two ends A and B. I give below (fig. 92) a record of responses btained in this way from the petiole of turnip. This petiole as somewhat conical in form, and owing to this difference tween the A and B ends, the responses given by one were lightly smaller than those given by the other, though the timuli were equal in the two cases. A few drops of a o per cent, solution of NaOH were applied at both ends. he record shows how quickly this reagent abolished the 134 COMPARATIVE ELECTRO-PHYSIOLOGY response of both. In the next figure (fig. 93) is given a photo- graphic record, showing the marked depression of response induced by a strong solution of KOH, and in order to show that under the given experimental conditions, the variation of resistance does not in any way affect the responses, the deflection produced in the galvanometer by the application of an E.M.F. of -i volt to the circuit is shown at the beginning and end of the record. The equality of these two deflections shows that the resistance in the circuit has remained practically the same throughout the experiment. Before \ After Fig. 92. Abolition of Response at both A and B Ends by the Action of NaOH Stimuli of 30° vibration were applied at intervals of one minute to A and B alternately. Response was completely abolished twenty-four minutes after application of NaOH. Therefore, the change in the amplitude of the E. M. responses recorded may be taken as due entirely to the variation in the excitability of the tissue. In the experiments just described, the stimulus was applied directly at the responding point. By the application of a chemical reagent, not only was the responsive excitability of the tissue modified, but its receptivity, or power of receiving stimulus, also underwent a change. It will be shown later that the receptive excitability and the responsive excitability are not necessarily the same. The records which have just been given show what is, strictly speaking, the EXCITABILITY UNDER CHEMICAL REAGENTS 1 35 effect of the reagent on both receptivity and responsivity jointly. If, however, we wish to study the effect of the reagent on responsive excitability alone, it will be necessary to separate the receptive from the responding point, and apply the reagent on the latter. This may be done by the method of trans- mitted stimulation described previously. Successive uniform I Before ^ After Fig. 93. Photographic Record showing the nearly complete Abolition of Response by strong KOH The two vertical lines are galvanometer deflections due to •! volt, before and after the application of reagent. It will be noticed that the total resistance remains unchanged. t Stimuli applied at a given point cause excitatory response at the separate responding point, the record of which is taken ; after this, the chemical reagent is applied locally at the responding point. It will be seen that the receptive excitability and the conductivity of the intervening tissue ^^pemain unaffected, changes being induced at the responding ^^■area alone. 136 COMPARATIVE ELECTRO-PHYSIOLOGY The specimen employed was the petiole of fern. The thermal stimulator was at a distance of 1-5 cm. from the proximal electrode. In fig. 94 is shown the stimulating action of a >A- y Fig. 94. Photographic Record showing the Stimulatory Action of Solution of Sugar 2 per cent, solution of sugar, inducing a continuous enhance- ment of response for some time. Another stimulating agent is a dilute solution of Na2Co3. This when applied in i per cent, solution induces an P'iG. 95. Photographic Record showing Continuous Action of 2 per cent. NajCO^ Solution Preliminary exaltation followed by depression. enhancement of amplitude of response, but when given in strong solution, induces depression. An intermediate strength of solution shows preliminary enhancement followed by de- pression (fig. 95). at' EXCITABILITY UNDER CHEMICAL REAGENTS 1 37 While pursuing another Hne of inquiry on the effect of various strengths of solution of NagCog on the natural current, I obtained results which were parallel (p. 122). It was there shown that dilute solution of NagCog induced a positive variation of the natural current ; a strong solution, a negative variation, and that a solution of intermediate strength induced a preliminary positive followed by a negative variation. Thus the positive variation in the last-named experiments, already shown to be indicative of increased xcitability, was here seen to correspond with heightened amplitude of response, while the negative variation on the other hand is seen to coincide with depression of excitability. The application of a strong solution inducing excitation, carries the molecular condition of the tissue to the stage iE, where, as we know, the excitability is depressed. Another fact elucidated by this and similar inquiries, pvhich I have pursued elsewhere,^ lies in the fact that the difference between stimulants and poisons, so called, is often one merely of degree. Thus a stimulatory reagent, if given in large quantities, will be found to induce a profound depression, whereas a poisonous reagent in minute quantities may be found to act as a stimulant. In carrying out a similar investigation with regard to growth response, 1 found that sugar, for instance, which is stimulating in solutions of, say, I to 5 per cent., becomes depressing when the solution is very strong. Copper sulphate again, which is regarded as a poison, is only so at i per cent, and upwards, a solution of '2 per cent, being actually a stimulant. The difference between sugar and copper sulphate is here seen to lie in the fact that in the latter case the range of safety is very narrow. Another fact, which must be borne in mind in this connec- tion, is that a substance like sugar is used by the plant for general metabolic processes, and thus removed from the sphere of action. Thus continuous absorption of sugar could not for a long time bring about sufficient accumulation to Iause depression. With copper sulphate, however, the case ' Bose, P/ani RespoTise, p. 488. : 138 COMPARATIVE ELECTRO- PHYSIOLOGY is different. Here, the constant absorption of the minimal stimulatory dose would cause accumulation in the system, and thus ultimately bring about the death of the plant. Fig. 96. Photographic Record showing the Depressing Action of 5 per c< nt. HCl Acid The effect of very dilute acids is often to induce an enhancement of excitability, while strong solutions induce depression and abolition. In fig. 96 is shown the depression Fig. 97. Photographic Record showing Effect of i per cent. KHO Note tlie preliminary positive twitch at the fourth response after application. and abolition induced by the application of a 5 per cent, solution of hydrochloric acid. In dealing with the question of electrical response, we have seen that two opposed electrical effects occur in the EXCITABILITY UNDER CHEMICAL REAGENTS 139 tissue subjected to stimulation. One of these is the positive effect, and the other, the true excitatory change of galvano- metric negativity. As the latter is, under normal conditions, predominant, the simultaneous effect of both is a resultant negativity. The positive effect may, however, be unmasked, as we have seen, by abolishing the true excitatory effect of negativity (p. 66). This positivity may also be un- lasked, if, by the action of a chemical reagent, the time- FiG. 98. Photographic Record of Effect of 5 per cent. KIIO Note the complete reversal of response to positive at the beginning, and its subsequent abolition. elations of the two responses are changed, so that instead of occurring simultaneously, the one is made to lag behind the * other. This case will be seen very strikingly illustrated in fig- 97> which exhibits the effect of a i per cent, solution of KHO, on response to transmitted stimulation in the petiole of fern. In the normal responses here given, we observe I the resultant response of galvanometric negativity. The ^feipplication of KHO is first seen to reduce the excit- ability, as indicated by the reduced height of the responses. Later, we observe that the true excitatory effect is delayed. II 140 COMPARATIVE ELECTRO-rHVSIOLOGY Hence the positive effect is no longer completely masked. Its existence is now seen as a preliminary downward twitch in a di-phasic response, in the case of the fourth and succeeding records, after the application of KHO. In fig. 98, a stronger, namely a 5 per cent, solution of KOH,was used. And here, by the almost complete abolition of the excitatory factor, the response has undergone an apparent conversion to positive ; this positive response is, however, subsequently abolished by the death of the plant. CHAPTER XII i VARIATIONS OF EXCITABILITY DETERMINED BY METHOD OF INTERFERENCE rrangement for interference of excitatory waves — Effect of increasing difference of phase— Interference effects causing change from positive to negative, through intermediate di-phasic — Diametric balance — Effect of unilateral application of K HO— Effect of unilateral cooling. I HAVE explained how the variations of excitability brought about by various agencies may be determined, by recording the corresponding amplitudes of response. I shall now pro- ceed to describe a new and interesting method of making such determinations, by means of which it will be found possible to elucidate certain questions which without it must remain obscure. This method is, moreover, of extreme delicacy, enabling the investigator to detect the slightest I™ variation of excitability, induced by any agent. V* Let two points in the experimental tissue, say A on the right, and B on the left, be suitably connected with the galva- nometer, and let the occurrence of excitation at A on the right be represented by an * up ' response record, the excita- tory effect at B, on the left, being represented as * down.' Kf now the two points, A and B, be excited simultaneously, he resultant electrical response will be due to the algebraical ummation of the two excitatory electro-motive effects E^^ and Ib, these standing for the individual electrical effects at the two points A and B. Now if the intensities of the two effects be the same, and if their time-relations be also the same, it is evident that these two excitatory electrical waves, being of equal amplitude and having the same phase but of opposite signs, wi\\, by their mutqal interference, i^eutralisp i 142 COMPARATIVE ELECTRO-PHYSIOLOGY I each other. Under such balanced conditions, therefore, on simultaneous excitation of A and B, the resultant response will be zero. If now, under the modifying action of any external agency, the excitability of A be enhanced, it is clear that the resultant response will be ' up,' showing the greater excitability of the right-hand point. A similar effect will also be produced if the excitability of B be depressed. Similarly the depression of the excitability of A, or enhance- ment of B, would cause a resultant response which would be *down.' If, again, the two waves of excitation be not of the same phase, we shall obtain various di-phasic effects resulting from the algebraical summation of the constituent response- curves. The resultant zero-response may thus be converted into di-phasic, by the action of any agency which is capable of changing the time-relations of either of the constituent responses. I shall now proceed to describe the experimental arrange- ments by which two points in connection with E and E' may be excited, and the resulting electrical disturbances made to interfere with each other. For this purpose we may use the vibrational stimulation which has already been described, with certain necessary additions (fig. 99). The angle of torsional vibration which regulates the intensity of excitation is determined by two stops, P and Q. An elastic piece of brass, B, projects from the torsion-head. When a single stroke is given to this, a quick to-and-fro vibration is induced, the backward pull being supplied by the attached spring, s. The amplitude of this vibration remains always the same, as determined beforehand by the setting of the stops P and Q. The stroke is given by the striking-rod R, set in motion by the turning of a handle. What has already been said about the excitation of the right-hand side of the specimen applies equally to the left-hand, arrangements for the purpose being a duplicate of those just described. After deciding on a suitable angle of torsional vibration for the right, and taking the response at that point, we proceed to adjust the torsional angle on the left, so that the response there may be exactly EXCITABILITY DETERMINED BY INTERFERENCE 143 the same as that on the right. If the excitability of the two points had been exactly the same, equal amplitudes of vibra- tion would have resulted in the equal stimulation of both. But in practice the excitabilities are found to be slightly different and the angle of vibration of the one must, therefore, be so adjusted as to induce an excitatory effect exactly equal to that of the other, ^fe The two striking-rods, one on the right, R, and the other ^B>n the left, r', can be adjusted so that both are in the same I Fig. 99. R, r', striking-rods for stimulation of two ends of specimen ; B, elastic brass tongue projecting from torsion-head. For producing phase-difference r is adjustable in azimuth. II ertical plane, or so that one is in advance of the other. The left rod is permanently fixed to the rotating axis, but the right can be set at any angle that is desired, with the other. When the right striking-rod is set, pointing to zero of the scale, the two rods are in the same vertical plane, and the rotation of the handle causes equal vibrational stimulus by the two at the same moment. The excitatory reactions on right and left are now, therefore, of the same phase and of equal intensity, but opposed to each other. In fig. 100, a^ are reproduced the two separate and equal constituent espouses given by a specinien of stern of Amaranth. The 144 COMPARATIVE ELECTRO-PHYSIOLOGY * down ' curve was given by the individual excitation of the left, and the * up ' by the right. On the simultaneous excita- tion of the two points, the resultant response was zero {b). But if the excitation of one — say, the right — be increased by increasing the angle of vibration, the resultant differential response is found to be ' up.' It is obvious that a similar effect would have been observed had the stimulation of the right been kept the same, while its excitability was increased by any external agent. In these cases we have two opposed excitatory waves of similar phase, and of the same or unequal intensities, interfering with each other. Fig. ioo. [a] Isolated response of left side (down) and right side (up) ; {b) null-effect when excitations are simultaneous ; (<:), {d), {e) di-phasic responses obtained with increasing difference of phase. We shall next take some simple instances in which, while the stimulation is maintained constant, there is an increasing difference of phase. If the right-hand striking-rod R, instead of being set at zero, be set to the right, or at a phis angle, the rotation of the handle will cause a slightly earlier excita- tion of the right than of the left. If, on the other hand, the rod be set at a minus angle, the excitation of the right will be later than that of the left. Under these circumstances, instead of the null-effect due to continuous balance, we shall have a di-phasic response. It is also clear that as the phase difference is increased, the neutralisation of effects will become less and less perfect, the separate constituent respon- ses being thus rendered increasingly apparent, In fig. lop, r, EXCITABILITY DETERMINED BY INTERFERENCE I45 is seen the di-phasic effect which was induced when the excitation of the right was made to lag slightly behind that of the left, by the adjustment of the striking-rod at a small minus angle. The first of the two twitches, which is downwards, indicates the relatively earlier excitation of the left-hand contact. As the phase-difference was increased progressively as in {d) and {e\ it is seen that the constituent elements of the di-phasic response are increased corre- spondingly. It is also clear from this that, having obtained the null-effect, if any agents were afterwards applied locally which would make the excitation of the one point earlier than that of the other, we must then expect the null-effect to be modified to di-phasic. An earlier * up ' twitch would now indicate that the right-hand contact, having had its re- action quickened, was the first to respond ; an earlier ' down ' «itch the opposite. We thus see how the conversion of the null-effect into a resultant ' down ' negative or * up ' positive, could be utilised as a test of the excitatory or depressing nature of a given reagent. We also see how the conversion of this null into a di-phasic effect would give us indications as to the change of time-relations induced by the reagent. I shall here, before going on to describe the results obtained with plants, give a photographic record (fig. loi) of certain positive, nega- tive, and di-phasic effects obtained in the electrical response of the inorganic substance, tin, under appropriate modification of the excitability of its two contacts by various chemical reagents.^ Turning now to the question of the determination of the effects of the various reagents by the Method of Interference, we may, as we have seen, cause simultaneous excitation of right and left, by means of the apparatus which has just been described, and which I shall distinguish as the Longitudinal Balance. There is, again, another and simple method of accomplishing the sanie object, by means, namely, of the Diametric Balance, the diagram of which has already ^ Bose, Response in the Living and Non-Living, p. 115. L II 146 COMPARATIVE fiLECTRO-PHYSIOLOGY been given in fig. 80. Using this arrangement, the specimen is clamped at one end, the vibration-head being at the other. Electrical connections are now made with the two dia- metrically opposite points, A and B, of which one, say A, is the upper, and B the lower. In a tissue which is isotropic, vibrational stimulus will induce equal and simultaneous excitation at the two points A and B. The effect of any given agency is tested by applying it locally, say at A, and observing the resultant variation of the response. I shall (a) (l>) ic) Fig. ioi. Photographic Record showing Negative, Di-phasic, and Positive Resultant Responses in Tin under appropriate modifications of excitation of the two contacts here give examples of results obtained by both these methods, thus affording an indication of the extent of their applicability in various investigations. We have seen in the previous chapter that the application of strong solution of potash will abolish the excitability of a tissue. Using the Longitudinal Balance, I took a petiole of Bryophyllum and first made such adjustments that the right * up ' and left 'down' responses were almost equal. On now producing simultaneous excitation of the two ends, a di-phasic response was obtained, due to the fact that the left-hand point was the quicker to respond. Strong solution tXClTABILlTY DETERMINED BY INTERFERENCE 14; of potash was next applied on the right-hand point, and from the record it is seen that the 'up' part of the di-phasic response, due to the excitation of the right-hand side, was thus completely abolished, the ' down ' response being at the same time increased by the suppression of this opposing response (fig. 102). In order to demonstrate the use of the Diametric Balance Method, I undertook to investi- gate by its means the influence of the lowering of tempera- ture on excitability. For this It Fig. 102. Photographic Records. {a) Di-phasic response of petiole of Bryophylliim, the up compo- nent being due to the excitation of right side. Strong application of KHO on the right abolished this responsive component, giving rise in {b) to enhanced down response Fig. 103. Photographic Record of Response of Petiole of Cauliflower by the Diametric Method A contact was naturally more excitable, hence resultant ' up '-response. Ex- citability of A being depressed by local application of ice, the re- sultant response became converted to ' down ' ; normal ' up '-response was restored on allowing the tissue to return to surrounding temperature. purpose, I took a petiole of cauliflower. In this instance, the natural excitability of the upper contact. A, was greater than that of the lower, B. Hence the resultant response was not zero, but * up,' The point A was now cooled locally by ice. This process so lowered its excitability that that of B was now relatively the greater, hence the resultant response was found to be reversed or ' down.' The point A was next L2 148 COMPARATIVE ELECTRO-PHYSIOLOGV allowed to return to the surrounding temperature of the room, records of the response being taken meanwhile, at intervals of one minute. It will be seen how, by means of the gradual restoration of the original excitability of A, the resultant response changes gradually from negative to zero, and then again from zero back to positive, indicating the restoration of the naturally greater excitability of A (fig. 103). . We have thus studied two different methods, both of which depend on interference, for the determination of the variations of excitability induced by different external agents. In a subsequent chapter we shall study a modification of this method, by means of which it is possible to demonstrate the variations not only of excitability but also of conductivity under various reagents. CHAPTER XIII f CURRENT OF INJURY AND NEGATIVE VARIATION Different theories of current of injury — Pre-existence theory of Du Bois- Reymond— Electrical distribution in a muscle-cylinder— Electro-molecular theory of Bernstein— Hermann's Alteration Theory — Experiments demon- strating that so-called current of injury is a persistent after-effect of over- stimulation — Residual galvanometric negativity of strongly excited tissue — Distribution of electrical potential in vegetable tissue with one end sectioned — Electrical distribution in plant-cylinder similar to that in muscle- cylinder— True significance of response by negative variation — Apparent abnormalities in so-called current of injury — ' Positive ' current of injury. I a section be made of an uninjured nerve or muscle, the transverse contact will be found to be galvanometrically negative, as compared with an uninjured longitudinal contact. I shall have occasion in the present chapter to give a simple explanation of this phenomenon and of the excitatory nega- tive variation of the current of injury. It is, therefore, only necessary to recapitulate briefly the three theories which have hitherto been proposed on this subject. The Pre-existence Theory of Du Bois-Reymond supposed that the smallest particle had the same electro-motive characteristics as the entire tissue, each such electro-motive molecule consisting of two bi-polar portions, the positive poles of any two molecules being always face to face with each other. This theory was based upon the fact that a muscle-cylinder, for example, exhibited a peculiar distribu- tion of electrical tension. There are in such a cylinder, one longitudinal and two transverse surfaces. Midway in the cylinder is the equatorial zone of the longitudinal surface, and this zone is positive to all the rest. Thus the electro- motive difference between one electrode placed on the I I50 COMPARATIVE ELECTRO-PHYSIOLOGY equator, and the other, is increased as the latter is moved further and further away, say towards the right transverse section. The distribution of electrical tension on the left side of the equator is symmetrical with this (fig. 104). On these facts was based the theory of Du Bois- Reymond ; but this has Fig. 104. Distribution of Electrical . , r j . 1 • Tension in Muscle-cylinder. smce been found to be m- adequate. I shall later return to the explanation of the particular distribution of electrical tension involved. According to the theory of Bernstein, known as the Electro-chemical Molecular Theory^ the fundamental attribute of the molecule is chemical. Its poles are supposed to attach to themselves electro -negative groups of atoms, while its sides attach oxygen, and stimulation is supposed to be attended by explosive chemical changes. According to Hermann's Alteration Theory^ finally, all the electro-motive activities of living tissues are supposed to be due to chemical rather than molecular changes of the substance. In amplification of this theory, Hering attributes all electro- motive phenomena to the disturbance of equilibrium by up and down chemical changes. It is my intention to show in the course of the present chapter that the current of injury is an after-effect of over-stimulation. And since excitation is fundamentally due to molecular upset, we shall best understand the plectro-Riotive changes concomitant with it, if we first study jt an(} its after-effect under the simplest conditions, namely those of inorganic substances. For here the action of such complicating factors as assimilation and dissimilation is clearly out of the question. We have found for example that a piece of well-annealed YV'ife was iso-electric throughout its length. In the first CURRENT OF INJURY AND NEGATIVE VARIATION 151 place, when a portion of it was subjected to any molecular disturbance, an electro-motive difference was induced, as between the molecularly disturbed or excited and the un- disturbed areas. The intensity of this electro-motive change in the second place, was seen to increase with intensity of excitation. And, thirdly, the recovery from excitation was seen to be delayed, where the intensity of stimulus was strong (fig. 105). This is shown in the electrical II Fig. 105. Photographic Record showing Persistent Electrical After-Effect in Inorganic Substance under Strong Stimulation. Note the tilt of base-line upwards The vertical line to the right represents -i volt. sponse of tin as a persistent after-effect, the sign of hich is the same as that of the excitatory electro-motive hange. A similar state of things is exhibited mechanically in a torsioned wire. When the torsion is moderate, and the molecular distortion slight, the released wire quickly re- covers its original position of equilibrium. But when the| torsion is excessive and the wire strained beyond a certain limit, it remains for a long time in a torsioned condition, 52 COMPARATIVE ELECTRO-PHYSIOI.OGY even after it has been set free. Recovery is thus, in such a case, indefinitely delayed. In other words, a molecularly over-strained substance exhibits a persistent after-effect. Turning next to plant response, we find a similar per- sistence of the after-effect to occur in consequence of over- stimulation. And first we shall take the simplest case — that in which the tissue is directly stimulated. Here the specimen was petiole of cauliflower, and increasing stimuli Fig. io6. Photographic Record exhibiting Persistent Galvanometric Negativity in Plant Tissue after Strong Stimulation Stimuli applied at intervals of three minutes. Vertical line = -i volt. were applied, at intervals of three minutes, by means of a gradually increasing angle of torsional vibration. It will be noticed that whereas the electrical recovery from moderate stimulation — as seen in the first of the series — is complete, it becomes, with increasing stimulus, more and more in- complete (fig. 1 06). In other words, the tissue, after strong stimulation, is seen to exhibit an after-effect of residual galvanometric negativity, which is really due to incomplete molecular recovery, in consequence of over-strain. In the cases just given, stimulation was applied directly. CURRENT OF INJURY AND NEGATIVE VARIATION 1 53 We shall now, however, take an instance in which excitation is transmitted and observe the persistent negative after-effect, due to strong stimulation. We know that a cut (mechanical section) or the application of a hot wire (thermal section) acts as a strong stimulus, and the effective intensity of such stimulation will obviously decrease with increasing distance from the point of stimulation. Hence, if we observe the persistent excitatory change of galvanometric negativity, which is induced as between an indifferent point — say, the surface of a leaf —and points increasingly near to the zone of section, we shall find that the electro-motive change is greatest at the point of section, and is progressively lessened as we recede from it. This induction may be verified experimentally by taking readings of the persistent negativity, as between an indifferent point, B, and points such as the contacts a, b, c, had m this case become attention towards an inquiry into reversed, the killed end having ,, r ,f 1 become gal vanometricallyposi- the causes 01 these anomalous jj^g ^ ^^ reversals. The subject therefore resolved itself into an investigation as to what conditions determined the negativity or positivity of a tissue at the onset of death. My first attempt, then, was to study a case in which the approach of death was natural, and not the result of any sudden or violent change, such as might conceivably give rise to abnormal reactions. And in my search for suitable specimens, I noticed that often, owing to local mal-nutrition or other causes, the leaves of plants exhibited spots or areas, from which, as centres, death pro- ceeded in constantly widening circles. Thus, in the leaves of Co/ocasia, for example, we find such dead and dying areas in otherwise fairly healthy leaves. The innermost of these I % l66 COMPARATIVE ELECTRO-PHYSIOLOGY patches may be quite dark and discoloured, while, as the living tissue is approached, this dark passes imperceptibly into yellow colour. And beyond this, again, we find the discolouration of yellow passing into the vivid green of living tissue. Proceeding thus in a radial direction inwards, towards the centre of such a patch from the living green, we shall find all possible stages of death, from its initiation, somewhere on the border-line between green and yellow, to its phase of completion, in the dark central area. On testing the electrical conditions of these different parts, I found that the border between green and yellow was negative to the living green surface. But the same point was also negative to the dead central area, and more negative to this than to the living tissue. Hence the dead was relatively positive to the living. 'Or if we make one fixed contact on the living tissue, and if the second exploring contact be made with various points sudcessively on a radial line passing from this to the centre of the dead area, these contacts will pass in succession through the living, the dying, and the dead. The variation of elec- trical potential will be found to be at its greatest along this line. The electro-motive difference between the point which has been fixed on the living tissue, and the exploring second contact, will at first be found to increase. The maximum difference is attained on reaching the border-line between green and yellow, or very little beyond this, this point being galvanometrically the most negative. On now passing further inward from this point, the maximum difference is found to decrease, till we come to a point in the dead tissue which is iso-electric with the living. On now again passing inwards, to the still more completely dead tissue of the central area, we find that we are approaching points which are more and more galvanometrically positive, as compared with the living tissue. The dying point on the border-line between green and yellow is thus the most negative, and points to the right or left of this are positive in comparison with it, the dead, however, being more positive than the living. It has been said that the electro-motive variation is most rapid along the CURRENT OF DEATH 167 radial line. On the other hand, we obtain series of equi- potential surfaces whose outlines closely follow those of the boundaries of the different degrees of discolouration. I shall next proceed to give quantitative measurements. The first point to be considered is that of the choice of a definite electrical level, which is to be used as a standard. If this point be selected in the living tissue, we shall find that our standard of comparison is extremely variable, since the tonic condition, on which its electrical level depends, is itself subject to change. The only condition which cannot be modified in any way is that of complete death. This may be taken, then, as the standard level. The method of experi- ent will thus consist in selecting a series of equidistant points, abed, and so on, 5 mm. apart, along a radial line, passing outwards from the central area, which is completely dead, to l^phe green tissue. The non-polarisable contacts E and e' are first placed on a and b, then on b and r, c and dy and so forth. The external circuit contains a high resistance, compared with which any difference of resistance, as between any 5 mm. of interposed tissue, becomes negligible. Hence, the successive deflections of the galvanometer indicate the electro-motive (difference that exists between a and ^, b and r, and so on. !■ One difficulty which is experienced, in these measure- ments of small electro-motive differences, lies in securing the iso-electric condition of the non-polarisable electrodes them- selves. Whatever precautions are taken in the construction of these, a small electro-motive difference will sometimes be found to exist between them. The existence of such a difference is easily tested by bringing the kaolin ends of the IHwo electrodes in contact, or by dipping both of them close together in a vessel of normal saline solution. Any electro- motive difference of the electrodes, however small, will now give rise to a large galvanometric deflection. IK This difficulty may be overcome by first taking special ^precautions as to the purity of zinc rods and the chemicals employed, and secondly, by keeping the electrodes for a long time short-circuited, with their ends dipped in normal I i68 COM PAR ATI VE ELECTRO-PHYSIOLOGY saline. In very obstinate cases, however, I succeeded in eliminating all differences by subjecting the electrodes to cyclic variations of alternating electro-motive force. By means of a Pohl's commutator, without cross-bars, the electrodes were put in connection with an alternating source of E.M.F., and with the galvanometer intended to test the resulting variation in the E.M. difiference, by turns. A small hand-driven alternating-current generator was used for this purpose. The speed of rotation of this machine was gradually raised to a maximum, and afterwards as gradually slowed down. Thus at each cycle the electrodes were subjected to ascending and descending intensities of alternating electro-motive variations. The effect of such cyclic changes, in diminishing the existing electro-motive difference between a pair of electrodes, specially selected for carelessness of preparation, will be clearly seen from the following tabular results : ■i c, •is Condition at starting Galvanometric deflection E.M. difference Original difference . After first cycle After second cycle . After third cycle . 360 divisions 40 „ •009 volt •001 ,, „ „ It will thus be seen that, after a very short time of this treat- ment, the two electrodes were rendered iso-electric. I next proceeded to determine the distribution of elec- trical potential in the various portions, living and dead, of the leaf. In order to remove any accidental strain, the leaf was placed in tepid water, and kept there for about half an hour, till the water was cooled to the surrounding temperature. The experiment was then carried out, in the manner already described, and the following tabular statement shows the results obtained. The electrodes, it will be remembered, were placed successively at points 5 mm. apart from each other, along a radial line proceeding from the dead tissue to the living, the first point being taken as zero ; CURRENT OF DEATH 169 m Position of Electrodes Galvanometric Deflection 0—5 mm. division 5-10 „ - 10 ,, 10- 15 „ — 20 ,, 15 - 20 „ - 45 „ 20 - 25 „ — 140 ,, 25 - 30 „ — 140 ,, 30 - 40 „ + no ,, 40 - 45 .. -h 20 ,, It will be observed that as we proceed from the dead to the dying, the negativity of the latter rapidlyincreases, the aximum being at 30 mm. from the zero-point taken on the \\y^^^::^:i^' \A '5 15 2D 25 30 35 40 200 250 300 350 4Q0 Fig. 113. Distribution of Electric Potential in Lamina of Colocasia along a radial line from dead to living through intermediate stages. Ab- scissa gives distance in mm. from chosen centre in dead tissue, ordinate represents galvanometric negativity in divisions. Dead tissue repre- sented dark, dying shaded, and living white. 'dead tissue. This point of maximum-negativity almost coincides with the visible border-line between the yellow and the green. Beyond this, however, there is an electrical II I/O COMPARATIVE ELECTRO-PHYSIOLOGY reversal, the living becoming increasingly positive, as com- pared with the dying. An inspection of the curve (fig. 1 1 3) shows that while there is a point in the tissue between the dying and the dead, which is equipotential with the living, the completely dead tissue is positive to the living. I next carried out an experiment in which death was artificially induced, by immersing a portion of the tissue in boiling water. In connection with this, I may say that it is extremely difficult to ensure the complete death of a thick tissue. It is only the outside layers which undergo death easily, but the interior tissues, from their protected position, are extremely resistant, and it is only after prolonged immersion in boiling water that death can really be ensured throughout. In the present experiment, however, where only a part of the tissue is to be killed, such prolonged immersion would cause death to encroach upon those portions of the tissue which were intended to be kept alive. This difficulty was met by choosing a specimen, the inside of which was accessible to boiling water. The peduncle of the water-lily {Nymphcea alba) in transverse section appears extremely reticulated, and there is thus no difficulty in exposing all its parts to the direct action of the hot water. The upper end of the peduncle was kept surrounded by a cloth moistened in ice-cold water, the lower end being immersed in boiling water for ten minutes. The specimen was then placed in tepid water, and allowed to cool down slowly. In this way a length of the peduncle was ob- tained, in which one end was completely killed, whereas the other remained fully alive, the intermediate portions showing all stages of the transition from the living to the dead condition. In order to determine the electrical distribution in its different parts, I now employed the potentiometer method of balance. One electrode was per- manently connected with that dying point which by a previous test had been found to exhibit maximum nega- tivity. The second electrode was placed at successive points. CURRENT OF DEATH 171 each of which was nearer than the last by 5 nim. to the dead end, which was to the left. The same process was now re- peated, the successive readings however being taken towards the right or living end. At each point, the electro-motive ■^ difference was balanced Kby the potentiometer. This straight form of potentiometer had a 3cale divided into one ■■thousand parts (fig. 114), and when its terminal electro-motive force was adjusted to i volt, each division of the potentiometer' was equal to -ooi volt. The following table gives the results obtained : Fig. 114. Straight Form Potentiometer A B is a stretched wire with added resistances, R and r'. s is a storage cell. When the key, K, is turned to the right, one scale division = -ooi volt, when turned to the left one scale division = -oi volt, p is the plant. li Towards left or dead end, E.M. difference in ysoo volt Distance from maximum negative point io the left (— ) or , to the right ( + ) Towards right or living end, E.M,. difference in ^oso volt 1-2 •5 cm. •9 5'o i-o „ 2-5 i8-2 I -5 » 47 1 22-0. 2-0 „ 6-8 23 -O' . 2-5 „ 10 -o 23-3 '■ 3'o „ 12-8 3'5 ,, 15-6 — 4'0 ,, i6-8 — 4-5 >. irs — 5-0 „ i8-o Here, also, as in the case of natural death, we find a point in the dying tissue which is most negative. From the curve given in fig. 115, it will also be seen that as we pass away from this point in cither direction towards the living or dead area, we find an increasing positivity ; the curve for the dead portion is, however, much steeper than that for the living. Thus two points, one i'^ cm. to the left in the dead tissue, and another 5 cm. to the right in the living tissue, are iso-electric. But \yhilq the maximum positivity of the living II 172 COMPARATIVE ELECTRO-PHYSIOLOGY is 'OiS volt, that of the dead is "0233 volt. Hence the dead tissue is here positive to the living, to the extent ot •0053 volt. We have seen that the prevailing idea is that the dead is negative to the living. But from the results here shown, we can see that this is not a complete statement of the case. Since then the electro-motive variation, instead of showing a Fig. 115. Distribution of Electric Potential in Petiole oi Nymphcea alba^ one end of which has been killed. The point of maximum negativity is taken as zero, distances to the left or towards the dead taken as minusy to the right or living, as plus. Ordinate represents potential difference in thousandths of a volt. progressive change from the living to the dead, exhibits a maximum difference, followed by a reversal, it may be asked, what is the reason of this anomaly ? Much light is thrown on this subject from the results given by another line of inquiry, to be explained in detail in Chapter XVI. It is there shown that the plant-tissue on the first onset of death exhibits a sudden contraction, indicative CURRENT OF DEATH 1^3 of a strong excitatory reaction. This corresponds with the rigor mortis of the animal, and by means of suitable apparatus, the concomitant mechanical response can be recorded. An electrical record of the same phenomenon may also be obtained, in the form of an electrical spasm of galvanometric negativity. Succeeding to this rigor of the dying tissue, a post-mortem relaxation takes place, with a concomitant change from galvanometric negativity to positivity. ^^ft Now in a tissue which has been killed unilaterally only, ^^Rrit will be understood that all possible gradations are to be ^^' expected. Passing from the completely dead to the fully alive, we must necessarily pass through various zones, l^pbeginning with the abnormally relaxed, through the inter- mediate highly contracted and rigored tissue on the death- frontier, to the living, which is not so contracted as the IBt dying, and not so relaxed as the dead. At the point where the onset of death is recent, the rigor, or excitatory contraction and galvanometric negativity, are at their maximum. Compared with this, the slightly tonically con- tracted living is positive, but not so positive as the abnormally relaxed dead. The death-frontier, however, is not fixed. It is con- 'itinually encroaching on the living. The line of maximum rigor and galvanometric negativity is thus also shifting in the same direction. Along with this, however, the opposite process of post-mortem relaxation is proceeding ; so that a point which was, in consequence of rigor, maximally negative, ; becomes gradually converted to positive. This positivity of dead tissue as compared with living, which has here been demonstrated in the case of the plant, I find to be also true of animal tissue, in those cases which il have investigated. Thus, while an injured and dying area .in a frog's nerve is negative, an already dead area is -positive, relatively to the living nerve. There is, moreover, an intermediate area, between the dying and dead portions of the nerve, which is iso-electric to the living. II t74 COMPARATIVE ELECTRO-PHVSIOLOGY Hence, having one contact fixed on a living area and the other on (i) the dying, (2) the intermediate, and (3) the dead tissue, we shall obtain three different types of what is known as the ' injury-current' In the first of these the second contact will be negative, a condition which has hitherto been assumed to be the . sole characteristic of the current of injury. But there are two other cases to be considered. Of these, when the second contact is made at a point intermediate between the dying and dead tissues, we shall find it to be iso-electric with the first, or living contact. And thirdly, when the second contact is on a dead area, the latter will be positive to the first, or living contact We thus find three cases of the current of injury — the first being negative, the second zero, and the third positive. Taking the first of these— that in which the injured contact is negative — the action -current, in response to stimulus, will bring about a negative variation of the so- called current of injury. In the second, the result will be indeterminate, since the injury-current is zero. In the third, the response will be by a positive variation of the current of injury. 1 give below three photographic records in illustration of these three cases, obtained with vegetable nerve. I may state here that I have often observed results precisely similar in the case of frog's nerve also. In the first record, in fig. 116, the thermal injury was moderate. The injured point was thus negative, and the current of injury is represented here by an up-line. The responses are seen to be by negative variation. In the second record the injury was greater, and the injured point was almost neutral ; that is to say, on making contact there was a slight up-twitch, which subsided to zero. There is here, then, no current of injury. The subsequent responses are, however, down, the action-current being away from the living contact. In the third record the injury was so great as completely to kill the injured point, which thus became positive to the living. The RESPONSE BY POSITIVE VARIATION I7'5 reversed injury-current is represented as down, the subse- quent excitatory responses are also down, and constitute a positive variation of the current of injury. It will thus be seen that an identical excitatory reaction of the living tissue appears to give rise to directly opposite Fig. ii6. Photographic Records of Responses of Vegetable Nerve, one end of which has been injured In the first injury was slight; current of injury represented up, response by negative variation. In the second, injury greater; injured point neutral, response down. In the third, injured point killed ; injury current reversed down, response by positive variation. effects— namely, a negative or a positive variation of the injury-current. I Br ^ S^^^ below a short summary of the diversities of response which may occur when either the natural, or the injury-current, is taken as the current of reference. (a) c "* B' J-R (5; B. —jnjur L V^m Fig. 117. Typical Cases of Variation of Current of Rest and Action- ^^B Current. Specimen originally isotropic (a) A, end slightly injured and negative ; c, current of injury ; R, action- current, a negative variation of c. {b) A, end killed and positive ; c, current of injury ; R, action-current, a positive variation of c. First — we take the case where the point A is slightly injured (fig. 117, a). The current of injury, C, is a->b, and the responsive current, R, is B -> A, constituting a negative variation. 176 COMPARATIVE ELECTRO-PHYSIOLOGY But when A is killed, the current of injury C is B -> A, the responsive current is also B -^ A, constituting a positive variation (fig. 117, d). Second — we take an instance where, owing to some physio- logical difference, an intermediate point A is less excitable than B or b' (fig. 118, a). The primary natural current will here be from the less to the more excitable : that is to say, C will be A-> b' and A -► B. If stimulus be now applied at X on the right, an identical excitatory current R, flowing away from the excited point from right to left, will cause seemingly opposite effects : that is to say, a negative variation of A->B' and a positive variation of A -> B. (a) (h) -< c *- >-c-« B'^ T ^B'A' f ' 'A- R^ = R< s Fig. 118. Typical Cases of Variation of Current of Rest and Action- Current ; intermediate point naturally less or more excitable than either of terminal. (a) Intermediate a, less excitable, shown by vertical shading ; current of rest A->B and A->b' ; when right-hand point x excited, action -current R from right to left, gives rise to negative variation of a->b', and positive variation of A-»B. {b) Intermediate B, more excitable, shown by horizontal shading ; current of rest a^-b and a'->b ; action-current on excitation at x , from right to left, giving rise to positive variation of a'->b and negative variation of a->b. Again, we may have the intermediate point B naturally more excitable than a' or A. The natural current C will be A-^B and A'->B (fig. 118, b). Stimulation at x will now give rise to an excitatory current R, from right to left. The results here will, however, appear to be exactly the reverse of those in the last case : that is to say, an identical current, R, will give rise to a positive variation of A' -> B, and negative variation of A->B. Instances of these effects will be given in Chapter XVIII. And, lastly, we may have a typically anisotropic tissue, composed of two halves, which are unequally excitable — as, for instance, the upper and lower halves of the pulvinus of Mimosa^ or the muscle and gland in a muscle-and-gland RESPONSE BY POSITIVE VARIATION 177 preparation. Under normal conditions the primary or natural current C is from the less excitable A to the more excitable B, represented by A -^ B (fig. 1 19, a). The action-current R, being in the opposite direction B -> A, constitutes a negative variation. But owing to the after-effect of excitation, such as may occur in isolating the specimen by section, the normal resting current C is reversed to B -> A (fig. 1 19, b). Here the end B may still be the more excitable of the two, hence the action-current »-> A will constitute a positive variation of the current of rest. But when B becomes fatigued, its excitability is reduced below that of A ; hence the action-current is from the relatively more to less excitable, i.e. A->B. In this case, (a) (b) (c) >c — l=jl R< B R^ B B ^R b ^V Fig. 119. Typical Cases of Variation of Current of Rest and Action- ^B Current. Anisotropic organ, b end originally more excitable than A ^V (a) Current of rest a^>b ; action-current, R, in opposite direction ; response ^B by negative variation, {b) Owing to excitatory after-effect, current of ^B rest reversed to B->A ; B nevertheless more excitable than A ; action- al current, r, is B->A; response by positive variation, {c) Current of ^B rest reversed b->a ; action-current also reversed a->b, by depression ^B of excitability of B, owing to fatigue ; response by negative variation. on account of the reversal of both the current of rest and action-current, the latter appears to constitute a negative variation of the former (fig. 1 19, c). It will thus be seen how intricate and diverse are the responsive variations of the resting current, induced by stimulus. Sometimes negative, sometimes positive, it would appear as if there were no guiding principle to regulate these phenomena. The so-called explanations hitherto attempted have consisted in assigning the positive variation to a hypothetical process of assimilation, and the negative to dissimilation. Such explanatory phrases reach the climax of absurdity when we find ourselves compelled to ascribe one identical excitatory reaction now to assimilation and hen to dissimilation. N If I7B COMPARATIVE ELECTRO-PHYSIOLOGY Indeed it must be said that, however suggestive the general theory of assimilation and dissimilation may have been found, its abuse has often stood in the way of physio- logical inquiry. The inquirer, when faced with any difficulty, instead of attempting to surmount it by patient inquiry, was tempted rather to evade it by invoking the aid of an hypothesis which could be made with equal ease to explain a given fact or its direct opposite. We must remember that in the investigation of obscure problems, the danger is always, instead of seeking an underlying law, to become satisfied with the mere registration of phenomena, and by naming these to imagine that they have been explained. The resulting chaos in the present case has served to deepen the impression that vital phenomena must always remain capricious and mystical. But when we come to survey the facts that have been described, we find the phenomena of response, however diverse they may at first sight appear, to be in no way governed by chance or caprice. They are, on the contrary, definite and uniform under definite conditions. As regards the so-called current of rest in a naturally isotropic tissue, of which one end has been subjected to injury, we must remember that the effect of injury is one of excitation, its sign, within limits, being contraction and galvanometric negativity. But we have seen that when a point is over stimulated, fatigue-changes appear which give rise to a reversal of its normal sign of response, from con- traction to expansion, from negative to positive {cf. fig. 64) The change at death, in which contractile rigor passes into post-mortem relaxation, is analogous to this. Thus when one end of the specimen is merely injured, that end becomes more or less persistently galvanometrically negative, the current flowing away from it. But when the same end is actually killed, the electrical change may be reversed, to one of galvanometric positivity. In an isotropic tissue, then, we may, by moderate injury, bring about a state of anisotropy, under which the uninjured RESPONSE BY POSITIVE VARIATION 1 79 end is rendered relatively the more excitable, and galvano- metrically positive, compared with the inexcitable injured end. In a naturally anisotropic organ, we have a state of things which is analogous. In this case, in the primary condition, the more excitable surface is galvanometrically positive. But under the excitation due to preparation, or accidental dis- turbance, this more excitable surface becomes the more excited, and, relatively to the other, galvanometrically negative. These varying changes in the direction of the so-called resting current, or current of reference, are the cause of the existing anomalies in the interpretation of response by the positive or negative variation. But the direction of the action-current under normal conditions is always the same. On diffuse stimulation it is always from the more excited B to the less excited A. The differential excitability or anisotropy, may be either natural, or artificially induced, as by injuring one end of an isotropic tissue. There are two different conditions under which the normal effect may undergo reversal, those, namely, of great sub-tonicity or excessive fatigue. But the statement that the responsive current is always from the more excited to t he less excited, remains universally true. Numerous illustra- tions, in verification of the cases laid down, will be met with in the course of subsequent chapters. N2 CHAPTER XV EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE General observation of effect of temperature on plant — Effect of fall and rise of temperature on autonomous response of Desmodium — Effect of frost in abolition of* electrical response — After-effects of application of cold, in Eucharis, Ivy and Holly — Effect of rise of temperature in diminishing height of response — This not probably due to diminution of excitability — Similar effect in auto- nomous motile response of Desmodium — Enhanced response as after-effect of cyclic variation of temperature —Abolition of response at a critical high temperature. We have now seen that the physiological activity of a Hving tissue may be gauged by means of its electrical response. We know further that the influence of temperature is of importance in the maintenance of a proper physiological condition. There is a certain range of temperature which is favourable to this, and above or below these limits physio- logical efficiency is diminished. If the plant be kept too long at or above a certain maximum temperature, it is liable to undergo death. Similarly, there is a minimum point at which physiological activity is arrested, and below which death is apt to occur. The plant has thus two death-points, one above the maximum and the other below the minimum temperature. Some can resist these extremes better than others, and length of exposure is also a factor which should not be for- gotten in the question of the ultimate survival of the plant under the given unfavourable conditions. Certain species are hardy, while others succumb easily. An unmistakable indication of the effect of temperature on physiological activity is found in the variations induced by it in the autonomous motile pulsations of the telegraph plant, Desmodium gyrans. Here, too great a lowering of the EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE l8l temperature abolishes the pulsation. In fig. 120 are seen (i) the records of normal pulsations; (2) their arrest under the application of ice-cold water ; and (3) their revival, as the plant regains the temperature of the room. In fig. 121 is shown the effect on similar pulsations of a rising tempera- ture. The records in this case were obtained with a different Fig. 120. Photographic Record showing Effect of Rapid Cooling, by Ice-cold Water, on V\x\?,a.\.\on?>'j:i{ Desfnodium gyraits Normal pulsations recorded to the left. Effect of application of ice-cold water is seen in the production of diminished amplitude and abolition of pulsation. Gradual return to the temperature of the room revives the pulsation in a staircase manner, the period remaining approximately constant. Note that cooling displaces the pulsation in a downward or contracted direction. Gradual warming, conversely, is seen to produce the opposite displacement towards expansion. Up-records represent the fall of the leaflet, down-records its rise. li specimen, and it is seen that the pulsations are diminished in amplitude while their period is quickened, with rise of temperature. When the temperature is raised still higher, ey come to a stop altogether. We shall next proceed to observe the effect of temperature on the electrical response of plants. As regards the influence of cold, for example, I have found, during the course of a research carried out in England, that after frosty weather, I 1 82 COMPARATIVE ELECTRO-PHYSIOLOGY t he electrical responses undergo an almost complete a boli- tionl During a certain week, for instance, the temperature was io° C, and the electrical responses then obtained from radish {Raphanus sativus) were considerable, giving an E.M. response which varied from '05 to 'i volt. Two or three days afterwards, however, as the effect of frost, I found the electrical response of this plant to have practically dis- appeared. A few specimens were found nevertheless which were somewhat resistant. But even in these the average E.M. response had only a value of "003 volt, instead of the normal mean of '075 volt. That is to say, their average sensitiveness had been reduced to one twenty-fifth. On now Fig. 121. Photographic Record of Pulsations of Desmodiiitn during Continuous Rise of Temperature from 30° C. to 39° C. warming these radishes to 20° C. there was an appreciable revival, as shown by their increasing response. But in those specimens which had been frost-bitten, warming effected no restoration. From this it would appear that frost killed some, which could not be subsequently revived, whereas others were reduced to a condition of torpidity from which, on \ warming, there was a revival. I have also investigated the effect of an artificial lowering of temperature on the electrical response of plants. The Eucharis lily is particularly sensitive to cold. In this case I took the petiole, and obtained response at the ordinary temperature of the room, which was at the time 17° C. I then placed it for 15 minutes in a cooling chamber at a EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE 1 83 temperature of —2° C. On now again trying to obtain response, it was found that it had practically disappeared. The same specimen was next warmed to 20° C, and this induced a revival of response (fig. 122). I was next desirous of studying the after-effect of lowered temperatures on different plants. For this purpose I chose three specimens (i) the |\ (a) petiole of Eucharis Lily, (2) the stem of Ivy, and (3) Holly. I took their normal responses at 17° C, and after- wards placed them in an ice-chamber at a temperature of 0° C. for 24 hours. The specimens were then taken out, and their responses under stimu- lation once more re- corded (fig. 123). From these it will be seen Hkhat while the respon- ^siveness of the delicate I Eucharis Lily was com- j pletely abolished, that of the hardier plants, i Holly and Ivy, exhibited complete revival. [ One interesting fact which I have noticed is that when a plant approaches its death-point, by reason of excessively high or low temperature, not only is its re- sponse, of galvanometric negativity, diminished to zero, but Iit is even occasionally reversed to positive. This effect is due to the unmasking of the positive, by the abolition (c) (b) Fig. 122. Diminution of Response in Eucharis by Lowering of Temperature (a) Normal response at 17° C. {b) The response almost disappears when plant is subjected to —2° C. for fifteen minutes. {c) Revival of response on warming to 20° C. 84 COMPARATIVE ELECTRO-PHYSIOLOGY Wc shall next study the effect on the electrical response .of the plant of a rise of temperature. The great difficulty of this investigation lies in raising the plant-chamber to the various determinate temperatures required. I was able, however, to accomplish this by means of electric heating. A spiral of german silver wire was placed in the plant-chamber {cf. fig. 2i), and by varying the intensity of the current the temperature was then regulated at will. In the process of this electric heating a complicating factor was found in the excitatory action of any sudden variation of temperature. But no such excitatory disturbance occurs if the rise of tempera- ture be not fluctuating, but gradual and continuous. I was able to secure this, by selecting at the beginning of the a. a i KK Ivy Holly Eucharis Fig. 123. After-effect of Cold on Ivy, Holly, and Eucharis Lily rt, The pormal response ; ^, response after subjection to freezing temperature for twenty-four hours. experiment, a suitable strength of current, such as to raise the temperature of the chamber continuously, at an approxi- mately uniform rate. Care had also to be taken that thermal radiation from the wire should not strike the specimen, since such radiation, as we shall see later, acts as a stimulus. The •^'^y 1 1 interposition of a sheet of mica, however, obviated this diffi- j I culty, mica being opaque to thermal radiation. While, under these conditions, the temperature was being raised, uniform vibrational stimuli were applied at intervals, and responses recorded, the temperature of the chamber at the moments of stimulation being carefully noted. In this way I obtained the following photographic record, with a petiole of Eucharis lily, affording a general idea of the effect of temperature on response. It will be seen that EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE l8i while the temperature was rising from 20° C. to 22° C. the ampHtude of the response was increased. After this, however, it fell rapidly in height with rise of temperature and became very small, at or near 60° C. On allowing the temperature to fall, however, the responses revived, with this peculiarity, that during the cooling, as compared with those given during the ascent of temperature, they were markedly en- hanced (fig. 124). kThe heating arrange- nents in this case were uch that the temperature /■as made to rise some- what rapidly. It will be noticed that response had ot here disappeared, yen at 65° C, though, s we shall see in the ext chapter, the death- int is at about 60° C. his apparent anomaly iS due to the fact that e plant, which is a bad onductor, was not al- wed time fully to attain e temperature of its urroundings. We shall ee that when the tem- erature is raised at a lower rate — about 1° C. n 1*5 minute — and when e specimen is not too thick, excitatory response disappears ith the approach of death, at a temperature very near 60° C. I give below a record of the effect of temperature arying from 30° C. to 50° C. on the response of the stem f Amaranth (fig. 125). In order to obtain perfect results, t is necessary that the specimen should not exhibit any Fig. 124. Photographic Record of Responses in Encharis Lily during Rise and Fall of Temperature Stimulus constant, applied at intervals of one minute. The temperature of plant- chamber gradually rose on starting current in the heating coil ; on breaking current, the temperature fell gradually. Tem- perature corresponding to each record is given below. Temperature rising : (i) 20°, (2) 20°, (3) 22°, (4) 38°, (5) 53°, (6) 60°, (7) 65° Temperature falling: (8) 60°, {9) 51°, (10) 45°, (11)40^(12)38°. 1 86 COMPARATIVE ELECTRO-PHYSIOLOGY fatigue, and I have found that this plant is but little subject to it. It will be noticed how the response is continuously depressed, as the temperature rises from 30° C. upwards. In this case, the thermal ascent took place at the rela- tively slow rate of 1° C. in i'5 minute, so that there was time for the plant to attain the temperature of its sur- roundings. A tabular statement is given below, showing the effect of temperature on amplitude of response, in two different specimens o{ Amaranth : Specimen I. Speci MEN XL Temperature Height of response Temperature Height of response 1 30OC. 50° 200 divisions 170 „ 95 M 72 „ 43 ,. 30° c. K 40° : 50° no divisions 90 40 25 ,. 10 The same fall in amplitude of response, when a certain point has been reached in the thermal ascent, to which I . have already referred, in the case of Eucharis lily, has also been noticed in that of muscle. From this it might be concluded that rise of temperature beyond 30° C. or so, induced depression of excitability. But here we are met by an anomaly. For growth, which I have SO" shown to be a phenomenon of excitatory response, in- creases, in the case of J V Fig. 125. Diminished Amplitude of the plant, throughout the Response with Rising Temperature, thermal ascent, up tO an (Stem of ^wara/zM) ' ^ optimum point at or near 35° C. Conductivity, again, which is, to a certain extent, correlated with excitability, undergoes enhancement with EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE 1 87 rising temperature. Thus in a certain specimen of Bio- phytuin^ for example, while the velocity of transmission at 30° C. was 37 mm. per second, it became enhanced to 9-1 mm. — that is, nearly three times — when the temperature was raised to 37° C. From these considerations, it would appear that the diminution in amplitude of electrical response, under a rising temperature below the rigor-point, might not always be due to a diminution of excitability, but to some other cause. IK In connection with this, it must be borne in mind that two factors are included in the process of response : namely, the external stimulus which induces contraction, or galvano- metrlc negativity, and the internal factor which brings about recovery. For I have already shown that whereas the action I of stimulus induces one effect — the contraction, for example, of pm excited tissue with galvanometric negativity — an increase ^ pf internal energy causes exactly the opposite— that is to say^ / U the expansion of the tissue and galvanometric positivity. ^ - • External stimulus and internal energy thus act antagonisti-^* cally. A steady rise of temperature causes, as we have seen,^^^^ (n increase of internal energy. Hence, increased energy^ Si^ ue to rise of temperature, enhancing the force of recovery I^L^^UJ^ lay cause a diminution of response, which is not due to diminution of excitability. 1^ The inference that it is the increased internal energy due ^Po rise of temperature which, by augmenting the force of recovery, diminishes the amplitude of response, appears the more probable from certain characteristics observed in the I autonomous pulsation of Desmodiuni gyrans. If rise of temperature increased the force of recovery, we should pxpect, conversely, that a fall of a few degrees would have the effect of diminishing this force of recovery, and con feequently enhancing the response. That this actually occurs \i\\\ be seen in fig. 126, in the first part of which is given a series of responses at the temperature of the room, which was 29° C. When the temperature of the plant-chamber 1 88 COMPARATIVE ELECTRO-PHYSIOLOGY to be diminished, since the amplitude of response was con- siderably enhanced. That the general excitability of the plant was not increased by the lowering of temperature, is seen from the fact that the frequency of pulsation was reduced on cooling to about two-thirds of its original value. The diminution of response with rising temperature may thus be due to an increase of internal energy, which tends Fig. 126. Photographic Records of Autonomous Pulsations in Des- modiuniy showing Increase of Amplitude and Decrease of Frequency, with Lowering of Temperature The pulsations to the left were recorded at the ordinary temperature of the room, 29° C. Those to the right, when the temperature had been lowered to 25° C. to cause antagonistic expansion and consequent galvano- metric positivity. This view finds support from the records seen in figs. 129 and 133, given in the next chapter. The first of these (fig. 129) shows the expansion, with consequent physical elongation, of the filament of Passiflora under a rising temperature. In the second (fig. 133) is seen the in- creasing galvanometric positivity of a specimen o{ Amaranth under similar circumstances. It is now clear that when the temperature of the tissue I EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE 1 89 is below a certain thermotonic minimum, the effect of a rise of temperature will be to enhance the amplitude of response by removing molecular sluggishness. This fact has been illustrated in the gradually heightened mechanical response of the autonomous pulsation of Desmodium gyrans when a plant artificially cooled was allowed to return to the normal temperature of the room (fig. 120). If similarly a plant tissue be first cooled and then allowed to return to the surrounding temperature, its electrical responses to suc- IIJKessive uniform stimuli being recorded throughout, a stair- ^:ase increase of response will be observed during the return. When the temperature, however, is raised above a certain l^fcptimum, a depression of the amplitude of response begins, not by the depression of excitability, but by the increasing force of recovery due to an augmentation of the internal factor. True depression only takes place when the plant IBb approaching a condition of heat-rigor. ■^ One very curious effect of temperature-variation which has been touched upon is the marked increase of sensi- tiveness which often makes its appearance as its after-effect. This is seen exemplified in the record given in fig. 124, showing the effect of a cyclic variation of temperature on Eucharis lily. In another experiment with Scotch kale^ the response at the temperature of 30° C. was eleven divisions, and at 50° C. eight divisions, during the thermal ascent. During the descent, however, the amplitude at 50° C. was sixteen, and at 30° C. twenty-three divisions. The sensitiveness was thus doubled. This enhancement _^nay be due in part to the increased molecular mobility IBlonsequent on the annealing effect, as it were, of temperature- variation. But it may also be regarded as partly due to the difference of the antagonistic forces which the excitatory response has to overcome during ascent and descent. During IB^e thermal ascent, the opposing expansive force is being rapidly accelerated. During the thermal descent, on the Ither hand, this is no longer the case, for the force of re- igo COMPARATIVE ELECTRO-PHYSIOLOGY When the temperature is raised above a certain critical point, the plant is killed, and its electrical response dis- appears at the same time. This is demonstrated visually in the accompanying photographic record (fig. 127). In this case, normal responses were first obtained at the usual tem- perature of the room. Steam was next introduced into the Before | After Fig. 127. Photographic record showing effect of Steam in abolishing Response The two records to the left exhibit normal response at 17° C. Sudden warming by steam induced at first an increase of response, but five minutes' exposure to steam killed the plant (carrot) and abolished the response. Vibrational stimulus of 30° applied at intervals of one minute ; vertical line = -I volt. plant-chamber, and kept streaming in during the course ot the experiment, electrical responses being recorded mean- while at intervals of one minute. It will be seen that at first a transitory augmentation ot excitability was induced. But this quickly disappeared, and in five minutes the plant was effectively killed, as is shown in the waning and final aboli- tion of response. This experiment affords us a qualitative demonstration of the abolition of response at death under EFFECT OF TEMPERATURE ON ELECTRICAL RESPONSE 191 the influence of high temperature. In the next chapter we shall enter upon an exact determination of this critical point of death. It is thus seen that temperature modifies the electrical response of plants. There is a temperature-minimum below which response is abolished. If the plant be kept too long at this temperature it is apt to be killed. In the case of a delicate species like Eucharis, which is highly susceptible of the injurious effect of cold, the electrical response is per- manently abolished by long exposure. But hardier plants, like Holly and Ivy, show revival of electrical response, on a return to a favourable temperature. The electrical response disappears also at a certain maximum temperature con- stituting the death-point. CHAPTER XVI THE ELECTRICAL SPASM OF DEATH Different post-mortem symptoms — Accurate methods for determination of death- point— Determination of death-point by abolition or reversal of normal elec- trical response — Determination of death-point by mechanical death-spasm — From thermo-mechanical inversion — By observation of electrical spasm : (a) in anisotropic organs : (<5) in radial organs — Simultaneous record of electrical inversion and reversal of normal electrical response — Remarkable consistency of results obtained by different methods —Tabulation of observations. It will be seen from the last chapter that there is in the case of every plant a certain high temperature which is critical, since above it life passes into death. Much difficulty has been experienced in the exact determination of this critical point, because no sure criterion of death was hitherto avail- able, such as would furnish an immediate and reliable indica- tion of its occurrence. The various symptoms of death, such as drooping, withering, discoloration and the escape of coloured cell-sap, do not manifest themselves at the onset of death, but at some time indeterminately later. Even when a plant has been subjected to a temperature in excess of the fatal degree, it continues to appear fresh and living ; and it is not till after some longer or shorter interval that the death symptoms are seen. To take, for example, the symptom of drooping, it is clear that the loss of turgidity on which this depends cannot at once make itself visible. In a thick tissue, again, death may take place in the superficial layers of the plant, the interior tissues, owing to feeble thermal conductivity, remain- ing comparatively unharmed. Or, if we employ the test of discoloration, which wc shall find to occur some time after the initiation of death we find that the exact moment at THE ELECTRICAL SPASM OF DEATH 1 93 which discoloration begins cannot be detected with sufficient precision. When we place the specimen in a thermal bath under a rising temperature, the beginning of discoloration after death is so slight as to be impossible of detection, and by the time it becomes marked, the temperature has already- passed several degrees above the fatal point. I have found, for example, that the colour of the milk-white style of Datura alba has changed to brown by the time that the temperature of the bath has risen to about 64° C. In the petals of Sesbania coccineum^ again, a striking change of colour is detected, under similar conditions. Rich crimson here turns into pale blue at a temperature of about 67° C. The fila- mentous corona of Passiflora quadrangularis^ finally, in which the filaments are barred by purple rings, loses its colour normally at about 68° C. In all these cases, the initiation of the loss of colour must have been imperceptible. Hence, all that can be determined from such experiments is that the death-changes must have commenced at some temperature lower than 64° to 68° C. I^b Before proceeding further, it is necessary to obtain a clear loea of what is meant by the death-point. In animals, an early symptom of death consists in the setting in of rigor mortis. But this does not synchronise throughout the body, certain parts of the organism undergoing the death- change earlier than others. Thus the only definition of the death-point which can be made at all precise is that which regards it as the point of initiation of some unmistakable sign of death. I shall next proceed to describe several death-symptoms and the modes by which they may be de- tected with certainty. With regard to such detection, I have pointed out else- where that, theoretically, it should be possible to make such a determination by watching the waning of some effect characteristic of the living condition, the death-point being known by its cessation at a given moment. Such a test, as we shall presently see, is afforded by the electrical responses. The ideally perfect method, however, would be by the II 194 COMPARATIVE ELECTRO-PHYSIOLOGY detection of some effect which at the moment of death under- went a sudden reversal to its opposite. There would not here be even that minor degree of uncertainty which is in- cidental to the determination of the exact vanishing-point of a waning effect. And such methods are afforded by my discovery of the occurrence of mechanical and electrical spasms at the moment of death. Turning now to the method of the waning effect, we have seen that response to stimulus by galvanometric negativity is distinctive of the living condition. When the plant is killed, this normal response disappears. At the moment of death from high temperature, therefore, we may expect to see the abolition of this normal excitatory response of negativity- . For this investigation I took a batch of six radishes. The specimens were kept for five minutes previous to each experiment in water at a definite temperature (say of 17° C), and were then mounted in the vibration-apparatus and their responses observed. Each specimen was next dis- mounted and replaced in the bath at a higher tempera- ture (say of 30° C.) for another five minutes. After this, a second set of responses, to the same stimulus as before, was taken. In this way observations were made with each plant, till the temperature at which response almost or altogether ceased was reached. I give below (p. 195) a table of the results obtained with the six radishes. From these experiments it would appear that in these cases the responses disappeared at about 55° C. It should be stated here that this investigation was carried out in the winter season in England, and it will be shown later that the incidence of cold has the effect of lowering the normal death-point by about 4° or 5° C. I was next desirous of substituting, for this method of] discontinuous observations, one which should be continuous. I, therefore, subjected the specimen — a stem of Amaranth — to a continuous rise of temperature, and took records of] responses to uniform stimuli after every few degrees of the ascent. 1 found here that not only was there a gradual THE ELECTRICAL SPASM OF DEATH 195 Table showing Effect of High Temperature in abolishing Response of Radish {Raphamis sativus) Specituen Temperature Galvanometrie response 100 divisions = 0*7 volt 53° c. ■r^<::\ 70 4 2 17° C. 53° C. 160 I 3 17° c. 50° c. 100 2 11' 17° c. 55° C. 80 . i 17= c. 5 60° c. 40 6 ^7°C. ^ 1 55° C. 60 decrease of response, tending towards its abolition, with rising temperature, but also that, at the death-point, it under- 40^ 50 58^ vv _ 60^ Fig. 128. Record of Electric Responses of Amai-anth at various temperatures The response undergoes reversal to positive at the critical temperature of 60° C. went actual reversal from the normal galvanometrie nega- tivity to positivity (fig. 128). This was due to the fact that o 2 I 196 COMPARATIVE ELECTRO-PHYSIOLOGY on reaching the death-point, the contained positive com- ponent in response was unmasked by the abolition of the true excitatory effect. But this positive response disappears also after a short time. It will thus be seen that by this method the death-point is capable of determination within very narrow limits, having been, in the present case, near 60° C. When the tissue is thin, this temperature soon proves fatal. But should it be thick, a very much longer exposure to it is necessary, if the interior of the tissue is to be killed effectively. I have also discovered another method of obtaining the death-point with precision, in the symptoms afforded by mechanical responses. For I found that a death-spasm occurs at a certain critical moment in a plant, which is analogous to the death-throe of the animal. The experimental plant — Mimosa^ for instance — was placed in a bath of water, whose temperature was being raised gradually, at a uniform rate of, say, 1° C. per one minute and a half, until the death-point was reached. During all this time there was no responsive fall of the leaf, for, we have seen, it is a sudden variation, and not a gradual rise of temperature, which acts as an excitatory stimulus. This gradual rise, on the other hand, increases the internal energy of the plant, by which the turgidity of the pulvinus is continuously augmented. In this process the increase of turgidity is more energetic in the more excitable lower half than in the upper. The greater expansion of the lower side of the pulvinus thus raises the leaf continuously. But immediately on reaching the death-point, there is a reversal of this movement, and an abrupt fall of the leaf. This spasmodic movement is sudden and well-defined. In \ a vigorous Mimosa the death-spasm is found to occur at or very near 60° C. This contraction of death is followed [after some time by d. post-mortem relaxation. That the death-response is an excitatory phenomenon is seen from the fact that any circumstance which lowers physiological activity lowers the death-point also. Thus, after a spell of cold weather, I found that the death-point i THE ELECTRICAL SPASM OF DEATH 197 of Mimosa was lowered from the normal 60° C. to about 53° C. This latter value, it will be remembered, was ap- proximately the same as that obtained with radish, in winter, by the method of electrical response. Again, 1 find fatigue to induce a lowering of the death-point, the extent of which depends upon the degree of fatigue. When this was moderate, I have found the death-point of Mimosa to be lowered to 6°C. This spasmodic contraction, indicative of the initiation of death, may express itself in diverse ways. For example, if the tubular peduncle of Allium be filled with water, and raised gradually in temperature, there comes a moment at which a sudden expulsion of the contained water occurs. A spiral tendril of Passijiora^ under the same circumstances, exhibits a sudden uncurling. The florets of the ray, in certain Compositce, show characteristic movements, either up or down. In all these cases alike, under normal circumstances, the death-point is found to be at or near 60° C. Turning next to the radial organs of ordinary plants, these also exhibit a sudden longitudinal contraction at the onset of death. I have shown elsewhere how, by means of the Morograph, an instrument which I devised for this purpose, a thermo-mechanical curve is recorded by the specimen, while it is being subjected to the continuous rise of temperature, culminating in the death-point. The ordinate of this curve represents the induced variation of length, and the abscissa the temperature. The expansion described in the case of Mimosa is seen here in the form of a gradual elongation, up to the moment of reaching the death-point. When this point is reached, however, a sudden contraction takes place, giving rise to an inversion of the curve. This turning point is very abrupt. The curve as a whole is thus one of life-and- death, in which the point of inversion separates the two. I give below a photographic record of this thermo-mechanical curve, obtained with the coronal filament of Passiflora. The death-point occurred here at 596° C. (fig. 129). The thermo-mechanical curve is very I 198 COMPARATIVE ELECTRO-PHYSIOLOGY similar in similar specimens under normal conditions. Fig. 1 30 gives two records of two different styles of Datura aibay obtained from flowers of the same plant. The death- point is seen to have occurred at 60° C. In recording the thermo-mechanical curve, there is found to be, normally speaking, a continuous expansion up to the death- point. In the case of vigorous specimens, in a good tonic condition, the inversion does not take place till about 59*6° or 60° C. But in less vigorous specimens, a certain hesitation, as it were, is seen to occur in the record at or near 55° C. With vigorous specimens, Fig. 129. Photographic Record of Thermo-mechanical Curve given by Coronal Filament of Passijlora The first or down part of th^ curve shows expansion, but on reaching death-point, at 59*6° C, there is a sudden inversion, due to spasmodic death-contraction. Fig. 130. Thermo-mechanical Curve of Two Different Specimens of Style of Datura alha, obtained from PMowers of the same Plant there may be the merest indication of this hesitation ; in other cases, with less favourable tonic condition, the hesitation is prolonged, but the expansion finally proceeds, and the death inversion takes place at the usual temperature of about 60° C. When the specimen, however, is enfeebled, or has been subjected to unfavourable circumstances, the point oi\ transient instability becomes fatal, and the inversion takes place there. As an example of what has just been referred to — namely, the influence of unfavourable external circum- II THE ELECTRICAL SPASM OF DEATH 1 99 stances in lowering the death-point — it may be mentioned here that the sudden incidence of cold weather will lower it by some 4° or 5° C. Intense fatigue will lower it by as much as 19° C. I have already said that this spasm, taking place at the moment of death, is an excitatory response. On this theory it occurred to me that it should also be possible to determine the onset of death by an electrical spasm. It may be well at this point, therefore, to examine some of the conditions under which such a spasm, supposing it to take place, might be displayed most conspicuously. We may suppose a radial org-an, with the usual electrical contacts, A and B, to have its temperature raised gradually up to the death-point. The excitatory effect of death may now be expected to cause the galvanometric negativity of a given point. But since these excitatory effects are equal and similar at A and B, they will balance each other, and there will be little or no resultant galvanometric response. In order to obtain a marked resultant effect, then, we must have an organ in which the excitabilities of the two points A and B are different. This difference of excitability, necessary to the exhibition of a resultant response, may be either natural or artificially induced. For the former, we may take a specimen which is not radial, but anisotropic, thus affording us two points of galvanometric contact, possessed of unequal excitabilities. We have seen that the inner surface of the petiole of Cucurbita maxima was more excitable than the outer. The same is true of the hollow peduncle of Uriclis lily. There is also a great difference of excitability as between the upper and lower sides of the scale of the bulb of the same lily, in the season of flowering, the concave surface of this scale being more excitable than the convex. Any of these specimens described I find to answer admirably for the purpose of this investigation. Taking the petiole of Cucurbita, then, I divided it longi- udinally, and rejected one-half, thus obtaining a half-tube, of which the inner concave surface was more excitable than I 200 COMPARATIVE ELECTRO-PHYSIOLOGY the outer convex. Electric contacts were now made through non-polarisable electrodes with equal and opposite areas on the two sides. These adjustments were made in a heating chamber containing electrical arrangements by which the temperature could be raised continuously. This was satisfactorily accomplished by an incandescent electrical lamp which was placed in a second chamber, vertically below the plant-chamber. There was a wooden partition between the two, by which the light of the lamp was excluded from the specimen (fig. 131). For the radiation itself will be **(]_) Fig. The Thermal Chamber Electric lamp in the lower compartment raises temperature of the upper. E, e', electrodes making contacts with the specimen ; t, thermometer. shown to constitute stimulus, and the object in the present case was to eliminate all exciting factors except death itself. By means of side-openings, the heated air was enabled to pass into the plant-chamber, thus raising the temperature. A rheostat included in the lamp-circuit made it possible to adjust the rate of this rise of temperature, its average being about 1° per minute. The natural current through the petiole is, under normal circumstances, from the less excitable outer to the more excitable inner surface: that is to say, the inner is galvano- THE ELECTRICAL SPASM OF DEATH 20I metrically positive. This is true when the excitation, due to section made for the purpose of preparation, has subsided. This stimulation, by causing greater excitation of the inner surface, is liable to induce there a temporary negativity. A gradual rise of temperature, as we saw, caused an increased turgidity of the more excitable lower side of the pulvinus of Mimosa, and this increased turgidity was exhibited mechanically by the erection of the leaf But the electrical - sign of increased turgidity is galvanometric positivity. We have also seen that electrical responses occur equally in motile and non-motile tissues. In the petiole of Cucurbita then, on its more excitable inner surface, we obtain, during the gradual rise of temperature, an increasing galvanometric positivity. This is true only, as has been said before, when the rise is continuous, and not marked by fluctuation. For any sudden variation will act as a stimulus, causing galvano- metric negativity of the more excitable inner side. For this reason it is necessary that the rheostatic resistance inter- posed in the lamp-circuit, for the adjustment of the uniform rate of rise of temperature, should be made at the beginning of the experiment, such tissue being very sensitive to this particular stimulatory action. At the commencement of my investigation I experienced much trouble from the erratic movement of the galvanometer spot of light, and the obtaining of a steady electrical curve seemed at that time almost hopeless. Later on, however, I found that these fluctuations were traceable to temperature-variations, unavoid- ably associated with the attempt to regulate the rise of temperature by movement of the rheostatic slide. It is for this reason, then, that the adjustment must be made, once for all, at the beginning. Carrying out the experiment in this manner, I obtained, with various anisotropic organs, a sudden inversion of the electric curve at the death-point. This death-point was found, in all vigorous specimens, from which traces of injury had been removed by previous rest, to occur accurately at 59*6° or 60° C. In these electrical curves, the same I 202 COMPARATIVE ELECTRO-PHYSIOLOGY point of instability, already noticed in the thermo-mechanical curve, was often found to occur at or about 55° C. And if the specimen were not in favourable tonic condition, or had been suffering from injury, the death-point was lowered to this degree. I give below an electrical curve showing the point of inversion at death (fig. 132). It was obtained with the sheathing petiole of Musa. The inner or concave side of this petiole is more excitable, as we have seen, than the outer. These responsive electrical variations were very large, and could not be represented within the limits of the photographic plate. I therefore took a photographic record between the temperature of 54° C. and 6f C. only. The first part of the curve represents the increasing galvanometric posi- tivity of the more excitable inner surface of the specimen. The same process of increasing posi- tivity under the continuous rise of temperature, had been going on previously, it is to be understood, before arrival at 54° C, at which the photographic record was com- menced. This increasing galvano- metric positivity corresponds to the gradual erection of the leaf in Mimosa^ and to the expansion of a radial organ, such as a coronal filament of Passiflora, all alike being due to the positive variation of turgidity. In order to give an indication of the particular temperature at each portion of the curve, the re- cording light was obscured for about 1 5 seconds after each degree of temperature. The successive gaps, then, are one Fig. 132. Photographic Record exhibiting Electric Spasm in the Petiole of Musa Sudden electric inversion takes place at the death-point, 59*5°. Record was com- menced at 54° C, and suc- cessive gaps in the record indicate 1° C. rise of tem- perature. THE ELECTRICAL SPASM OF DEATPI 203 degree centigrade of temperature apart. These interruptions, however, were not made after the occurrence of the inversion. As soon as the death-point was reached, in the present case at 59'5° C, there was a sudden inversion of the electrical curve (fig. 132), corresponding with the point of inversion of the thermo-mechanical curve (fig. 129). Each of these curves is seen to bear a striking resemblance to the other. In both cases, the inversion was due to the same fact of sudden excitation, finding expression in the one, in induced galvanometric negativity, and in the other, in mechanical Vntraction. In the case of organs which are more or less radial, and in which there is little differential excitability, it is necessary to abolish the excitability of one contact, as, say, by previous scalding. For this experiment I took a leaf of Amaranth, and injured a portion of the lamina by immersion in boiling water. The two contacts were made, one with the petiole, and the other with the injured lamina. On raising the temperature continuously, the more excitable petiole became increasingly positive. The photo- graphic record in this case was com- menced only on reaching 55° C, and the death-inversion took place at ^^'^J^^^:^^^^^. 59*5° C. (fig. 133). version at Death-point, We have already seen that, besides Amaranth this electrical reversal, there is another means of detection of the death-point, afforded by the con- version of the response to external stimulus from the normal negative into positive. Now it occurred to me that it would be interesting, if in the same specimen, both these tests could be applied at the same time. We could then see whether two methods so independent of each other furnished mutual corroboration or not. For this purpose I took a stem of I 204 COMPARATIVE ELECTRO-PIIVSIOLOGY Amaranth, and abolished the excitability of one of the two contacts — a lateral leaf — by scalding. The electrical curve, under a continuously rising temperature, was now taken in the usual manner. The existing electro-motive difference between living and injured contacts underwent the usual 40" 50 H 55 H i 59 I. ,'58 57" Fig. 134. Record showing Inversion of Electric Curve (represented by dotted line) and Simultaneous Reversal of Electric Response in Stein of Amaranth \ indicates current of injury from injured to uninjured contact, which, reaching a maximum, undergoes reversal at death-point. Normal response up, also becomes reversed to down after death-point. increase, reaching a maximum at the death-point. Meanwhile electrical responses to uniform vibrational stimuli were taken at intervals a few degrees apart. It will be seen (fig. 134) that the electrical inversion took place at 57° C, this moderate lowering of the death-point being due, in all probability, to the slight depression caused by scalding at THE ELECTRICAL SPASM OF DEATH 205 the distal contact, which had not had time to pass off. The test-responses to uniform mechanical stimulation which were being taken meanwhile show a continuous diminution towards the death-point. When this, however, had been passed, the response is seen to be reversed in direction to positive. As has been said before, this positive response also disappears after a time. We thus obtain a very striking demonstration of the fact that the reversal of the electrical curve and the reversal of the sign of response are concomitant. VSk It may be mentioned here, in anticipation of a future chapter, that the death-point may also be obtained by the Iudden inversion of the curve of electrical resistivity, and hat the value obtained in this way coincides with those Iready given. I give below a table showing the death-points deter- nined by various methods : Table showing Death-points Obtained by Different Methods Specimens Method Death- point I. Flower of French Marigold . Opening or closing of flower C. 59° 2. Peduncle of Allium Expulsion of contained water 59° 3- Spiral tendril of Passijlora Movement of uncurling 59° 4- Pulvinus of Mitnosa Spasmodic lateral movement 59°-6o° 5-8. Style of Datura (four speci mens). Each gave . Morograph 60° 9-12. Style of Hibiscus (four speci mens). Each gave . )> ... 60° 13-18. Coronal filament of Passiflorc (six specimens). Each gave ' ". . 60° Anisotropic organs Electric inversion 19. Petiole of Cauliflower . >> 5J • • 60° 20. Scale of Uriclis bulb J> }> • • 60° ' 21. Petiole of Musa . 3> }> • • 60° 22. Radial organ : petiole f Amaranth J> JJ • • 59-6° 23- VetiolG of Amaranth Reversal of sign of electrical response 59-6° 24. Style of Hibiscus . Resistivity variation . 60° It is thus seen that employing different methods and using plant-organs which are equally diverse — flowers, bulbs, petioles, and others — a death-point is determined which II 206 COMPARATIVE ELECTRO-PHYSIOLOGY is very definite and practically the same for all phanero- gamous plants. Among the mechanical methods, that which depends on the thermo-mechanical curve, giving the death-point by a sudden inversion, is specially accurate. Of an equally precise character is the death-point obtained by the inversion of an electrical curve, and also that which is given by reversal of the electrical response. And it is in the highest degree remarkable that the points of inversion in the mechanical and electrical curves respectively, together with the point of reversal of the electrical response itself, should so exactly coincide. CHAPTER XVII MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE Repeated responses under single strong stimulus — Multiple mechanical response in Biophyttim — Multiple electrical responses in various animal and vegetable tissues — Continuity of multiple and autonomous response — Transition from multiple response to autonomous, and vice versa — Autonomous mechanical response of Desmoduim gyrans and its time-relations — Simultaneous mechanical and electrical records of automatic pulsations in Desmodium — Double electrical pulsation, principal and subsidiary waves — Electrical pulsation of Desmoduim leaflet under physical restraint — Growth-pulsation — So-called current of rest in growing plants. E have seen that when a plant organ is acted on by a iingle stimulus of sufficient intensity, it exhibits a single sxcitatory effect, which may show itself in two independent ^ays, as mechanical and electrical response. We have also feen that part of the impinging stimulus may become latent, \o find appropriate expression later. It was also shown lat with increasing intensity of stimulus the amplitude of response reaches a limit. It may thus happen that a very ^strong stimulus, not finding adequate expression in a single response, will exhibit itself by means of repeated responses. 'he incident energy in such cases is held latent for a time,^ to manifest itself later in a rhythmic manner. I have been able to demonstrate the occurrence of this lultiple excitation, in response to a single strong stimulus, >y several different and independent methods. The simplest md most striking of these depends on the recording of the lotile effects in the leaflets oi Biophytum. In fig. 135 are seen 10 less than sixteen multiple pulsations resulting from a single strong thermal stimulation of the petiole bearing the leaflets. ^ For more detailed account see Bose, Plant Response ^ pp. 279-357. 208 COMPARATIVE ELECTRO-PHYSIOLOGY The average period of each pulsation is here about thirty seconds ; but this may vary in different cases from half of this to one minute. 1 have also been able to detect these multiple excitatory waves, during their transit through non-motile con- ducting tissues such as stems. The imperceptible volumetric changes which occur on the arrival of excitation were here detected electrically by variations of pressure induced in an enclosing microphonic contact. In fig. 136 is seen such Fig. 135. Multiple Mechanical Response of BiophyluiHy due to a Single Strong Thermal Stimulus multiple electro-tactile response In the stem of Mimosa^ due to a single thermal stimulus. 1 have also been able to record such multiple excitatory effects by means of electro-motive response. In fig. 137 is seen a photographic record of a series of such responses, given by the leaf of Biophytmn, the individual thermal stimuli being here applied at intervals of five minutes. It will here be noticed that each single stimulus gave rise to from five to eight responses, the average period of which was thirty seconds. From the corresponding mechanical MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 209 responses, it will be remembered that the average period of these had also this value. These multiple responses to a single strong stimulus, while very strikingly manifested by such plant-tissues as the pul- vinules of Biophytum, and in the animal, by the cardiac tissue, are also exhibited by almost all kinds of tissues under favourable circumstances. In fig. 138 will be seen records which show this in the case of different vegetable organs under diverse forms of stimu- , . ^ r • • ^^^- 136- Multiple Electro-tactile lation. In tig. 139 is given Response in Stem of Mimosa, IK series of multiple responses stimulus ^'"^^^ ^^'°"^ Thermal which I have obtained from (OrTgLTl record reduced to 1 ) frog's stomach. I have also detected the occurrence of multiple responses in nerves of animals, which will be described in a later chapter. Fig. 137. Photographic Record of Multiple Electrical Response in Leaf of Biophytum First series of eight responses to a single thermal stimulus ; second stimulus, after interval of five minutes, evoked five responses ; third stimulus, after second interval of five minutes, gave six responses. Average period of each response, thirty seconds nearly. We have seen that multiple response takes place in msequence of some sufficient increase of internal energy. P 2IO COMPARATIVE ELECTRO-PHYSIOLOGY In the cases mentioned, this was derived from impinging stimulus. The internal energy of a tissue may, however, be Fig. 1 38. Multiple Electrical Responses under Different Forms of Stimulus in Different Organs {a) In Mimosa due to thermal, and [b) to chemical stimulation ; (r) in peduncle of Hiophytum^ due to thermal stimulus ; [N.B.— This series persisted for two hours.] {d) in hypocotyl of Tamarindus indica^ due to stimulus of cut. increased, and multiple response may consequently be initiated in other ways, as, for instance, by adequately raising the temperature of the plant. This is seen in the following record of pulsatory responses as induced in a young leaflet of Biophytum, when the temperature was raised to 35° C. With the increase of internal energy, the turgidity of the tissue was en- hanced, and the excessive hydro- static tension thus brought about induced autonomous pulsations (fig. 140), as in a quiescent snail's heart similar pulsations are in- duced by increase of the internal hydrostatic pressure. I have else- where shown that the energy which expresses itself in pulsatory movements may be derived by the plant, either directly from immediate external sources ; or from an excess of such Fig. 139. Photographic Record of Multiple Electrical Re- sponse to Single Thermal Shock in Frog's Stomach MULTIPLK AND AUTONOMOUS ELECTRICAL RESPONSE ail e!ieri:j\' already accumulated aud held latent in the tissue^ nded by the incidence of external stimulus; or from the \cessive accumulation of such latent eneri^y alone. There IS thus a continuity between multiple anil autonomically responding; plants. Biaphytum^ which mulcM i>rdinary cir- cuinst.uu c-s brKMu;s ti> the former of tluv^c^ rl.iNses, becomes converted into the latter under e\ic'i>tion.dly favourable Inic conditions. That is to s;n\ it le.spoiuls hv a single ■"'° '° " """ ■ * ~ '• " tFio, 14a Induction of Autonomons Response in Bi$^i^yh$m Al Mixierately Hiub TdnjvDivtre of 35® C. Noic ihc diminution of amplitiK^o . i k [vnsr with falling tcmpemiure. The pulsalions -p tuK.u .'.)■■ (\ ipoiisfs to A stuMii; stimulus. Under cxrcniuMi.dly favour- Ic tonic conditions, however, it cxhihi : hk. us or tonomous responses. Ar^ -•'■-- u otlui h.nul lich ortlinarily exhibits .^ isc, will. ihuUm favourable circumstances, cease to exhibit spont.mc c)us movements. It then exhibits a single resjxMisc t(> .1 in;'1(« Ioderate stimulus, and multiple .responses lu a :>iu^lc .strong hnulus : I When the leaflet of this plant, owinir t.> iK t'u it .f inic mal jcrgy, is in a state of standstill, a renewal oi tlu> iipi^U of iniulus will restore it to a condition ol" autonomous u spiMisi llis is seen in the followint^ record (fig, 141) ot tlu u jnn ( 212 COMPARATIVE ELECTRO-PHYSIOLOGY of the leaflet of Desmodium. The leaflet was in a quiescent condition, but under the action of stimulus of light, it ex- hibited multiple responses ; and these, owing to the increasing absorption of energy, showed a staircase enhancement of amplitude. On the cessation of light, the energy absorbed maintained the pulsation for some time. It is thus the absorption of energy which is the cause of the so-called autonomous movements. The energy, as already stated, may be derived by the plant either directly from ex- ternal sources ; or from the excess already accumulated and held latent in the tissue, aided by incident external stimulus ; or from an excess of latent energy previously accumulated. Fig. 141. Initiation of Multiple Response in Lateral Leaflet of Desmodium originally at Standstill Light applied at x and continued till the end of the sixth response, as shown by the thick line. The responses show a staircase increase with increase of absorbed energy. Pulsations persist for a short time even on the cessation of stimulus. It would be impossible to conceive of movement without an exciting cause. Only under the action of stimulus can a living tissue give responsive indications. An ex- ternal stimulus may either give rise to an immediate responsive expression, or be partly or wholly reserved in latent form for subsequent manifestation. * Inner stimuli ' are simply external stimuli previously absorbed and held latent. A plant or animal is thus an accumulator which is constantly storing up energy from external sources, and numerous manifestations of life — often periodic in their character— are but responsive expressions of energy which has been derived from external sources and is held latent in the tissue. MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 213 M '\ f\ M In Desmodium gyrans, as is well known, we have the typical example of autonomous response, its secondary leaflets executing periodic up and down or elliptical movements. The movement of the leaflet in some instances takes place by jerks, in others it is more uniform. The period of a com- plete up-and-down movement varies between two and four minutes. The length of this period is much affected by temperature, being less when this is moderately high. From the normal, or highest position, the leaflet sinks somewhat rapidly ; having reached its maximum depressed position, it iHrirests for a while. There l^fes next a rather slow I^Hise to its original posi IHtion. This up-and-down motion is in some cases approximately straight. n others, the pulvinule f the leaflet is slightly isted after its descent, nd the corresponding rve described becomes ore circular. In view of certain culiarities to be ob- rved in the electrical sponse of DesmodiuiUy is necessary here to enter into some detail regarding the me-relations of its mechanical response during the two hases of down and up movements. The great difficulty recording the pulsatory movements of Desmodium lies the extreme slenderness of the lateral leaflets. This such that the friction of a light recording-lever against e recording surface is sufficient to bring these movements a stop. This difficulty has, however, been overcome, as ated elsewhere, by means of the Optical Lever.^ Fig. 142 ii i/ y y y y n m Fig. 142. Photographic Record of Autonomous Mechanical Pulsation in Desmodium Leaflet Period of each complete pulsation = 27 minutes. ' See also Bose, Plant Response^ p. 5. 214 COMPARATIVE ELECTRO-PHYSIOLOGY gives a photographic record of a series of autonomous pulsations exhibited by a leaflet of Desrnodium. For the accurate observation of the rate of movement of the Desmodium leaflet during its different phases I have also been able to make records by means of a series of punctures produced by electrical sparks on a recording- surface. The sparks occur at the short gap between the end of the recording arm of a very light aluminium lever and the drum, these being connected respectively with the two electrodes of a RuhmkorfPs coil. The electrical dis- turbance does not afiect the plant, as the pulsating leaflet is separated from the other arm of the lever by a long silk thread. The primary current in the Ruhmkorff's coil is broken at intervals of five seconds. Hence succes- sive punctures in the record represent intervals of five seconds each. I give here (fig. 143) a record obtained in this manner, of a single mechanical pulsation of a leaflet of Desmodium. It is to be understood that the up-movement in the record represents the down-movement of the leaflet. These movements are produced by excitatory con- tractions of the lower and upper halves of the pulvinus alternately. An inspection of the record given shows that after a pause in the highest position a sudden excitatory impulse is developed in the lower half, which is gradually exhausted as the lowest position is reached. The maximum rate of movement to which this excitation gives rise is in this particular case 7 mm. per second. After the lowest position is attained there is a pause. The up-movement Fig. 143. Spark-record of Single Pulsation in Leaflet of Desmodium Showing lime-relations of down- and up-movements in single pulsation of leaflet of Desmodium. Up- curve represents down-movement of leaflet and vice versa. In- terval between successive dots = 5 seconds. MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 215 then takes place more gradually and at a much slower rate. This movement is due to natural recovery, aided by a moderate excitatory contraction of the upper half of the pulvinus. 1 give herewith a table showing the characteristic rates of movement in the different phases of the entire pulsation. Table showing Rates of Movement at Different Stages of Pulsation in Desmodium. Down-movement Up-movement Total period . . 45 seconds Average rate . '61 mm. per second Maximum rate . 7 ,, ,, ,, Duration of pause . 40 seconds Total period . . 70 seconds Average rate . -4 mm. per second Maximum rate . '5 ,, ,, ,, Duration of pause . 35 seconds Several facts are brought out in this table which are of jpecial importance, and first we observe that the excitatory impulse which causes the down-movement is brief and quickly ;xhausted. This is seen by the great distance covered luring the first ten seconds, after which the movement gradually slows down. This indicates a short-lived impulsive Lction, the subsequent movement of the leaflet being mainly lue to inertia. There is then a pause in the down-position, after ^hich the up-movement commences. It will be noticed here that this movement is more gradual and prolonged than the lown-movement. From the indications given by these :haracteristic movements, we may conclude that the excita- )ry reaction by which the down-movement is caused is relatively more intense and more quickly exhausted than lat which brings about the up-movement. We may gauge ^he relative intensities of the two impulses approximately, jither from the maximum or the average rates of the down ind up motions. The former gives the ratio of -'-=r4 ; the 5 tter gives =1*52, The intensity of the downward Impulse may therefore be taken to be roughly one and a-half :imes as great as that which occasions the up-movement. 2l6 COMPARATIVE ELECTRO-PHYSIOLOGY The total duration of the down-movement is again much less than that of the up. We have seen that a single excitation has a single con- comitant electrical pulsation. We have also seen the multiple electrical responses corresponding to multiple excitations. It remains, then, to find out whether autonomous pulsations have any electrical concomitant, and if so, of what nature. I shall here, therefore, describe experiments for the recording of the electrical pulsation of the Desmodium leaflet. For this purpose I selected specimens in which the movement of the leaflet was not spasmodic, but gradual and continuous, and where the up and down movements were approximately in a straight line. In order to obtain those responsive electro-motive changes which might accompany the automatic movements of the leaflets, it was necessary to make one of the electric contacts at a point on the tissue which was free from excitation, the second contact being made at a place where the excitatory reaction was at its maximum. I have shown elsewhere ^ that the seat of autonomous excitation in Desmodium is neither central nor peripheral, but localised at the slender pulvinated joints to which the leaflets are attached, the latter thus serving merely as indicating flags. Acting on these considerations, I made one contact with the slender pulvinule of a lateral leaflet, the other being made with the common petiole. These electrical connections were made securely by means of cotton threads moistened with normal saline solution, and attached to non-polarisable electrodes. The electro-motive variations induced in the plant now gave rise to correspond- ing deflections in the galvanometer in circuit. On taking records of these electrical responses, I was surprised to find that, corresponding with each complete mechanical vibration, there was a double electrical pulsation — a large principal followed by a smaller subsidiary wave. In a given case, where the period of the complete mechanical vibration was about 35 minutes, the period of the principal of these two ' Bose, Plant R espouse ^ p. 299. MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 21/ waves of electrical response was slightly less than i minute, and that of the subsidiary wave a little over 2*5 minutes. These double electrical pulses, corresponding to a single mechanical vibration, are at first very puzzling, and I under- took special investigations to ascertain the reason of this peculiarity. In order to obtain an insight into the relation between these mechanical and electrical responses it was necessary to take simultaneous records of the two on the same recording drum. This was accomplished by having the two recording spots of light — one from the galvanometer ,nd one from the optic lever — thrown on the same horizontal lit, in front of the revolving drum, round which was wrapped ; sensitive photographic film. The galvanometer spot of ight, and consequently the electrical response record, was he lower of the two. The vertical movement of the spot of ight which records the mechanical response is to be under- stood as converted into horizontal by reflection from a second mirror suitably inclined. This experimental arrange- ment is similar to that employed for simultaneous mechanical d electrical records in the case of Mimosa, as shown in g. 12. The record given in fig. 144 exhibits the simultaneous echanical and electrical responses thus obtained. It will seen that the minor electrical wave took place while the aflet was moving up from (a) and com.ing to its highest sition at (^). This was followed by a wave of higher mplitude but shorter period, which coincided with the ovement of the leaflet again from its highest to its lowest sitions. It will thus be seen that the subsidiary electrical ave of small amplitude and relatively long period coincided ith the slow up- movement of the leaflet, and that the rincipal wave, characterised by large amplitude and short period, corresponded with the quick down-movement of the iiaflet. These galvanometric deflections indicated, it must be nderstood, a condition of galvanometric negativity of the ulvinule at the moments of its excitatory up and down 2l8 COMPARATIVE ELECTRO-PHYSIOLOGY understand why two electrical waves correspond to one mechanical pulsation. First, we know that an excitatory- change at a given point will have, as its concomitant, an electro-motive variation of galvanometric negativity. Second, the intensity of this electro- motive variation depends on the intensity of the excitatory change. And lastly, on the cessation of excitation there is an elec- trical recovery. Now we have seen from the spark-record (fig. 143) that the leaflet during one complete mechanical pul- sation is subjected to two excitatory impulses, occur- ring in the upper and lower halves of its pul- vinule alternately. It is these two excitations which give rise to the two electrical disturbances of galvanometric negativity. And the different ampli- tude and period in the two cases are fully accounted for by the different period and intensity of the two excitatory impulses. It will be remembered that the excitatory impulse which produced the up-movement was the feebler and more protracted of the two. It is consequently attended by an electrical disturbance of moderate intensity and correspond- ing persistence. On the cessation of the up-movement, as we have seen, there is a pause, and during this time we find that Fig. 144. Photographic Records of Simultaneous Mechanical and Elec- trical Pulsation of Desmodium Leaflet a b (upper figure) represents up-movement of leaflet ; a b (lower figure) corre- sponding electrical subsidiary wave ; b a' (upper figure) down-movement of leaflet ; b a' (lower figure) corre- sponding principal electrical wave. MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 219 electrical recovery takes place ; but before this is complete, and while there is still a certain residual galvanometric negativity, there occurs that more intense and short-lived excitatory reaction which finds mechanical expression in the downward-movement. The same reaction finds electrical expression in a brief and intense response of galvanometric negativity ; on the expenditure of this excitatory impulse there is again an electrical recovery, which becomes prac- tically complete. In the instance given in the spark-record, it was found :hat the relative intensity of the down to the up impulse was Lpproximately as 1*5 is to i. And it is interesting to see that in the photographic record of the electrical responses [fig. 144), the ratio of the amplitudes of the corresponding ilectrical waves is also the same. In other instances, the relative intensity of the principal wave is still higher. I give )elow two tables showing the absolute values of the electro- lotive variations in two different cases. Table I. Table U. I Number of obser- vation E. M. variation. Principal wave E.M. variation. Subsidiary wave I •0014 TOlt •0013 „ •0014 „ •00055 volt 2 3 •00051 „ •00054 „ 4 •001 5 „ •00054 „ 5 •0016 „ — f^"™!*' iE.M. variation. E.M. variation. Subsidiary wave I -0024 volt •0016 volt 2 -0025 „ •0015 „ 3 -0025 „ •0016 „ 4 -0025 „ •0017 „ 5 j -0026 „ — It might be thought that these two electrical waves had ►een induced by the mechanical movement of the leaflet as such. We have seen, however, that the electrical response is a concomitant of the excitatory condition, whether such Kxcitation be followed by any mechanical response or not. 'his we saw in the absence of mechanical movement in the 220 COMPARATIVE ELECTRO-PHYSIOLOGY case of sensitive plants when responsive mechanical move- ments were prevented from taking place by physical restraint (p. 20). T he mechanical an d elec trical respons es are thus independent nio des of expression of a singl e fund a- mental excita tory process. In order to demonstrate this in the case of the autonomous pulsation of Desmodiuin, I first obtained simultaneous mechanical and electrical responses of Fig. 145. Photographic Record of Simultaneous Mechanical and Electrical Pulsation in Leaflet of Desmodium, before and after Physical Restraint of Leaflet. The first part of this record shows both mechanical and electrical pulsa- tion. In the second part, leaflet was physically restrained, as seen in the mechanical record, becoming horizontal. Electrical pulsation now seen to persist with even greater vigour than before. the leaflet (fig. 145). In the next part of the same record the mechanical movement of the leaflet was restrained, as seen in the upper mechanical record, which here becomes a straight line. But the lower record, which gives the electrical re- sponse, still shows the double electrical pulsation unimpeded. Indeed, so far from the mechanical response having been the cause of the electrical, we find that on its arrest, at least in this particular case, the latter becomes very MULTIPLE AND AUTONOMOUS ELECTRICAL RESPONSE 221 much enhanced. In fact, it appears as if the fundamental excitatory reaction, being now deprived of one of its two modes of expression, exhibited the other with the greater energy. We thus see that not only does the electrical response give us a means of detecting the action of external stimulus on a tissue, but that the same mode of indication enables us further to demon- strate the existence of those internal ex- citations which may find mechanical ex- pression in the so- [called * autonomous ' ovements. One such autono- ous pulsation pre- lent in all plants is ;hat of growth, and jby means of the Ihighly magnified re- ^cord given by the Crescograph. I have shown this to consist f the additive effects of multiple minute pulsatory movements. e see this in fig. 146, which gives a series of records of multiple growth responses obtained with the peduncle of Crocus at different times of ;he day. In the present case, the average period of each lulsation is twenty seconds. In the case of relatively slow pulsations like these, if one electrical connection be made with the growing-point, where such movements are in pro- w I Fig. 146. Crescographic Record of Multiple Growth-responses in Peduncle of Crocus The ordinate represents the extent of responsive elongations in mm. ; the abscissa, time in seconds. 222 COMPARATIVE ELECTRO-PHYSIOLOGY ceased, the galvanometer-spot of light is thrown into a state of oscillation, indicative of the local excitatory reactions at the growing-point. These multiple pulsations of growth consist of alternating po sitive and negative turgidity-variations. In dealing with the concomitant electrical response, however, we have seen (p. 64) that the induced galvanometric negativity, owing to its greater intensity, always overpowers the galvanometric positivity, if the two occur in rapid succession. In growth- pulsation, the constituent pulses are often extremely rapid. Hence a growing-point may be expected to exhibit, galvano- metrically, a resultant negativity. This consideration may explain the observation of Johannes Miiller-Hetlingen, other- wise unexplained, that the growing-points of both shoot and root, in the seedling of Pisum sativum^ are^ negative, as compared with the indifferent cotyledons. CHAPTER XVIIl RESPONSE OF LEAVES Observations of Burdon Sanderson on leaf- response in Diofuea—'Lea.f-Sind- stalk currents — Their opposite variations under stimulus — Similar leaf-and- stalk currents shown to exist in ordinary leaf of Fzcus ixligiosa — Opposite- directioned currents in Citrus dectunatia — True explanation of these resting- currents and their variations — Electrical effect of section of petiole on DioncEa and Ficus religiosa — Fundamental experiment of Burdon Sanderson on lamina of Dionaa — Subsequent results — Experimental arrangement with symmetrical contacts— Parallel experiments on sheathing leaf of Mtisa — Explanation of various results. It was pointed out in Chapter 11. that progress in the in- vestigation of the subject of excitatory phenomena in plants Iiad been long delayed, in consequence of the prevalent idea hat only motile plant-organs were ' excitable.' The atten- ion of investigators was thus mainly confined within the larrow range of the so-called * sensitive ' plants, such as DioncBa. It was also shown, in the same place, that the i'esults already arrived at by observers in this field had not been altogether concordant, and presented many anomalies. As it has now been demonstrated, however, in the course if previous chapters, that ordinary plants are fully sensitive, it ill be well to proceed to show that the various effects observed in the 'sensitive' Dioncsa may be still better studied in ordinary leaves. It will be possible, moreover, by follow- ing this line of inquiry, to determine those general laws, of 'hich the peculiarities observed in Dioncea are only instances ; ind thus we shall be the better able to offer an explanation ►f such cases as now appear anomalous. Before doing this, I shall briefly recapitulate the principal iffects observed by Burdon Sanderson in the leaf of Dioncea. 224 COMPARATIVE ELECTRO-PHYSIOLOGY These observations relate firstly to the existing current of rest in the petiole and midrib, and the variations of this resting-current, whether under excitation of the lamina, or by section of the petiole, or again, by the action of electro- tonus; and secondly, to the induction of variations of a transverse current between the upper and lower surfaces, of the lamina. As regards the current in the petiole and its prolongation the midrib, which I shall distinguish as ' the longitudinal petiolar current,' Burdon Sanderson found this 5■ < R ^ e ^y A ==^!ik= ^ -^ ^> N -R > ■"^- R <: Fig. 147. Natural and Responsive Currents in Leaves {a) Leaf-Jand stalk-currents in Dioncra. Natural current, n, flows out- wards <- A->, s being stalk-current, and l leaf-current. Stimulation of lamina at x gives rise to responsive current, R, from right to left, inducing negative variation of leaf-current and positive variation of stalk-current. When stimulus, however, is applied on left at x , responsive current, R, is from left to right, inducing effects exactly opposite of former, viz. negative variation of stalk-current and positive variation of leaf-current ; {b) Leaf- and stalk-currents in Ficits nli- giosa similar to those of Dionaa. Natural current flows outwards <-A->. Stimulation of lamina at x gives rise to responsive current, R, inducing negative variation of leaf- and positive variation of stalk- currents ; (f) Leaf- and stalk-currents of Citrus decumana, opposite to those of Ficus and Diomea, -> A *-. Stimulation at x induces positive variation of leaf and negative variation of stalk-currents. to flow in the midrib, from the end proximal to the stalk to the distal end. This he designated as the ' normal leaf- current' He further found that if electrical connections were made, so that one contact was near the lamina, and the other away from it, the stalk-current was opposite in direction to the leaf-current (fig. 147 (a) ). On stimulation of the lamina, these resting leaf-and -stalk currents were found to undergo responsive variations. But these changes were exactly opposite to each other. That is to say, the leaf-current underwent a negative, and the stalk- RESPONSE OF LEAVES 225 tf current a positive, variation. No explanation has as yet been offered, regarding either the existence of these opposite- directioned currents of rest, or the apparently anomalous result, that an identical stimulus would induce, in one case a negative, and in the other a positive, variation of them. As regards these peculiar currents of rest we have seen (p. 176), that if an intermediate point be physio- logically less excitable than either of the two terminal points, then a resting current will flow from the less to the more excitable. This is the particular current-distribution in the leaf of Dioncea. It is not a unique phenomenon, for I ave noticed other such instances in ordinary leaves. The int of junction of the petiole with the lamina of Ficus religiosa, for example, is galvanometrically the most negative point in that petiole-and-midrib. The currents here also, then, as in the case of Dioncea^ flow outwards from the point f junction — the leaf-current towards the tip of the leaf, and e stalk-current in the opposite direction (fig. 147 (J?)). We also saw, however, in the same place, that there may instances in which an intermediate point is more excitable an either of the two terminal. When this is so, the rrents of rest will be reversed in direction,, and flow wards. This I find to be the case in the leaf of Citrus xumana (fig. 147 ic) ). Next with regard to the excitatory variation of these sting-currents in leaf and stalk, we must remember at the e ffect of stimulation is to give rise to a t rue citatory current, flow ing away from the excited. If then lere be already a resting-current, the responsive current will added to this algebraically. When the lamina to the [ght is excited, the responsive current flows from right to ift. This would naturally, in the case of Dioncea^ induce a fegative variation of the leaf-current, and a positive variation the stalk-current (fig. 147 {a)). The same thing is seen m stimulating the lamina of Ficus religiosa, where also the excitatory current, being of opposite sign to the leaf-current, id of the same sign as the stalk-current, induces a negative Q 226 COMPARATIVE ELECTRO-PHYSIOLOGY variation of the former, and positive variation of the latter (fig. 147 (d)). The same stimulus thus induces effects which are apparently opposite. Or an interesting variation of the phenomenon may be obtained, on repeating the experiment with the leaf of Citrus. Here, on stimulating the lamina, we observe a positive variation of the leaf-current, and a negative variation of the stalk-current (fig. 146 {c) ). This is because the currents of reference or resting-currents are the opposite of those in Dioncea and Ficus religiosa. Another series of variations exactly the reverse of these, and therefore at first sight anomalous, is caused by simply changing the point of application of stimulus, from the right end on the lamina, to the left end on the stalk. The direc- tion of the excitatory current is thus reversed, being now from left to right (fig. 147 (^)). By algebraical summation, there now occurs a negative variation of the stalk-current, and a positive variation of the leaf-current, in Dioncea and Ficus religiosa^ while the very opposite takes place in Citrus. I shall here draw attention once more to those errors to which an investigator becomes liable when he infers that positive and negative variations must necessarily be the expression of assimilatory and dissimilatory processes. For we have just seen that the same responsive current, by alge- braical summation with two opposite-directioned resting- currents, may appear to be both positive and negative, at one and the same time. Again, with a single resting-current, it is possible to obtain either a positive or a negative varia- tion, according as the same stimulus is applied to the right or the left. It is now abundantly clear that the one uni- versal effect of stimulus is to give rise to a responsive current which flows from the more to the less excited portions of the tissue. If there be already an existing current, the responsive current is added to this algebraically, and induces, according to circumstances, either a positive or a negative variation. Much confusion, and many erroneous inferences would be avoided, if instead of looking at these variable indications attention were centred on the one constant criterion, namely I RESPONSE OF LEAVES 227 that the excitatory current always flows from the more to the less excited portions of the tissue. Another effect observed by Burdon Sanderson was, that on cutting the petiole across, the existing normal leaf-current was increased, the amount of this increase being determined by the length of the petiole cut off, in such a way that the shorter the petiole left, the stronger the leaf-current became. In Natu7'e (vol. x. p. 128), he suggested an explanation of this phenomenon. In the leaf of Dioncea^ as already said, there is a resting-current in the stalk, opposed in direction to that in the leaf Thus ' the electrical conditions on opposite sides of the joint between stalk and leaf are antagonistic to each other ; consequently, so long as the leaf and stalk are united each prevents or diminishes the manifestation of electro- motive force by the other.' He thus inferred that the pro- gressive removal of the antagonistic element, by section of the stalk, would serve to enhance the intensity of the leaf-current. Taking the ordinary leaf of Ficus religiosa^ I have myself been able to obtain results precisely similar to those described in Dioncea^ by making successive sections of the petiole, at shorter and shorter distances from the point of junction. The leaf-current at each section underwent an increment. The parallelism of the two sets of effects will be seen from the following table. It Ficus Leaf. DioN^A Leaf (Burdon Sanderson). Length of stalk Galvanometric deflection 7 cm. 16 divisions 36 50 60 1 Length of stalk | Galvanometric deflection 1 2-5 cm. 1-25 „ 0-6 „ 40 divisions 50 „ 65 » 0-3 „ 90 „ 1.4,, Burdon Sanderson's suggested explanation that the suc- essive augmentations of the leaf-current were due to suc- cessive removals of the antagonistic element, by section, is Q2 II / 228 COMPARATIVE ELECTRO-PHYSIOLOGY quite untenable. He failed to see that the effect was, on the contrary, due to the increasing excitatory action of the sections themselves. Similar results may be obtained, even without the bodily removal of the supposed antagonistic element, if, instead, we apply an increasing intensity of stimulus, as say, by contact of a hot wire at points nearer and nearer to that of junction. In the case of the trans- verse section, the cut acts as a stimulus, and the respon- sive current flows from the left to the right. Algebraical summation of this with the existing leaf-current, which is also from left to right, causes an increase, or positive variation of it, in a manner exactly the converse of the negative varia- tion induced in the leaf, when the stimulus was applied on the lamina. As the section is made nearer and nearer to the point of junction, the degree of stimulation, and the con- sequent positive variation of the resting-current, must become greater and greater. And lastly, in the case of the longitudinal leaf-current, Burdon Sanderson found that if a current from a battery were directed through a leaf-stalk, at the same time that the two ends of the midrib were led off to the galvanometer, the difference previously existing between the ends of the midrib would be increased, if the current led through the leaf-stalk were in the same direction with the leaf- current, and diminished, if it were in the opposite direction. A similar effect, as seen in the conducting tissues of ordinary plants, will be studied in detail, when we take up the question of the extra-polar effects induced by electrotonic currents (Chap. XXXIX.). We have already seen that, by means of induced varia- tion of the longitudinal stalk-current, under the stimulation caused by section of the petiole, it is easy to obtain an un- mistakable indication of the nature of the true excitatory electrical change. Burdon Sanderson, however, laboured under the disadvantage, as already said, of having failed to recognise that a section acts as a stimulus. His investi- gation, therefore, on the character of the excitatory variation, RESPONSE OF LEAVES 229 was chiefly carried out by means of experiments on electrical variations induced in the lamina. These depended (1) on variations in the cross-difference of existing potential between the upper and lower surfaces, according to his ' fundamental experiment/ and (2) on electrical variations in the led-ofifs of symmetrical surfaces of contact on the under-side of opposite lobes. The results which he obtained, however, by these methods, appear to the reader to have been very conflicting, and in fact the experimental methods described by him would seem to have been open to many sources of complica- tion of which he himself was unaware. 'IG. 148. Burdon Sanderson's Funda- mental Experiment on Diomva Leaf Electrical stimulus applied on distal I lobe, ?, induces responsive effect on led-off circuit ftn. Upper or internal surface, /, more excitable than lower, in. Fig. 149. Parallel Experiment in Sheathing Petiole of Musa Thermal stimulus applied on distal side induces responsive effect on led-off circuit. Upper or 'internal' surface more excitable than lower. I^= I shall deal first with Burdon Sanderson's ' fundamental experiment,' of which the excitatory electrodes are seen on the left lobe, and the led-off on the right in fig. 148. In fig. 149 is given a diagram of a parallel experiment carried out by yself on the petiole of Musa. According to Burdon- Sanderson, as the result of excitation, a f current is induced in the right lobe of Dioncea (fig. 150). This means, of course, that the upper or more excitable surface of the right lobe has become positive to the lower. This current, how- ever, he termed * excitatory,' regarding it as the analogue of the 'action-current' known to animal physiology. After I" ■' *■ ~ ■ " " " ~ " 230 COMPARATIVE ELECTRO-PHYSIOLOGY observed a second phase to set in, in which the upper surface became relatively negative to the lower. This negative change, which he called the * after-effect,' he described as taking place at that moment at which the mechanical effect of excitation also made itself evident. This negative phase — called by him the ' after-effect ' — Burdon Sanderson regarded as connected with those electrical changes which had been observed by Kunkel to be induced by movement of water in the tissues. The first effect on the Figs. 150, 151, 152. Recordslof Electrical Responses of Different Leaves of Diomea according to Fundamental Experiment of Burdon Sanderson Fig. 150. Positive response of certain leaves of Diomea. Time-marks 20 per second (Burdon Sanderson). Fig. 151. Diphasic response of leaf of Diomea ' in its prime.' Positive followed by negative. Time-marks 10 per second (Burdon Sanderson). Fig. 152. Positive response of same leaf when 'modified' by previous stimulation. Time-marks 10 per second (Burdon Sanderson). The above records were obtained with capillary electrometer. contrary, which immediately preceded this, and was charac- terised by relative positivity of the upper surface, he regarded, as already mentioned, as the true excitatory or action-effect. The following is from his summary : * The first phase of the variation --the effect which immediately follows excitation, and has an opposite sign to the after-effect, and a much higher electro-motive force — does not admit of a similar explanation : for it cannot be imagined that a change which spreads over the whole lamina in less than one-twentieth of a second can be dependent on migration of water. The excita- tory disturbance which immediately follows excitation RESPONSE OF LEAVES 23 1 is an explosive molecular change, which by the mode of its origin, the suddenness of its incidence, and the rapidity of its propagation, is distinguished from every other phenomenon except the one with which I have identified it — namely, the corresponding process in the excitable tissues of animals. Of the nature of this preliminary disturbance (to which alone the term ex- citatory variation ought to be applied, it alone being the analogue of the 'action-current' of animal physio- logy) we know nothing. . . . The direction of the ex- citatory effect in the fundamental experiment is such as to indicate that in excitation, excited cells become (positive to unexcited, whereas in animal tissues excited parts always become negative to unexcited. The ap- parent discrepancy will probably find its explanation in the difference of the structural relations of the electro- motive surfaces.' ^ From this quotation it will be seen that Burdon Sanderson had fallen into the basic error of mistaking what I have demonstrated to be the hydro-positive, for the true I excitatory effect, and vice versa. In a subsequent Paper again {Phil. Trans, vol. 179, 1889) Burdon Sanderson published certain results, which differed from those referred to above. He had previously found that usually speaking the upper surface of each lobe was negative to the lower. Later, however, he came to the conclusion that in the leaf of DioncEa in its * prime,' the upper surface 1^^ was positive to the under. On repeating his * fundamental ( experiment' moreover, with these vigorous leaves, he found that instead of the pronounced positive response which he had ^^fcpreviously observed, he now obtained a short-lived positive effect succeeded by a strong negative (fig. 151). He was unable to offer any definite explanation of this difference between the two sets of results, but suggested that it might I arise, in some way, from changes of the resting-current. i, ' Phil. Trans. 1882, vol. 173, p. 55. 232 COMPARATIVE ELECTRO-PHYSIOLOGY ' In the leaf, observed facts show most conclusively that the two sets of phenomena — those of the excited and those of the unexcited state — are linked together by indissoluble bands : that every change in the state of the leaf when at rest conditionates a corresponding change in the way in which it responds to stimulation, the correspondence consisting in this, that the sign, that is the direction, of the response is opposed to that of the previous state, so that, as the latter changes sign in the direction from t to I , the former changes from \ to t •' ^ In making this statement, Burdon Sanderson was prob- ably guided by the prevalent opinion that response takes place by a negative variation of the existing current of rest. We have seen, however, that this supposition is in fact highly misleading. For, owing to such fluctuating factors as age, season, previous history, or excitation due to prepara- tion, the so-called current of rest may and frequently does undergo reversal. Thus a single excitatory effect might, as we have seen (pp. 175-177) under different circumstances, appear either as a positive or a negative variation of the existing current. The assumption of the universality of response by negative variation is thus seen to be unjustifiable. Indeed, it would appear from the description of some of the experiments actually related by Burdon Sanderson him- self, that response did not, even in these cases, always take place by negative variation of the existing current. For instance, while in the leaf of Dioncea in its * prime ' (upper surface positive) the response is negative, and while this latter becomes reversed to positive, as he tells us, in conse- quence of* modification' due to previous excitation (fig. 152), yet he admits that even in these circumstances the upper surface had first returned to positivity {ibid. p. 447). Thus, though the resportses of the leaf in its * prime,' and of the ' modified ' leaf are opposed, yet the antecedent electrical condition of the modified leaf has not in this case undergone reversal. Phil. Trans. 1889, vol. 179, p. 446. RESPONSE OF LEAVES 233 The suggestion, therefore, that the reversal of response is due. in some way unexplained, to a reversal of the electrical condition of the leaf, cannot hold good. Nor does the use of the term ' modification ' in any way assist in the elucidation of the phenomenon. A satisfactory explanation of this reversal of response, then, still remains to be found. So much for the * fundamental experiment' The next experi- mental arrangement employed by Burdon Sanderson consists of a leaf l^irhich is led off by symmetrical 11 Fig. 153. Experimental Con- nections with DiofKBa ac- cording to the Second Experimental Method of Burdon Sanderson IS "contacts on the under surfaces of its two lobes (fig. 153). If now the ght lobe was excited, by touching one of the sensitive filaments (on the upper surface) with a camel's-hair pencil, in the neighbourhood of the leading-oft contact, it was found that the under-surface of the right lobe became first positive, and subsequently negative (fig. 154), :elatively to the left {ibid. p. 440). Fig. 154. Response of Under-surface of Leaf of Z)/tf«^?a, with Electrical Connections as in Fig. 153 Mechanical excitation of upper surface of right lobe shows relative positivity of under surface of same right lobe (up curve), followed by its relative negativity (down curve). Time-marks 20 per second (Burdon Sanderson). Summarising these various observations, then, we find jsults which are very much at variance. First, according to the ' fundamental experiment,' certain leaves are seen to give rise to the positive response ; other leaves, in their 234 COMPARATIVE ELECTRO-PHYSIOLOGY first positive and then negative. These latter again, after previous excitation, become so modified as to show only- positive changes. And lastly, using the experimental arrangement of symmetrical contacts, a diphasic variation is obtained — positive followed by negative — on the under- surface, instead of the upper, of the lobe excited. No theory is advanced, however, by which a comprehensive explanation might be afforded of these apparently anomalous results. But from the generalisations which I have already esta- blished, regarding the electrical signs of the hydro-positive and true excitatory effects respectively, and from the results of certain experiments on ordinary leaves which I shall presently describe, it will be found easy to arrive at a true explanation of the various observations related by Burdon Sanderson, which would otherwise have appeared inexplic- able. The fact that hydrostatic disturbance induces galvano- metric positivity, and that true excitation induces negativity, has already been clearly demonstrated under conditions from which all possible sources of complication had been elimi- nated (p. 6i). The experimental arrangement adopted by Burdon Sanderson, however, laboured under the double dis- advantage, not only of a liability to confuse the hydro- positive and true excitatory effects, but also of the com- plexity arising from the differential excitability of the responding organ. It is only indeed by the closest analysis that it is possible to discriminate, in his results, between such as are due to true excitation and those arising from the hydro- positive effect. The various electrical phenomena which are possible in an anisotropic organ in consequence of the hydro-positive and excitatory effects respectively, may be clearly exhibited, as I have already shown, by means of the mechanical response of the leaf of Mimosa. With regard to this, we have seen (pp. 59, 60) that direct stimulation of the pulvinus induces a negative mechanical response, or fall of the leaf, by the greater contraction of the more excitable lower half of the organ. The corresponding electrical variation would RESPONSE OF LEAVES 235 thus consist in the greater galvanometric negativity of this more excitable lower, in relation to the less excitable upper half. If the stimulus, however, be applied at some considerable distance, so that true excitation cannot reach the responding point, then we have an erectile or positive mechanical response of the leaf. This is brought about by the relatively greater expansion of the more excitable. The corresponding electrical response will be the galvanometric positivity of this more excitable, in relation to the less excitable half of the organ. Between these two extremes lies that experi- ment in which stimulus is applied at some intermediate point, the consequence of which is that the hydro-positive Ifcwave, with its greater velocity, reaches the responding organ earlier than true excitation, thus bringing about a pre- liminary erectile or positive response, followed by the ex- ■■fcitatory negative or fall of the leaf The corresponding electrical response would therefore be diphasic, positive (followed by negative. But the occurrence of this second or negative phase is bnly possible when the conductivity 4s so great as to allow the wave of true excitation to reach the organ. We may imagine that in a very vigorous plant, with its great con- IBtiuctivity, we have found a point, at the maximum distance from which the true excitatory effect of a given stimulus is capable of transmission to the organ. With such a speci- men, in its ' prime,' we shall observe a diphasic effect — pre- liminary positive followed by negative. But if we took a less vigorous specimen, and applied the stimulus at the same distance from the responding point, the true excitatory wave ould fail to reach the responding organ, and we should see here, only the positive effect due to hydro-positive action, ence, two different specimens, treated in exactly the same ay, may exhibit two different effects, one diphasic, and the ther positive alone ; this difference being due to their nequal vigour, and concomitant inequality of excitability nd conductivity. This will account for the diphasic and ositive responses which were exhibited by the more and 236 COMPARATIVE ELECTRO-PHYSIOLOGY less vigorous leaves respectively of Dioncsa^ when stimulation was applied on the distal lobe, according to the fundamental experiment of Burdon Sanderson. We must next refer to the reason why a leaf that origin- ally gives diphasic response — positive followed by negative — undergoes such ' modification,' in consequence of previous excitation, as thereafter to give only positive response. We have seen that the negative element of the diphasic response is due to the arrival at the responding point of the true excitatory w^ave originated at the distant point of stimula- tion. Now it has been shown (p. 65), that if by any means the conductivity of an intervening region should become diminished, we may expect that the hydro-positive effect will continue to be transmitted, although the passage of true excitation is partly or wholly blocked. By means of this selective block, I was able to unmask the hydro-positive component present in resultant response (cf fig. 49). I have shown elsewhere ' that the conducting power of a tissue will be impaired by the fatigue consequent on previous stimulation. Thus, in the petiole of Biophytum^ I found that while the plant, when fresh, had a conductivity measured by the velocity of transmission of excitation, at a rate of r88 mm. per second, the same plant, when partially fatigued by four successive stimulations, had its conductivity dimi- nished, the velocity of transmisson being now only 1-54 mm. per second. The diminution in this case, then, was about 18 per cent. I shall moreover . show in a later chapter that in consequence of growing fatigue the passage of true excitation may at a certain stage be arrested, the hydro- positive effect alone being then transmitted. It is thus easy to explain how it was that in Burdon Sanderson's experi- ment, of stimulus applied on the distal lobe, the wave of true excitation became blocked, and the 'modified' leaf gave positive response alone. These considerations will be found as I think, to offer a satisfactory explanation of the conflicting results arrived at by Burdon Sanderson. ' Plant Rest)onse, p. 244. RESPONSE OF LEAVES 237 I shall now, however, proceed to describe a series of ex- periments exactly parallel to the ' fundamental experiment ' on Dioncea, carried out on ordinary plants. We have seen that the inner or concave surface of the sheathing petiole of Mzisa is relatively more excitable than the outer or convex. Thus it corresponds with the ' internal ' or upper surface of the leaf of Dioncea. The more excitable internal surfaces of both these, again, correspond with the more excitable lower half of the pulvinus of Mimosa. In fig. 149 is shown an experi- mental arrangement with a specimen of Musa which will be seen to be parallel to that ot Burdon Sanderson's funda- mental experiment on Dioncea. In order to avoid any such disturbance as might conceivably arise from current-escape, if the electrical form of stimulus were used, I employed the thermal mode of stimulation. A momentary heating-current passed through a thin platinum wire gave the thermal varia- jtion required, and was found to furnish a very satisfactory brm of stimulus. The led-off circuit was at first placed at a istance of 16 mm. from the point of stimulation. As the timulation was moderate, and as the conductivity of the issue was not great, the effect induced at the respond- ng circuit was hydro-positive, the more excitable concave urface becoming positive (fig. 155 W)* This response is the same as the positive responses given by the 'unmodifiable leaf of Dioncea (fig. 150), as well as that of a vigorous leaf hich had been 'modified' by fatigue (fig. 152). On next taking a second pair of led-off points, at the shorter distance of 8 mm., the hydro-positive effect reached the led-off points earlier, and was followed by the true excitatory wave. This is seen as a preliminary positive response, followed by the excitatory negative (fig. 155 {b) ). This again is the same as the di-phasic response of a Dioncea leaf in its 'prime' I (fig. 151). In the experimental arrangement with J///j-^ the led-off circuit was now brought still nearer to a distance of r4 mm. There was now little interval between the arrival at the led-off points of the hydro-positive and true excitatory lei 238 COMPARATIVE ELECTRO-PHYSIOLOGY expression, the former is masked by it, and we obtain here only the excitatory negative variation (fig. 155 (c)). It only remains to consider the responses which Burdon Sanderson obtained with symmetrical contacts (fig. 1 52) on the under-surfaces of the two lobes. In the next figure (fig. 153) is reproduced his record of electrical response, obtained on mechanical stimulation of a sensitive filament situated on the upper surface of the right lobe, vertically above the right-hand led-ofif. This response is, as will be seen, di- FiG. 155. Photographic Records of Positive, Diphasic, and Negative Responses ot Petiole of Musa depending on the Effective Intensity of Transmitted Stimulus (a) Here stimulus was applied at a distance and hydro-positive effect alone transmitted ; {d) Stimulus was applied nearer, and the positive effect was succeeded by the true excitatory negative ; {c) Stimulus was applied very near, with the result of true excitatory negative response. phasic, its first phase being one of relative positivity of the under-surface of the excited lobe, and the second representing its subsequent relative negativity. This first phase is clearly due to the earlier transmission of the hydro-positive or indirect effect of excitation, from the stimulated point on the upper surface. It was supposed by Burdon Sanderson that the second phase of this re- sponse represented the later arrival of the same positive effect at the distal second contact, which would thus induce reversal. But it appears much more probable that this second ph&se of negativity is due to the arrival at the RESPONSE OF LEAVES 239 under-surface of the wave of true excitation, initiated vertically above. This relative negativity of the under- surface may or may not be helped by the induction of positivity at the distal, due to the transmission of the hydro- positive effect. This view is supported by the fact that in a corresponding experiment on an ordinary leaf, in which the second contact was at a distance too great to allow of the effective transmission of any hydro-positive wave, the stimulation of the upper surface induced a similar diphasic response at a point diametrically opposite, on the under side. In this case the second or negative component of the response could not be due to anything but the subsequent arrival of the true excitatory wave with its concomitant negativity. ! It is now clear that among the various results ■■tbtained from the study of the electrical responses of the leaf of Dioncsa, there are some which do not represent I true excitation at all, while in others it is only one of the two phases which is significant of this, the other being due to the hydro-positive effect. We have also seen that Burdon Banderson at starting fell into the error of wrongly identify- ing the true excitatory electrical effect with that which was Bue to the hydro-positive effect, and vice versa. We have seen that there is not a single response given by the so-called excitable leaf of Dioncea^ which cannot be obtained under similar conditions from the leaves of ordinary plants also. |^k[n fact it has been by means of experiments carried out on the latter that we have been enabled to unravel all the intricacies which were offered by the recorded responses of e lamina of Dioncea. It has further been shown in the course of the present hapter that the leaf and stalk currents observed in Dioncea re also found in, for instance, the leaf of Ficus religiosa. hese have been shown, moreover, to be due to physiological ifferences between an intermediate and the terminal points, he negative variation of the leaf-current, and the positive ariation of the stalk -current, on the stimulation of the 240 COMPARATIVE ELECTRO-PHYSIOLOGY lamina, were both alike shown to be the result of the alge- braical summation of a definite excitatory current with the two opposite-directioned resting-currents. The positive variation of the leaf-current, again, on section of the petiole, has been traced to the same cause, namely the stimulatory action of mechanical section, giving rise to an excitatory current which was summated with the existing leaf-current. Finally the positive response of the concave surface ot Dioncea has been shown to arise, not from any specific difference between plant and animal response, but from the fact that in this particular case it was the indirect hydro- positive effect of stimulus that was transmitted, inducing an action opposite to that of true excitation. CHAPTER XIX THE LEAF CONSIDERED AS AN ELECTRIC ORGAN Electrical organs in fishes— Typical instances, Torpedo and Malepterurus — I Vegetal analogues, leaf of Pterospertmim and carpel of Dillenia indica or , pitcher of iW/^«//z^— Electrical response to transmitted excitation —Response . to direct excitation — Uni-directioned response to homodromous and hetero- \ dromous shocks — Definite-directioned response shown to be due to differential \ excitability — Response to equi-alternating electrical shocks — Rheotomic ob- f servations — Multiple excitations — Multiplication of terminal electromotive effect, by pile-like arrangement, in bulb of UricHs lily. T has been shown that by a study of the peculiarities of electrical response in plants, it is possible to obtain an insight into the obscurities of similar responses in the animal tissue. Among animal structures, there is one — the elec- trical organ of certain fishes — the explanation of whose action offers unusual difficulties to the investigator. But I shall attempt to show in the course of the present chapter, that there are also, on the other hand, vegetable structures, the study of which will be found to elucidate the electro-motive action here involved. Taking that of the Torpedo as type, we find that the electrical organ is disposed in the form of columns, each column consisting of numerous electrical plates, arranged in series, one over the other, like the plates in a voltaic pile. Each electrical plate consists of a rich plexus of nerve-fibres imbedded in a gelatinous mass. There are thus two surfaces, one nervous and the other non- nervous. Each disc then becomes electro- motive under the impulse from the nerve. Though the induced electro-motive force in each plate is small, yet in consequence of their serial arrangement in columns, the elements are coupled for inten- sity, and the resulting E.M.F. of discharge becomes high. I 242 COMPARATIVE ELECTRO-PHYSIOLOGY Fritsch estimates the total number of these plates in some of the Torpedos to be over 1 50,000. From the point of view of their development, these electrical organs in general constitute modified muscles, containing nerve-endings. The electrical fish known as Malepterurus of the Nile is an exception to this rule, inas- much as morphological evidence goes to prove that in its case it is glandular, rather than muscular, elements which have been so modified. The peculiar characteristic of the discharge of electrical organs in general, is that it takes place in a definite direction at right angles to the plates. It was Pacini who tried to establish the generalisation that the direction of the dis- charge would be found to be dependent on the morphological character of the organ. He found that as a general rule the discharge takes place in a direction from that surface of the disc which receives the nerve (henceforth to be referred to as the anterior surface) to the opposite non-nervous, or posterior^ surface. Thus in the Torpedo^ where the plates are horizontal, and the anterior or nervous surface constitutes the verttral aspect of the disc, the discharge is from the ventral or anterior, to the dorsal or posterior surface. In Gymnotus again, the plates or discs are vertical to the long axis. The anterior or nervous surface is here towards the tail-aspect, and the discharge is from tail to head. If these cases had been all, Pacini's generalisation, as regards the direction of discharge — from the anterior nervous to the posterior non -nervous — would have been complete, and from it some attempt might have been made to offer an explana- tion of the phenomena. Unfortunately, however, this is not so, since Malepterurus presents a hitherto inexplicable ex- ception to the rule. In this fish, though the anterior or ' nervous surface is towards the tail-aspect as in Gymnotus^ yet the discharge is in the opposite direction towards the head : that is to say, from the posterior surface to the anterior. The difficulties in the way of an explanation of the activity of these electrical organs of certain fishes are thus seen to be THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 243 very great. Is the activity something specific occurring in these fishes alone, and unrelated to other electro-motive phenomena in the animal tissues? Or is it related to the electromotive action already observed in excited muscles ? In support of the latter view, it is urged that most of the electrical organs consist of modified neuro-muscular elements. Against this argument, however, as we have just seen, is the instance of Malepterurus, in which, from a morphological standpoint, the organ is to be regarded as a modified gland, and therefore not muscular in character. I^K There are certain peculiarities, further, about the action l^pf these organs which call for elucidation. Among these is the question of the character of the natural current of rest, about the significance of which there have been differences of opinion. There is also the fact that the organ, under a single strong excitation, gives rise not to one, but to a series of electrical responses. -^ We have seen that the apparently unique character of I^Piis group of organs constitutes an added difficulty in arriving at a correct theory on the subject. But it is clear that if we could succeed in discovering among vegetable organs any cases which showed similar characteristics, IHire should then be so much the nearer to the determination of that fundamental reaction on which the phenomenon in animal and vegetable alike depends. In the typical case of Torpedo^ it has been seen that the conducting nerve, when entering into an electrical plate, breaks into an extensive ramification, and thus forms the nervous surface, in contradistinction to the jelly-like sub- stance in which it is imbedded, forming the opposite, and here indifferent surface of the plate. Now this arrangement js closely imitated by many ordinary leaves, in which the l^ascular elements break, on reaching the lamina, into a pro- fuse arborisation. I must here anticipate matters to say that I have discovered in the fibro-vascular bundles of plants (see Chap. XXXII.) Hements which are in every way analogous to the nerves of 244 COMPARATIVE ELECTRO-PHYSIOLOGY animals. For an exact vegetal analogue to the electrical plate of Torpedo, we may take certain leaves in which the ventral, or anterior, surface is formed of a prominent network of highly excitable nervous elements, while the upper consists of an indifferent and relatively inexcitable mass of tissue. An example of this may be found in the leaf of Pterospermum suberifolium (Rox.) whose lower surface is characterised by a remarkably perfect venation, while the upper or posterior is dry and leathery. Thus the nerve passing into an elec- trical plate of Torpedo corresponds with the petiole attached to the leaf just described, since in the two cases alike, it is the ventral surface which contains the highly excitable nervous elements. In the exceptional Malepterurus, on the other hand, it is, as we have seen, a modified gland, and not a modified muscle, which forms the posterior surface of an individual electrical element. Morphologically speaking, the vegetal analogue is found in such organs as the carpellary leaf of Dillenia indica, or the pitcher of Nepenthe , both of which are glandular on their upper or inner surfaces. In point of structure, then, these leaf-organs are analogous to single discs or elements of the electrical organs of Torpedo and Malepterurus respectively. But we have still, in the course of the present chapter, to inquire whether the electrical reactions are equally correspondent — that is to say, whether, on stimulation, the excitatory current in the type of vege- table organ represented by the leaf of Pterospermum is or is not, from the lower or anterior surface to the upper posterior, as in the electrical plate of Torpedo ; and, con- versely, whether in the type represented by the carpel of Dillenia or the pitcher of Nepenthe the excitatory current is from the posterior to the anterior surfaces, corresponding with the discharge in the electrical element of Malepterunis, from the posterior glandular to the anterior non-glandular surface. While dealing with the theory of the action of electrical organs, I shall be in a position to show that the characteristic THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 245 reaction of each of these two types is governed entirely by the question of the relative excitabilities of the two surfaces. The physiological anisotropy on which the distinctive effect of the type depends is very pronounced in the representa- tive cases of the vegetal analogues which have been named. In many other cases, however, though the results under normal conditions are fairly definite, and approach one or other of the two types, yet the characteristic responses are liable to be reversed under the physiological modifications induced by age and surrounding conditions. In this way it may be said of the leaf of water-lily {Nymphcea albd)^ of Bryophyllum calcineum^ and of Coleus aromaticus that when vigorous, and in their proper season, their responses are of the first of these two types, while those of the bulb-scale of Uriclis lily, with its glandular inner surface, are of the second type. The electrical organ of the fish may be excited indirectly by means of stimulus transmitted through the nerve ; or direct stimulation may be applied, as by means of induction- shocks. Under either of these conditions the excitatory discharge is definite in its direction. In the case of Torpedo^ as already mentioned, this is always from the ventral and (anterior to the dorsal or posterior surface. Turning then to ^■the corresponding vegetable organ of the first type, I shall show that transmitted stimulus induces an effect exactly similar ; and I shall demonstrate this experimentally by means of the leaf of Nymphcea alba. Suitable galvanometric connec- tions were made with the ventral anterior and with the dorsal posterior surfaces of the lamina. Thermal .shocks, by means of the electro-thermic stimulator, were applied on the petiole, IBdose to the lamina, at intervals of one minute, records being taken photographically of the resulting responses. It should be remembered here that excitation is transmitted to the lamina by the conducting nerve-like elements present in the petiole. The records (fig. 156) show that the effect of this periodically transmitted stimulation was a series of respon- sive currents, whose direction was like that of the discharge 246 comparative: ELECTRO-PHYSIOLOGV Of great importance was the investigation carried out by Du Bois-Reymond on the effects induced in the electrical organ by the passage of currents in different directions. Polarising-currents in the direction of the natural discharge of the organ are distinguished, in the terminology introduced by Du Bois-Reymond, as homodromous, and those in the opposite as heterodromous. Polarisation -effects in the direction of the natural discharge he distinguishes as ' absolutely positive polarisation,' and against that direction * as absolutely negative.' A polarisation-current in the same direction as the polarising- current he calls ' relatively positive/ and in the opposite direction ' rela- tively negative ' polarisation. It was found by him that polarising- currents of fair intensity and short duration, whether homodromous or heterodromous, would always give rise to polarisation-currents in the same direction as the natural dis- charge. He believed this to be due to the occurrence in the electrical organ of two different polarisation- effects, positive and negative. This will be understood from his own dia- grammatic representation (fig. 157) of the effect which he supposed to take place immediately on the passage of the polarising-current. In the upper figure the ascending arrow represents the homodromous polarising-current. This gives rise, according to Du Bois- Reymond, to two opposite polarisation-effects. The de- flection seen in the galvanometer is the resultant of these, represented by the shaded part of the figure. The resultant of a homodromous current, then, is positive polarisation, both absolute and relative. The heterodromous current, on the other hand, induces absolutely positive and relatively Fig. 156. Electrical Response of Lamina oi Nymphiea alba due to Transmitted Excita- tion from Petiole Direction of responsive current from anterior or lower to posterior and upper surface. THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 247 3L E negative polarisation. According to Du Bois-Reymond, further, heterodromous shocks induce no relatively positive polarisation, or only infinitely little (see down curve in lower part of figure). On sending congruent alternating currents from Saxton's machine, he obtained only the absolutely positive polarisation - effect. This he accounted for by supposing the relatively nega- tive polarisations in both irections to cancel each Dther ; the heterodromous positive to be so small as to be practically negligible ; and the homodromous posi- Iive therefore to be alone iffective. Du Bois-Reymond failed o recognise the element of ixcitation in these phe- iiomena. What he calls positive polarisation has been shown by subsequent workers to be due to local polar ex- ( citation. But the question Ls to how polarising-currents in both directions could give rise to a single-directioned responsive effect has not up llktp the present, so far as I am aware, been explained fully and satisfactorily. The ex- periments carried out on leaves, Fig. 157. Diagrammatic Representa- tion by Du Bois-Reymond for Ex- planation of Electrical Response in Organ of Torpedo The natural discharge is here supposed to be from below to above, h homodromous current | (upper half of figure) is supposed to induce two opposite polarisations, positive and negative. The resultant, repre- sented by shading in figure, is absolutely and relatively positive. A heterodromous current \ , on the other hand, is regarded as inducing a resultant absolutely positive and relatively negative polarisation (lower part of figure). to which I am about I [escribe, will, however, throw much light on this subject. It has already been shown, from anatomico-physiological onsiderations, that there are certain leaves which approxi- mate to the character of single plates of such electrical organs i 248 COMPARATIVE ELECTRO-PHYSIOLOGY as that of Torpedo. One such leaf, already mentioned, was that of Pterospermum. When induction- shocks are sent in both homodromous and heterodromous directions through such a leaf, between upper and lower surfaces, the leaf being, it is understood, in a normal condition, a responsive current is found to be evoked, always in one direction — that is to say, from the lower or anterior to the upper or posterior surface. This is strictly parallel to the electrical reaction observed by Du Bois-Reymond in Torpedo. That this result is really due to the excitatory effect is proved by the fact that the same is found to occur when other forms of stimulation are used. Thus, if we place the leaf of Coleus aromaticus within a surrounding thermal helix, suc- cessive thermal shocks, acting simultaneously on both surfaces, give rise to responsive currents which are, as in the last case. Fig. 158. Photographic Records from the lower anterior to the 1:^Sr^^Ji::X^tl upper posterior surface. Fig. 158 Surfaces are Excited Simul- gives a series of such responses. taneously by Thermal Shock ,^ .^ r . i • i i , ^ . , . ^ , r rom the fact which has already Resultant responsive current from _ ^ more excitable anterior to less been fully established, that on excitable posterior surface. ^^^^ simultaneous excitation of two points the responsive current is always from the more to the less excitable, it is quite clear that in the present case it is the lower or anterior surface of the leaf which is the more excitable. These responsive currents, obtained under a non- electrical form of stimulus, and similar to those evoked by electrical shocks, completely demonstrate the fact that the result is brought about, not by polarisation, either positive or negative, but by the differe7iti(il exxitability of the tissue itself. The response of electrical organs in general, then may be summarised in the following law : The excitatory discharge is determined by the physiological anisotropy of the organ, its definiteness of direction being deter- I I THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 249 mined by the fact that the responsive current is always from the more to the less excitable of the two surfaces. Referring once more to the definite-directioned after- current which we have seen to be induced as the result of polarising-currents, whether homodromous or heterodromous, it is now clear that these currents act as an electrical form of stimulus. The intensity of the after-current here seen in the galvanometer, however, is not wholly due to the excitatory electro-motive change, but in part also to physical polarisation, which is added to it algebraically. Thus, an exciting homodromous shock gives rise to an electrical after-effect, in which the excitatory current is opposed by l^p counter-current of negative polarisation. Under a hetero- dromous shock, on the other hand, the excitatory electrical change becomes summated with the negative polarisation, which is now in the same direction as itself. In these cases, though the preponderating nature of the excitatory effect determines the definite direction of the after-effect, yet it is difficult to know how much of the latter is actually due to excitatory action as such, and how much to ordinary Klarisation helping or opposing this. Very much greater complexities ensue again in practice m the difference between anodic and kathodic actions on the two unequally excitable surfaces. In Torpedo, for instance, according to Du Bois-Reymond, the electrical organ responds better to a homodromous than to a heterodromous l^fcciting current, while in Malepterurus, according to Gotch, the reverse is the case, the heterodromous being more efficient than the homodromous. Such diversity of results is prob- ably to be accounted for by the considerations to which I have referred. If we take, tor example, the simplest case, that in which the anterior surface is more excitable than the posterior, and if we suppose an induction-current of moderate intensity to be sent in a homodromous direction, we may assume that Pfliiger's Law — the kathode excites at make, and the anode at break — will hold good. We shall here, for the sake of I 2 so COMPARATIVE ELECTRO-PHYSIOLOGY simplicity, neglect any effects that may accrue from anode- make and kathode- break. Under a homodromous induction- shock, then, two different excitatory electrical changes will be induced, on the lower and upper surfaces respectively, the consequent currents through the tissue being in opposite directions. On these, moreover, will be superposed again the polarisation-current. Calling the effect induced by anode- break as A,, and that of kathode-make as K,,^, we shall obtain a resultant consisting of A^ on the more excitable anterior surface, minus K^ on the less excitable posterior, minus the negative polarisation-effect. Under a heterodromous shock^ on the other hand, we shall have K,„ on the more excitable anterior surface, minus A^ on the less excitable posterior, ///^j the negative polarisation-effect. Even this, however, does not exhaust the possibilities of complication. For I shall show in a subsequent Chapter, and have already shown elsewhere, that under a high E.M.F. Pfliiger's Law does not apply. The relative excitatory values of anode and kathode may indeed undergo one or more reversals, according to the intensity of the acting electro-motive force. Thus, under a moderately high E.M.F. in what I have designated the A stage, both the anode and kathode are found to excite at make, and either kathode or anode at break. In the B stage, under a still higher E.M.F., it is the anode which excites at make, and the kathode at break. It will thus be seen what a number of complicating factors may be present when an organ is excited by currents of varying direction and intensity. If, then, we wish to study the purely excitatory reaction of an organ, as dependent solely upon its individual characteristics, uncomplicated by defects inherent in the method of excitation, we must see first that the applied stimulus is equal on both surfaces, and, secondly, that such factors as are not excitatory — that is to say, negative or counter-polarisation —are eliminated. These ends may be accomplished by subjecting the responding organ to symmetrical and alternating equal and opposite shocks, THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 2$ I following each other in rapid succession. For the resultant negative polarisation will in practice be neutralised, if the primary polarising currents are similar, equal, and opposite. The stimulus applied on the two surfaces, moreover, will be equal, if the two rapidly succeeding and opposite-directioned shocks be so symmetrical as to be interchangeable. Which- ever may be the factor of excitation will then act equally on both surfaces. Tlie response, therefore, will now be determined solely by the natural difference of excitability as between the two surfaces. I^L It has been said that in order to accomplish these experi- '^Riental conditions, the two opposite shocks should be equal in intensity and in point of time-relations. An ordinary make- and-break Ruhmkorff's shock does not fulfil this condition, since the break-shock is there the quicker and more intense of the two. Moreover, owing to the varying residual magnetisation in the iron core, successive shocks may not be equal. These defects are overcome by sending round the primary, with a constant rapidity, two equal and opposite currents in alternation. During one semi-cycle, then, the primary current l^kries from + C to — C, and during the next from — C to ^^% c, and since these two changes are effected with the same rapidity, the induced currents are symmetrical, equal, and opposite. Such reversal of current is accomplished by means of a tating reversing-key. The key R is wound up against the tension of a spring S, being maintained in this set position by the electro-magnet E, acting on the armature. When the current in the electro-magnet is broken, the alternating double shock from the induction coil I is passed through the experimental leaf L, by means of non-polarisable electrodes Nj Ng. In the case just described the sequence of the current through the primary coil was, say, right-left-right. In the next experiment, by means of the Pohl's commutator, K4, this sequence may be made left-right- left (fig. 159). Employing this method, I have carried out rheotomic observations for determining the time-interval after the shock It I 252 COMPARATIVE ELECTRO-PHYSIOLOGY at which the E.M.F. attained its maximum. The general arrangement here is similar to that described in Chapter IV. (cf. fig. 37). C is the compensator by which any existing electro-motive difference is compensated at the beginning of each experiment. The striking- rod A breaks the current in the electro-magnet E, by which the rotating reverser R is actuated, which brings about equal and opposite shocks to the leaf. The galvanometric after-effect, at any short Fig. 159. Experimental Arrangement for Rheotomic Observations A, B, striking-rods attached to revolving rheotomic disc; K,, key for electro-magnetic release of rotating reverser R ; K^, key for unshunting the galvanometer when pressed by rod, B, for a definite period ; K3, key for preliminary adjustment ; E, electro-magnet with its armature by which rotating reverser, r, is set against antagonistic spring, s ; K^, Pohl's commutator; c, compensator; P, primary, and i, the secondary, of the exciting induction-coil ; N,, N.^, non-polarisable elec- trodes, making electrical contacts with posterior and anterior surfaces of leaf. interval after excitation, is obtained by the un-shunting of the galvanometer, caused by the striker B impinging against the key Kg (fig. 159). We have seen that, owing to the presence of various complicating factors, as well as to the occurrence of negative polarisation, successive responses to homodromous and heterodromous shocks are unequal. By the employment of equi- alternating induction-currents, how- THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 253 ever, we obtain true excitatory effects, unmodified by any such elements of uncertainty. In order to show how perfect the results obtained by this method become, I give here (fig. 160) the records of two successive excitatory responses obtained from a leaf of Bryophylluni cafycinum^ the responsive current being from the lower or anterior surface to the upper or posterior. In this mode of stimulation, by equal and opposite shocks, as already said, no advantage is given to either surface l^ftver the other. Neverthe- less, I thought it well to take two successive records under shocks, in which the l^hternating currents in the ^>rimary circuit were first right-left-right, and then left- t[ght-left. In the electrical organ f Torpedo Gotch found the maximum electromotive change to be attained in about •01 second after the application of the excitatory shock. In leaves, again, I find the rapidity with which the maximum effect is at- tained to depend on the nature of the tissue, and also on the intensity of the exciting shock. In sluggish specimens this may be as long as '2 second. It should be remembered that in the case of mechanical stimulation of moderate intensity also, this period was, similarly, about '2 second (p. 51). With very vigorous leaves of Nymphaa alba^ however, and employing a stronger electrical stimulus, the maximum effect was Fig. 160. Records of Two Successive Responses in Leaf of Bryophyllujn calycinuju under Equi-alternating Electrical Shocks I 254 COMPARATIVE ELECTRO-PHYSIOLOGY attained in a much shorter time —that is to say, in about •03 second. I give below a table showing the rheotomic observations made on such a leaf at gradually increasing intervals after the exciting shock. It should be remembered Table of Rheotomic Observations. Mean interval after the shock Galvanometric deflection •01 of a second •03 „ „ •05 „ „ •07 „ „ •I „ » 20 divisions 63 „ 17 ,, 15 M 20 ,, •2 ,, ,, •3 » ., •5 » „ 9 » 8 , 5 ' that the recording galvanometer was un-shunted for 01 second. The curve given in fig. 161 has been plotted from these results. The maximum electro-motive change took place, as already pointed out, in *03 second after the application of stimulus. This curve shows multiple apices, as was also the case, it will be remembered, after a strong mechanical stimulation (cf fig. 40). This point will be referred to in greater detail in the next chapter. In the course of half a second after the shock, the excitatory electro-motive change had subsided to about one-twelfth of the maximum. It has been said that the excitatory current depends for its definiteness of direction on the physiological anisotropy of the organ. In those leaves in which the physiological differentiation of the upper and lower surfaces is not strongly marked, the differential Fig. 161. Response-curve from Rheotomic Observation on Leaf of Nymphcea alba Ordinate represents galvanometer deflection ; abscissa, time in hundredths of a second. r THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 255 excitability of the two is liable to undergo reversal, under the changing conditions of age, season, and fatigue induced by previous stimulation. In the leaf of Pterospernmm, however, which I have here taken as the type corresponding with Torpedo, the normal differential excitability is generally very persistent Here the excitability of the posterior surface, which is leathery, is slight, and practically negligible. But the anterior surface, with its rich and prominent venation, is highly excitable. The excitatory discharge of such a leaf is thus from the anterior to the posterior. I give in fig. 162 a series of its responses to equi-alternating electric shocks. It will be seen that these are very uniform, and exhibit ractically no signs of fatigue. We have thus found a vegetable organ whose re- sponses are exactly parallel to those of a single plate of the electrical organ of Torpedo and its type. We shall next I study the responsive pecu- ^^\ '%^- r^^f^^f Responses given ' "^ ^ ^ by Leaf of Ftei'ospermutn suoeri- liarities of the vegetable organ folium to Stimulus of Equi-altemat- L , J ing Electrical Shocks at Intervals ot ^nose responses correspond j Minute with those of the organ of McLleptei'urus. It has been mentioned that the posterior surface Ipf each single element of this electrical organ is regarded as consisting of a glandular, rather than a muscular, modification. Among corresponding leaf-organs, then, the carpellary leat pf Dillenia indica might, as we also saw, be taken as the type, its posterior surface being glandular. Or the analogy will be still more perfect if we take as the vegetal type the pitcher of Nepenthe. Here the internal or posterior surface IHb richly provided with glands. The next point to be deter- l^fcnined is whether, in these cases also, on excitation the ■ "responsive current is from the posterior surface to the anterior, as in the electrical element of Malepterurus. And on sub- jecting them to equi-alternating electrical shocks, I found ■ 256 COMPARATIVE ELECTRO-PHYSIOLOGY this to be the case. The responsive current here flowed from the glandular posterior to the non-glandular anterior surface. From this experiment we see that a glandular surface is exceptionally excitable, a conclusion which will be found to be supported by the numerous experiments on glandular organs in general, to be described in Chapter XXIV. I give in fig. 163 a series of photographic records, obtained on excitation of Dillenia indica. In the next record (fig. 164) are seen the responses given by the pitcher of Nepenthe. Fig. 163. Photographic Record of Re- sponses of Carpel of Dillenia indica Natural current from posterior to an- terior, and responsive current from anterior to posterior surfaces. Fig. 164. Photographic Record of Normal Responses given by Pitcher oi Nepenthe, under Equi- aiternating Electric Shocks Responsive current from internal glandular to external non- glandular surface. Note ten- dency to multiple response. An interesting fact to be noticed in the latter is the tendency to multiple response. Similar results were also obtained on taking any single scale of the bulb of Uriclis lily about the time of flowering. In each of these the lower or outer surface is invested with a more or less dry and glistening membrane, while the upper or concave is moist and glanduloid. The moisture observed inside each scale is in fact exuded from this inner surface. On subjecting one of these scales, then, to the electrical THE LEAF CONSIDERED AS AN ELECTRIC ORGAN 257 excitation already described, it is found that a very strong- responsive current is obtained, whose direction is, as in the last case, from the glanduloid to the non-glanduloid surface. The effect of the serial arrangement, again, in enhancing the electro-motive force — as seen in the pile-like arrangement of the electrical organ of fishes — may be exemplified, in the parallel instance of the plant-organ, by means of the superposed scales of the bulb, as found in nature. The bulb may be divided longitudinally into halves, of which the right-hand half is mounted, for experiment, with the scales vertical. It will be understood that all the glanduloid surfaces here face the left, while the non-glanduloid are l^fcurned to the right. Thus the left aspect of this pile corresponds to the head aspect of the organ of Malepterurus. I The latter, on excitation, responds by a current in the jdirection of head to tail — that is to say, from glandular to non-glandular ; and similarly, in the pile-like half-bulb of Uriclis lily, the responsive current is from glanduloid to non- glanduloid — that is to say, from left to right. I^P Another interesting way to perform the same experi- ment is without making any section of the bulb. We take a bulb of Uriclis^ with the peduncle rising out of the Iiiddle. When this hollow peduncle is cut across, it allows of n electrical connection being made with the centre of the iterior of the bulb. An equatorial belt makes the second, r outer, connection. On subjecting this to equi-alternating bocks, the resulting response will be found to be from the inner surface to the outer, through the numerous intermediate scales, the individual effect in each being concordant and iKdditive. H We have thus seen how the response of a leaf gives us an ■nsight into the action of a plate of an electrical organ ; how ^■he differential excitabilities of the two surfaces give rise on ^■timulation to an induced E.M.F. as between the two ; how ^a nervous and indifferent-tissued surface will give rise to a response in one direction, and a glandular and non-glandular I 258 COMPARATIVE ELECTRO-PHYSIOLOGY in the other ; and finally how, by a serial arrangement, the terminal electro-motive effect becomes enhanced. The light thus thrown on the two types of response, known to occur in the electrical organs of fishes, is evident. Further con- siderations, relating to the theory of electrical organs, will be given in detail in the next chapter. CHAPTER XX THE THEORY OF ELECTRIC ORGANS Existing theories— Their inadequacy — The 'blaze-current' so called — Response Iuni-directioned, to shocks homodromous or heterodromous, characteristic of electric organs— Similar results with inorganic specimens — Uni-directioned response due to differential excitability —Electrical response of pulvinus of Mimosa to equi -alternating electric shocks — Response of petiole of Afusa — Of plagiotropic stem of Ctuurbita — Of Eel — The organ-current of electric fishes — Multiple responses of electrical organ — Multiple responses of Biophytum. NE of the most perplexing problems in connection with, tne phenomena of electrical organs is the question as to I whether the activity of such organs is specific — that is to say, ■|>eculiarly characteristic of them — or falls into line with the other electro- motive reactions observed in animal tissue. Many arguments have been brought forward for and against the identity of these phenomena with the excitatory reactions tf the nerve and muscle. From the experimental results which I have described, owever, it would already appear that such reactions as these of the electrical organ are not specifically characteristic; even of the animal structure, but may equally well be observed in plant tissues. It is therefore essential, if we are to determine that basic reaction which is common to all alike, that we IH^hould find a wider generalisation than has hitherto been contemplated. This basic reaction, as we have already seen, depends upon the differential excitability of an anisotropic organ, and this aspect of the case we are now about to study in greater detail. Before doing this, however, we shall briefly glance at various theories which have been suggested, but are generally admitted to be inadequate. I 26o COMPARATIVE ELECTRO-PHYSIOLOGV Bell, for example, thought it might be possible to explain * the discharge of the electrical organ solely by the negative variation of the nerve-current, concomitant with innervation. In this arrangement the dorsal surface of the electrical plate (of Torpedo) would, at the moment of innervation, become positive, the ventral surface negative, as actually occurs.' As against this, it was pointed out by Du Bois-Reymond that this hypothesis in the first place predicates the existence of a current of rest, caused by the * natural ' cross-sections (acting like artificial sections) of the nerves in the plates, and accordingly heterodromous to that of the discharge. Instead of this permanent current — which must correspond in E.M.F. with the discharge, if the nerve-current is to disappear in the negative variation — there is only an inessential P.D. during rest, and the resulting ' organ-current ' is always homodromous with the discharge.* Even had these objections not existed, however. Bell's hypothesis would have failed to explain the electrical action of MaUpterurtis, Du Bois-Reymond himself tried to explain the action of the electrical organ, ' not by the negative variation of the nerve current, but by a process in the electrical plates transformed from muscles, comparable with the negative variation of tJu muscle-current y as set forth from the standpoint of the pre-existence theory.' Here, however^ it is perhaps sufficient to point out that the pre-existence theory, on which the hypothesis was based, is now held to be invalidated. There remains only the Chemical, or Alteration Theory, which associates all electrical changes with the corresponding chemical processes of assimilation and dissimilation. But it has not been made clear in what way these can bring about the characteristic discharge of the electrical organ. There is another point, not altogether unrelated to this subject, which may be dealt with on the present occasion, I allude to the so called 'blaze current ' of Dr. Waller. By this is meant an after-current in the same direction as the ' Biedernuinn, Eltctro- Physiology (English translatioD), vol. II. p. 462. IK THE THEORY OF ELECTRIC ORGANS 261 exciting current. It is, in, fact a new name for that phe- nomenon which Du Bois-Reymond indicated as * positive polarisation-current' Du Bois-Reymond had also shown that this particular effect was most markedly exhibited when the functional activity or ' livingness ' was at its highest. Under opposite conditions, again, it would disappear. The intensity of this homodromous after-effect was thus dependent on the degree of vitality of the tissue under experiment. Hermann and Hering, however, afterwards showed that what Du Bois-Reymond called * positive polarisation ' was in reality ^excitatory reaction. These excitatory effects are known to caused by either the anode or the kathode * ; and I have, the course of the last chapter, demonstrated the fact that It is the differential excitability of a tissue which determines juch uni-directioned response. It is difficult, therefore, to see the necessity of a new name for these phenomena. Dr. aller himself, however, offers the following as an important ;ason : ' The great mass of living things, whatever else they may give and take from their surroundings, take oxygen and give carbonic acid ; they may live slowly or they may live quickly — sluggishly smoulder or suddenly blaze. A muscle at rest is smouldering : a muscle in its contraction is blazing ; the consumption of carbohydrate and the production of COg, never absolutely in abeyance, even in the most profound state of rest, are sharply intensified when the living machine puts forth its full power, and there is then a sudden burst of heat, and an electrical discharge. . . .' ^ This amounts to another way of saying that the cause of the excitatory galvanometric effect is some explosive dis- similatory change, a view which I have already shown in * Within a given " physiological " range of strength of current the negative athodic must, equally with the positive anodic, be designated an "irritative" after-current, due entirely to " polar current-action." ' — Biedermann, Electro- I'hysiology (English translation), vol. I. p. 448. 262 COMPARATIVE ELECTRO-PHYSIOLOGY previous chapters to be quite untenable. I shall presently describe experiments which will further show that galvano- metric responses, not to be distinguished from this, take place when there is no possibility of any consumption of carbo- hydrates or production of COg. The fact, however, that the excitatory after-effects de- scribed, disappear on the death of the tissue, has led Dr. Waller to put forward the generalisation that this so-called * blaze-current ' is the final distinction between living and non-living matter. His formula, with regard to this, is, ' If the object of examination exhibits blaze in one or in both directions, it is living.' He admits, nevertheless, that a sub- stance which is undoubtedly living will not always exhibit the ' blaze-current' But it is contended that the occurrence of ' blaze ' is an undoubted ' sign of life,' and that thus a strong distinction is to be made between vitalistic and non-vitalistic, or physical, reactions. Hence, as there is supposed to be no excitatory reaction possible in non-living or inorganic matter, it would follow that electrical shocks passed through such a substance, in either direction, should give rise only to those counter-polarisation currents which are known to physicists. In such cases, on reversing the direction of the shock, the direction of the after-current is also reversed ; but in the living substance, it is maintained, the case is quite different. If the direction of the shock be here reversed, the after-current will still appear, with direction unchanged, because in this latter instance it is not generated by the shock, but is, on the contrary, an inexplicable function of the living material, set in action by it, in the same way as a loaded gun is fired by pulling the trigger. The possibility of obtaining from the given substance such a uni-directioned after-current, independently of the direction of the shock, is thus to be taken as the test and token of * vitality.' Now, while it is certainly true that the domain of physio- logical phenomena has not yet been so thoroughly explored as that of the physical, it is nevertheless equally true that no one could venture to claim that even physical phenomena THE THEORY OF ELECTRIC ORGANS 263 had up to the present been exhaustively studied. It is, then, somewhat hazardous to declare that because a particular phenomenon has not yet been observed to occur in inorganic matter, it is by that fact demonstrated to be hyper-physical in its nature, and must be relegated to the different and mystical category of the exclusively vitalistic. The very foundation of such a statement would be swept from under it, the moment it was shown that the same phenomenon followed, under the same circumstances, in conditions which were admitted to be purely physical. I have shown, it will be remembered, in the previous chapter, that the uni-directioned response to electrical shocks in either direction was due to the differential excitability of the structure. The response of any aniso- tropic organ would .always be from the more to the less excitable, the more excitable becoming relatively galvano- metrically negative. There may here be various cases of excitation, all giving results of the same type, say, a respon- sive current from B to A. The first is that in which, on excitation, both B and A become galvanometrically negative, A being the less so of the two. In the second case, the excit- ability of A being slight, or negligible, B alone becomes negative. And in the third case, excitation induces positivity of A and negativity of B. In all these cases the relative negativity of B being greater, the responsive current will flow from B to A. The resultant current is made up, in the first case, by subtracting the galvanometric negativity of A from that of B ; in the second case, it consists of the galvanometric negativity of B, that of A being zero ; and in the third case, it is produced by the addition of the effect at A to that at B. Examples of the last of these will be found in certain animal and vegetable skins, described in Chapter XXII. These being the conditions, then, for the induction of the uni-directioned responsive current, it appeared to me probable that the same result could be obtained with inorganic substances, provided that the specimen were so 264 COMPARATIVE ELECTRO-PHYSIOLOGY ir prepared as to be anisotropic, one side having a greater potentiality of galvanometric negativity under excitation than the other. In that case, further, it was clear that the strongest resultant current would be obtained, if one surface of the structure became galvanometrically positive, and the other negative, on excitation. I have already stated, in Chapter T., that different inorganic substances give electrical responses of opposite signs. Thus the response of lead is positive, while that of brominated lead is negative. If, then, we take a lead wire A B, clamped in the middle at C, as represented in the upper diagram (fig. 165), and stimulate the right-hand end B, say by mechanical vibration, a responsive current will be induced, which will flow to- wards the stimulated, B thus becoming gal- vanometrically posi- tive. The same will be the case with A, on stimulation. When both A and B are simul- FiG. 165. Responsive Currents in Lead Wire Upper yf^«;-XejaJc. The com- plete law, as will be shown in a later chapter, is complicated by the fact that the result depends on the intensity of the electro- motive force. With feeble and again with excessive E.M.F., the actual facts are the opposite of conclusions arrived at by Pfliiger. Under these circumstances it would appear at first sight impossible that any reliable results could be obtained by the employment of the electrical form of stimulation. I shall now, however, proceed to show in what manner all these difficulties may be overcome, and the electrical form of stimulus made extremely reliable. This will perhaps be best understood if we take a concrete example, Let us suppose II 274 COMPARATIVE ELECTRO-PHYSIOLOGY that a single electrical shock of moderate intensity enters an isotropic tissue at the point A, and leaves it at B ; A will then be the anode, and B the kathode. Here the true excitatory effect is found to take place only at the kathode, probably because the anode-break excitation takes place at much higher intensities of E.M.F. than the kathode-make. At these higher intensities, then, the anode-break effect also will occur. At an excessively high E.M.F. again, these relations, for reasons already explained, may undergo reversal. The point of such reversal would depend on the nature and excitability of the tissue. Though, for all these reasons, the relative excitations of A and B remain a matter of doubt, yet we may be sure of the excitation of both points, if two, or any equal number, of exactly equal shocks be sent through the tissue in opposite directions, in rapid alternation. If, again, instead of two alternate shocks only, we give n alternating shocks, abso- lutely equal, and if, further, the natural excitability of the two points A and B have been the same, then there will be nothing to distinguish the excitatory effect induced at A from that at B. In other words, the two excitations will be exactly equal. These strictly equal and opposite alternating currents, more- over, can have no resultant polarisation-effects, for the effect arising from an induction shock, in either direction, will be counteracted by that caused by the opposite. We thus see that by the employment of this method the only change induced at the two electrodes will be the excitatory change, the physical polarisation-factor being eliminated. Thus, on subjecting two points, A and B, to equal stimulation, the induced galvanometric negativity, at both the points, will be equal, if the natural excitabilities of the two have been the same. But if the tissue be anisotropic, and the natural excitability of one point, say B, greater than that of A, then we shall obtain a resultant responsive current, which will flow in the tissue from the more excited B to the less excited A, the induced galvanometric negativity of B being relatively the greater. We have here what is merely a repetition, by electrical stimulation, of the results which DETERMINATION OF DIFFERENTIAL EXCITABILITY 275 were described in Chapter IX., as obtained by thermal and mechanical stimulus. How perfect and how consistent, due precautions being taken, these results may be rendered, will be seen from the numerous records in this and the following chapters. I have postponed till now the consideration of the mode of application of these alternating shocks. The usual alter- nating current from a Ruhmkorff's coil would be entirely unsuitable for delicate and crucial experiments ; first, because the excitatory values of the slow make- and quick break-shocks are unequal ; and, secondly, because such currents leave their residual polarisation effects. I^b These defects I have been able, as stated in the last chapter to avoid, by quick reversals of the primary current which actuates an induction-coil. When the primary current is reversed from the plus to the minus direction, we obtain an induction-current due to magnetic varia- tions of lines of force, from, say, plus n to minus n. When the primary current is re-reversed, from minus to plusi we obtain an opposite induction-current, due to magnetic variation, from minus n to plus n. It will be seen that if these reversals of the primary current are made with equal rapidity, the alternating induced currents will be equal and opposite. The reversals are accomplished by means of a Pohl's commutator, worked up and down by a crank, in connection with an electric revolving motor (fig. 170). The intensity of the induction-shock may be varied by sliding the secondary nearer to, or further away from, the primary. Having now described the general means of producing equi- alternating electric shocks, it still remains to explain two distinct methods of applying them, for the determination of the differential excitability of the tissue. These may be distinguished as : (i) the Method of the After-Effect ; and (2) the Method of DiRECT-ErFECT. According to the first of these —the method of the after-effect — the tissue is excited for a definite length of time, and the excitatory effect observed, by connecting it immediately afterwards with the II 276 COMPARATIVE ELECTRO-PHYSIOLOGY galvanometer-circuit. The manner in which this is done will be understood from fig. 170. We have a highly insulating electrical key, K, of ebonite. P and Q are connected with two points A and B of the tissue, whose relative excitabilities are to be determined. A spiral spring keeps the key down, con- necting the two points in the specimen with the galvanometer. Any existing difference of potential, as between the two points Fig. 170. Experimental Arrangement for Determination of Excitatory After-effect of Equi-alternating Electrical Shocks M, electrical motor working Pohl's commutator for alternate reversal of current in primary, p. Note that the connecting-rod, A, works simply up and down, causing reversals of current ; s, secondary coil ; K, ebonite key kept, down by elastic spring, the two surfaces of the specimen being thus in circuit with galvanometer, G. When key is pressed, these are put in circuit with exciting coil, s. When key is released, after-effect of excitation on specimen exhibited by galvano- meter deflection, c, the compensator. . is balanced by the compensating potentiometer c. Under these circumstances, the galvanometer spot of light would remain steady, whether the key was up or down. By pressing the key K, the galvanometer-circuit is broken, and the tissue is put in series with the exciting circuit S, which forms the secondary of the induction-coil. When the key is again released, the galvanometer-circuit is rapidly made, at a definite short interval after the cessation of the exciting shock, and the resulting deflection of the galvanometer indicates the differ- DETERMINATION OF DIFFERENTIAL EXCITABILITY 277 ential excitation as between A and B. In this way records may be taken of a series of the after-effects of brief excitations, at intervals of, say, a minute. The direction of this responsive current, which in the tissue is from the more to the less excitable, enables us to determine the relative excitabilities of the two points, A and B. lib Much more delicate is the -second method, that, namely, ■^vhich depends on the record of the Direct-Effect of equi- alternating shocks ; but for its perfect working, certain difficulties have to be overcome. One of the first conditions to be fulfilled lies in the perfect equality of the alternating shocks. The importance of this will be understood on I^Bbserving the effect of the alternating shocks given by a Ruhmkorff's coil, when actuated by a vibrating hammer. Here, the make- and break-shocks are of unequal intensity and duration, and the following sources of disturbance come into play: (i) a galvanometric drift in one direction or the other ; (2) a resultant inequality of polarisation- effects ; and (3) the inequality of the excitatory values of the two shocks. IBF The galvanometer-drift, owing to the inequality of the mduction shocks, becomes very troublesome, when we have to employ, as is necessary, an instrument of high sensibility. If the differential excitability of the specimen be very great, this drift may be masked by the predominant excitatory effect. In other cases, however, the excitatory effect itself may be overpowered by the drift. The difference of intensity as between the make- and break-shocks in the Ruhmkorff's coil described, thus becomes a strongly dis- l^urbing element. The necessity to make the two shocks Ufcsolutely equal will be understood when we find that alternating telephonic currents, which are generally re- garded as equal and opposite, induce a drift of the galvanometer in one direction or the other, on account of a slight difference of intensity between the two alternating ^^rrents. ^B The difficulty arising from inequality of polarisation- 278 COMPARATIVE ELECTRO-PHYSIOLOGY more important, moreover, is the disturbing factor last mentioned, that of the inequality of excitatory values as between the make- and break-shocks themselves. This element of uncertainty is very clearly seen in the experi- ments of Von Fleischl on the response of nerve. The resulting deflection he found to be in the direction of the break-shock. The explanation of this phenomenon has hitherto been regarded as a matter of great difficulty, authorities being much divided on the subject. In these experiments, alternating currents from a Ruhmkorfif's coil are sent through the galvanometer and the nerve, in series. The two points on the nerve, A and B, are presumably of equal excitability. A make-shock of relatively lower E.M.F. passes, say, from A to B, followed by a break-shock of higher E.M.F. in the opposite direction, from B to A. Confining our attention to the excitatory effects of these shocks, we have at A, during make and break, the following four effects : (a) feeble anode-make ; {b) feeble anode-break ; {c) strong kathode-make ; and {d) strong kathode-break. At B, on the other hand, at the same time, we shall have : {a') feeble kathode-make ; {b') feeble kathode-break ; {c') strong anode- make ; and {d') strong anode-break. Now, since the excitatory value of anode-break is probably the stronger and more persistent, and since the intensity of this effect will depend, within certain limits, on the intensity of the anodic shock, it follows that the {d') or strong anode-break effect at B will, as a general rule, be the most conspicuous of these. That is to say, as a result of all these excitatory effects in combination, greater galvanometric negativity will be induced at B than at A, the responsive current being thus from b^a, in the same direction as the break shock, which was the actual result. In any case, whatever may have been the cause of this, it is clear that the employment of such unequally exciting shocks of make and break would be fatal to any attempt to determine accurately the natural difference of excitability as between the two points. I may state here, that when I have employed absolutely equal alternating shocks on a specimen DETERMINATION OF DIFFERENTIAL EXCITABILITY 2/9 of nerve, I have obtained no resultant deflection whatever, showing that such shocks induce exactly equal excitations in an isotropic tissue. But if the excitability of one of the two points be first abolished by killing, then a definite resultant responsive current is obtained, from the excitable living to the inexcitable dead. So perfect were in fact the results secured by^ means of these equi-alternating electric shocks, that I was desirous not only to detect, but also to record photographically, the responses thus obtained. In this a certain difficulty is experienced, inasmuch as the alternating shocks are apt to render the recording spot of light tremulous, and thus to spoil the photographic im- pression. This may, however, be overcome by making the alternation frequency so high, in reference to the period of the needle or suspended coil of the galvanometer, that the unsteadiness of the deflection ceases. IB I shall now describe the practical means employed to obtain equi-alternating shocks of any frequency that may be desired. This I have been able to do in several ways, and^ among others, by using a Rotating Reverser. This consists of an ebonite disc, on the periphery of which there are strips of metal of equal breadth, and separated from each other by equal distances. The odd strips (i, 3, 5, and so on) are connected together and led to a metallic ring on the left of the disc. The same is done with the even strips, which are led to the right. The two electrodes of a battery are led through a key, K, to these two metallic rings and are con- nected with them by means of brushes. Thus one ring, with all the odd strips, is connected, say, with the positive, and the other, with all the even strips, with the negative pole of the battery. The current is led off* by a second pair of brushes, placed diametrically opposite to each other on the disc, in the primary circuit, P, of an induction coil (fig. 171). Let us suppose the upper brush to be connected with an odd strip, the lower will then be connected with an even. The current in the primary coil now flows in one direction. When the disc is rotated, so as to bring the next pair of strips in I 28o COMPARATIVE ELECTRO-PHYSIOLOGY contact with the brushes, the upper will then be connected with the even strip and the lower with the odd. Thus the direction of the current will be reversed, and rapid rotation of the disc will give rise to equi-alternating currents in the primary of the induction coil. This will in turn induce equi-alternating induction currents in the secondary, the intensity of which can, as already said, be varied within wide limits by appropriate changes of distance between the primary and the secondary. The number of strips in the apparatus used is fifty, and when the disc is rotated, by Fig. 171. Method of Direct Effect of Excitation by Equi- alternating Shocks R, rotating reverser, in circuit with primary coil, P. Duration of stimula- tion determined by metronome, M. s, secondary coil in series with specimen and galvanometer. means of an electrical motor, at a rate of one revolution per second, there will be fifty alternations of current in a second. The duration of the application of the stimulating shock to the tissue is regulated by a metronome, which completes the primary circuit for a definite short length of time. When the metronome, M, is so adjusted as to complete the circuit for '5 second, then a stimulus of that duration will be imparted at each stroke. A second interrupting key, not shown in the figure, is included in the circuit. When this key is closed, a single beat of the metronome gives a stimu- lating shock of "5 second's duration. The key is now opened DETERMINATION OF DIFFERENTIAL EXCITABILITY 28 1 for one minute for recovery. In this way, records of response and recovery are obtained, at intervals, say, of one minute. Another very effective means of producing equi-alter- nating shocks is by the employment of an alternating- current dynamo, driven by an electrical motor, M (fig. 172). The alternating current is led to the primary of a Ruhm- korffs coil in the usual manner. The motor is driven by an electrical supply from the street mains, its speed being adjusted by a regulation of the current, which is effected by P' Fig. 172. Excitation by Equi-alternating Shocks M, motor, rotating armature of alternating-current dynamo, D ; R, liquid rheostat, in circuit with street-mains, for regulating speed of rotation of motor ; p', idle coil ; P, primary coil ; i, resonating index ; s, secondary coil, in series with specimen and galvanometer. Duration or excitation determined by pressure of key, k. n electrolytic rheostat, R. As the dynamo is provided with a permanent horse-shoe magnet, the intensity of the alter- nating current is determined by the speed of rotation of its armature. If the speed be kept alw^ays constant, the number of alternations will also be constant, and the ex- l^piting value of the electric shocks will depend simply upon the distance of the primary from the secondary. It is thus possible day after day to use the same intensity of stimu- lation, and thus to compare the relative excitabilities of ■ 282 COMPARATIVE ELECTRO-PHYSIOLOGY different tissues. The constancy of the speed of rotation of the alternating-current dynamo is secured by means of the resonating index, l. This consists of a short steel spring with a long index. When the frequency of alternation is the same as the natural period of vibration of the spring, the resonator is thrown into strong sympathetic vibration. At first the rheostatic resistance, which determines the speed of the motor, is made slightly too large. The movable plate is now gradually brought nearer, till the proper speed has been arrived at, and this point is at once indicated by the induced vibration of the resonator. A further difficulty has to be overcome in the main- tenance of the uniformity of speed. When the open circuit of the alternating dynamo is closed, by the interposition of the primary of the Ruhmkorff's coil, the speed undergoes a sudden diminution, owing to the work which the dynamo has now to perform. In order to avoid this fluctuation, then, the dynamo circuit is kept closed by means of an idle primary coil, P', which is a duplicate of the primary, P, of the Ruhmkorff's coil. When the key, K, is pressed, the alter- nating current is transferred from P' to P. There is thus no fluctuation in the speed of the dynamo, and the duration of the closure determines that of the stimulus. I may mention here, that instead of employing a separate motor to drive the alternating-current dynamo, I have sometimes used, with equal success, a motor transformer, giving rise to alter- nating currents. It is easy to construct a very compact and •portable form of this latter apparatus. In this manner we may apply uniform stimuli of equi- alternating shocks at regular intervals of time, say of one minute. The usual preliminary test of the successful elimination of all sources of disturbance may here be made in the following way. The kaolin ends of the non-polaris- able electrodes are connected with each other, without the interposition of a specimen, and alternating shocks from the secondary are passed through the circuit. These should give rise to no deflection in the galvanometer. It may be DETERMINATION OF DIFFERENTIAL EXCITABILITY 283 said here that I use a D'Arsonval type of galvanometer, in which, instead of a suspended needle, we have a suspended coil. There is thus here not even the remote contingency of disturbance which might arise from the demagnetisation of the magnetic needle. Having thus tested, by null action, the symmetry of the electrodes and the galvanometer, the differentially excitable tissue, say the sheathing petiole of Musa, is interposed, with its concave and more excitable surface upwards. On now applying excitation by equi- alternating shocks, the responsive current will be found to flow downwards, from concave to convex, giving a deflection of the galvanometer, say to the right. And this deflection l^^ill continue to be to the right, even if the battery current ^fig. 171) be reversed by means of key K. The direction of the excitatory current, moreover, depending solely, as it does, IHbn the relative excitabilities of the two surfaces of the specimen, will remain constant, even if the connections with the secondary coil, S, be reversed. The zinc rod, N, of the non-polarisable electrode in connection with the concave surface (fig. 172) has thus, up to the present, shown induced galvanometric negativity, the galvanomctric deflection being to the right. But if we exchange the zinc rods of the non- polarisable electrodes, it will then be N' which will be con- nected with the more excitable concave surface, and it will now be this electrode N' which will show galvanometric nega- tivity. This reversal of the galvanometer deflection with the reversal of the electrodes affords additional confirmation of the greater excitability of the concave surface of the specimen of Musa. Im In these experiments the existing current' of rest may t)e balanced previously by a potentiometer. But this is not absolutely necessary. I give below a series of records obtained with a specimen of the sheathing petiole of Musa (fig- I73)> in which we know the inner or concave surface to be more excitable than the outer or convex. The responsive current is seen under this form of electrical 284 COMPARATIVE ELECTRO-PHYSIOLOGY thermal stimulation, to flow from the more excitable concave to the less excitable convex. In order next to demonstrate the physiological character of these responses, I subjected the tissue to the action of chloroform, and the record in the second part of the figure shows the consequent depression of the response. The great delicacy and pliability of this mode of applica- tion of stimulus enable us to attack many difficult problems, on the difference of excit- ability between two points in a tissue, with perfect ease. To -how many distinct in- vestigations it can be suc- cessfully applied will be set forth in detail in succeeding chapters. As there is nothing to prevent the two exploring electrodes from being applied on any two points, however distant, of the same organism, it is seen that we have here a means of determining, not only the differential excit- ability of any two points of the same organ, but also that of any two organs of the same specimen. For the present I shall, however, content myself with giving a few instances only in illustration of the ex- treme delicacy of this method in detecting physiological differences as between two points. We shall first turn our attention to those physiological modifications which are due to the a-symmetrical action of the environment on the organism, and here we shall select the case of the plagiotropic stem of Cucurbita. We have seen that in the recumbent stem of this plant the tissue of the upper side is rendered relatively fatigued by the con- tinuous action of sunlight, and thus becomes permanently less excitable than the lower side. We have also found that Fig. 173. Photographic Record of Re- sponse of Petiole of Mtisa to Equi- alternating Electric Shocks, before and after Application of Chloroform. DETERMINATION OF DIFFERENTIAL EXCITABILITY 285 while the natural resting-current was from the less excitable upper to the more excitable lower side, the responsive cur- rent under mechanical stimulation was in the opposite direc- tion — namely, from the lower to the upper (p. 112). Using now the electrical form of stimulus, we obtain results which are identical. Fig. 174 gives a series of such responses under equi-alternating electrical shocks. Curiously enough, as pointed out in the last chapter, I have detected a similar plagiotropy in the case of the eel. The head of the fish was cut off, and voluntary action thus eliminated ; electrical connections were then made, after a period of rest, with the dark dorsal upper surface, and the colourless skin of the ventral or lower. A natural current was now found to flow from the upper MWL IIG. 174. Photographic Record of Responses of Plagiotropic Stem of Cucurbita to Equi- alternating Electric Shocks Irection of responsive current from ventral to dorsal surface. Fig. 175. Electrical Responses of Eel to Equi-alternating Electrical Shocks Current of response from ventral surface to dorsal. surface to the lower, as in the case of the plagiotropic stem of Cucurbita. Electrical excitation was now applied, and the result was a responsive current from the more excitable lower to the less excitable upper surface again, as in the QdL^^ o{ Cucurbita. In fig. 175 is seen a series of records in illustration of this. Another investigation which I thought might be interest- ing had reference to the variegated colouring of certain foliage leaves. A striking example of this is found in the ■ 286 COMPARATIVE ELECTRO-PHYSIOLOGY tropical elephant creeper {Pothos), the rich green of which is barred by longitudinal streaks of milk-white. This dis- tribution of colour is found even in the youngest and most vigorous leaves. The question whether such colouring was accidental or associated with physiological differences could, I thought, be determined by the delicate mode of investiga- tion which was now at my disposal. On making electrical connections, then, with the green and white portions of a leaf, I found that the natural current of rest was from white to green through the tissue, and, on further testing the differ- ential excitability in the usual manner, the responsive current was observed to flow from the green to the white. This showed that pallidity was here associated with a depressed physiological condition. CHAPTER XXII RESPONSE OF ANIMAL AND VEGETAL SKINS Jurrents of rest and action — Currents in animal skin — Theories regarding these — Response of vegetal skin — Stimulation by Rotary Mechanical Stimula- tor — Response of intact human skin — Isolated responses of upper and lower surfaces of specimens — Resultant response brought about by differ- ential excitability of the two surfaces — Differences of excitability between two surfaces accounted for — Response of animal and vegetal skins not essentially different — General formula for all types of response of skin — Response of skin to different forms of stimulation gives similar results — Response to equi-alternating electric shocks : (i) Method of the After Effect ; (2) Method of Direct Effect — Response of grape skin — Similar response of frog's skin — Phasic variation of current of rest induced as result of successive stimulation in (a) grape skin ; (d) frog's skin ; {c) pulvinus of Mimosa — Phasic variation in autonomous mechanical response of Des- viodhim ^jrawj-— Autonomous variation of current of rest — True current of rest in skin from outer to inner— This may be reversed as an excitatory after effect of preparation — Electrical response of skin of neck of tortoise — Electrical response of skin of tomato —Normal response and positive after- effect—Response of skin of gecko — Explanation of abnormal response. In this and the next few chapters it is my intention to make an inquiry into the responsive peculiarities of the skin, epithelium, and glandular tissues, alike in plant and animal. By the study of such simple cases as are found in plants, it should be possible to obtain a clear insight into the various f"actors which go to make the corresponding phenomena l^ki animal tissues so complicated and obscure as to be |B|ifficult of reconciliation with each other. y^k It is not possible in a short space to give any but the [^Briefest summary of the work hitherto done on this extended subject in animal physiology. All that can be attempted is to indicate some of the leading theories and results, at the same time drawing attention to those outstanding questions 288 COMPARATIVE ELECTRO-PHYSIOLOGY employed in the investigation of plant phenomena, moreover, have proved so highly satisfactory that records will be given of the results obtained by their means in the case of animal tissues also. And this will, I hope, show the great reliability and simplicity which it is thus possible to intro- duce into the investigation as a whole. With regard to the electrical effects in animal skin, epithelium, and glands, the inquiry resolves itself into the determination of, (i) the direction of the current of rest ; (2) that of the excitatory current ; and, lastly, (3) a consideration of theories regarding these. The first of these, the current of rest, was found by Du Bois-Reymond and Engelmann in the skin 'of frog to be * ingoing ' — that is to say, passing from the outer surface to the inner. Hermann also found a similar current in the skin of eel. He regarded the source of electro-motive action as lying in the partial mucin-metamor- phosis of single cells. From the fact that in the toad, where the ingoing current is specially strong, the skin glands are vigorously developed, and from the discovery by Rosenthal that in the mucous glands of the stomach the current is also ingoing, it was assumed that the observed electro-motive forces were due to the glandular nature of the tissues. The skm current of the frog and of the fish, and the glandular current of the stomach, are thus usually regarded as due to the same cause. There is, however, a serious discrepancy in this view, inasmuch as, while local stimulation of the upper surface of the frog's skin induces a positive change, a similar stimula- tion of an unmistakably glandular surface is found to bring about a negative. If then the electrical effect on the skin of frog be the same as on a glandular surface, the dis- crepancy of their responsive reactions becomes inexplicable. As regards the excitatory change, very diverse results have been recorded when stimulus has been applied indi- rectly—that is to say, through the nerve. This fact is not to be wondered at, since the responsive effects are subject, as will be shown, to numerous modifying influences. It is generally RESPONSE OF ANIMAL AND VEGETAL SKINS 289 supposed, in the preparations made for these experiments, that it is one surface only which is electro-motive. I shall show, however, that the responsive effects are brought about by the differential excitability of the two. This response, again, is modified by relative changes induced in the two surfaces. And, in addition to these, still further compli- cations are introduced when the stimulus is indirect — that is to say, applied through the nerve. In this case, the relative excitations of the two surfaces will be determined by the particular distribution of the nerve-endings. Again, we shall see that, in an isolated preparation, the nerve itself is liable to undergo certain changes by which its trans- mitted effect may be modified even to reversal (p. 530). Thus, so long as it remains highly excitable, the transmitted effect is one of true excitatory galvanometric negativity. But l^kith physiological depression, the conductivity of the nerve ^s very much lowered, and the effect transmitted becomes reversed to positivity. IB, For all these reasons, if we wish to study the specific reactions of skin, epithelium, and gland preparations, it is I J^etter to do so by observing them under direct stimulation. I^p Engelmann, in studying the responsive reactions to direct stimulation of frog's skin, found a negative variation of the current of rest. Since the latter was naturally ' ingoing,' as regards the upper or epidermal surface, this meant that the responsive current was 'outgoing.' Reid, again, work- ing on the skin of the eel, obtained ingoing response, or positive variations of the resting-current, by single induction shocks in either direction. Biedermann, in the mucous membranes of the tongue and stomach, obtained both positive and negative variations of the current of rest. Waller, using single induction shocks in either direction, found in the digestive mucosa both ingoing and outgoing responses, the former being much predominant. Even under the simple conditions imposed by direct stimulation, then, the results obtained are seen to be in- Insistent. They would appear to show both that the 290 COMPARATIVE ELECTRO-PHYSIOLOGY responsive effects in the different preparations are different and that, even in the same preparation, they may be reversed under unknown changes of circumstances. It appeared to me, as already said, that much h'ght might be thrown on the questions thus raised by means of an investigation carried out on plants. The most perfect method of experiment here would consist in observing the separate responsive effects on upper and lower surfaces of the preparation. Waller employed single induction shocks for this purpose, observing the after-effect. But in this case, the action of polarisation was not excluded. It would thus be more satisfactory, in order to eliminate this unknown element, to employ either a non-electrical mode of stimulus or an electrical form which would leave no resultant polarisation effect. The latter condition, as we have seen, was fulfilled by the employment- of rapidly alternating currents, whose alternating components were absolutely equal. As regards the application of a non-electrical form of stimulus, both thermal and mechanical forms may theoreti- cally be employed. Engelmann and others used heated metals in the proximity of one of the electrodes, for the production of thermal stimulus. This, however, has the disadvantage of thermo-electrical variation, due to unequal heating of the two contacts. Besides this, there is also the effect of a rising temperature, which, as we have seen, is opposite to that of sudden variation, the latter alone consti- tuting the excitatory effect. I have already explained how these difficulties may be overcome by using thermal shocks in which a sudden thermal variation is made to act on both contacts at once. The resultant response thus obtained was shown to be determined by the differential excitability of the two contacts under examination. As regards the mechanical mode of stimulation, previous observers have employed pressure or friction. Such stimulus, however, is at best merely qualitative. If it be applied at the contact itself, objections may be taken to the effect, as RESPONSE OF ANIMAL AND VEGETAL SKINS 291 due to, or modified by, the variation of contact resistance. And if to avoid this the mechanical stimulus be applied, not at the electrode, but at a neighbouring point, the results will be quite different, according as the conductivity of the intervening tissue is great or slight. In the former case they will consist of the transmitted effect of true excitation ; in the latter of the indirect effect, whose electrical sign is the exact opposite. Fig. 176. Rotary Mechanical Stimulator 'Specimen of skin pinned on hinged platform which is pressed against electrodes by elastic india-rubber. Electrodes rotated by cord, c c'. s, antagonistic spring, made of elastic. Enlarged view of electrode seen to the right. T, outer fixed brass tube ; t', inner rotating tube, holding non-polarisable electrode. P, pumice-stone. A perfect method of direct mechanical stimulation has been described in Chapter III., the stimulus being vibrational. But for investigations on limp structures, such as skin, this method is not practicable, and the modification which I am now about to describe is necessary, in order to meet the difficulties of the case. The apparatus consists of a hinged platform, P (fig. 176), on which the specimen is securely pinned. The two electrodes, E and E', rest with a definite pressure on the two points A and B, whose excitatory re- actions are to be studied. These electrodes have at their II u 2 292 COxMPARATlVE ELECTRO-PHYSIOLOGY lower ends projecting pumice-stone cylinders of equal sec- tions, soaked in normal saline. When the electrode E is rotated, the mechanical friction induces local excitation of the point A. B may also be subjected to similar isolated excitation in the same way. In order that such successive excitations may be quantitative and uniform, it is necessary first that a definite area at A or B should be stimulated. In other words, there must be no lateral slip. For this reason the electrodes are passed through tubular holders which, from the description presently to be given, will be seen to allow rotation about a definite vertical axis. The extreme bases of the pumice-stone cylinders are, as has been said, of equal section. The glass electrode tube is tightly fixed, by means of a cork, inside a brass rotating tube. The latter, again, plays inside an outer brass tube, which is fixed. The inner brass tube is provided with two collars, one below and one above, by means of which rotation can take place without up or down movement. A string is also wound round it, by pulling which rotation is produced. The electrodes are perpendicular to the plane of the platform which carries the specimen. It will thus be seen that any variation of the surface subjected to stimulation is prevented. The next difficulty to be overcome is that of liability to variation in the pressure of the contact. It will be remembered that the platform is hinged. It is further held up against the electrodes by the tension of an elastic piece of india-rubber or a spiral spring of steel. This pressure can be regulated to a suitable value, and kept constant. The final difficulty is to apply successive stimuli of equal value, and to render them capable also of graduation from low to high values. This could be secured by rendering the successive rotations of the exciting electrodes equal in number and in time of execution. The intensity of stimulus might then be increased by increasing the number of rotations or the pressure of the electrodes on the specimen. In order to apply successive rotations of definite number, one end of the string wound round the inner brass tube is RESPONSE OF ANIMAL AND VEGETAL SKINS 293 attached to a piece of stretched india-rubber, which is fixed by its other end to the apparatus. The second end of the string is tasselled, after being passed through a fixed ring. This position of the string is adjusted by means of a knot, so that the india-rubber at its other end is ah'eady in a state of tension. When the tassel is now suddenly pulled and let go, it gives rise to a number of rotations in the positive, followed by an equal number of rotations in the opposite direction, the latter work being performed by the stretched antagonistic spring. It should be remembered that a mechanical rotation, whether //«.y or minus, gives rise to the same excitatory reaction. Next, to make the number of rotations definite, let us suppose the inner brass tube to have a circumference say of i cm. If the string be now pulled through a distance of 5 cm., and let go, there will be five rotations in the positive, followed by five rotations in the negative direction. A second knot in the string, at the distance of 5 cm. from the first, exactly limits the length of the pull ; and increase or decrease in the intensity of the stimulus can be brought about by a change in the distance of the regulating knot. The distance between the two electrodes being always the same, the resistance of the interposed tissue remains approximately constant. To nullify any accidental variation, a high and constant external resistance is interposed in the galvanometer circuit. When the excitatory electro-motive variation of the specimen is very great, it is possible to use an external resistance as high as one million ohms. It should, however, be remembered that even if there be any unavoid- able variation of resistance, it will not in any way affect the discrimination of sign of the characteristic electro-motive response. For the excitatory effect at either electrode may be tested by repeating the experiment with the other. The experiments which will be described afforded definite and characteristic records, which were found capable of repetition. The physiological character of such responses was further demonstrated by repeating the experiment, after killing the I 294 COMPARATIVE ELECTRO-PHYSIOLOGY tissue with boiling water, when these electro-motive variations were found to disappear. How very reliable these responses can be rendered is shown by the photographic record in fig. 1 80, which is of very special interest, giving as it does the record of responses afforded by the intact human skin. Turning now to the nature of the response of the skin, it has been found by Engelmann and others, as already said, that in the frog, while the natural resting current is from the outer surface to the inner, the responsive current is from inner to outer. Dr. Waller, again, undertook to analyse the constituent elements of this response, by passing induction shocks along each of the surfaces, first upper and then lower, and in both directions. He then observed the after-effect at one of the excited points, in relation to an indifferent point. In this way he found that an excited point on the upper surface becomes galvanometrically positive, the current being thus outgoing. The inner surface, however, he found to be ineffective. When an induction shock is passed across the tissue, the resultant response from lower to upper is thus, according to Dr. Waller, due to induced positivity of the upper surface. With vegetable specimens, however, such as the outer skin of apple, the results obtained by him were opposite to those of the frog's skin. The responsive currents were here found to be ingoing, the excited point being galvanometrically negative. The explanation offered, in regard to these results, is that living tissues have the peculiarity of responding by ' blaze currents ' to electrical shocks. The use of this phrase^ however, as already said, offers no real explanation ; but even apart from this point, the question remains, Why should the blaze currents, so called, be directly opposed in the cases of frog's skin and of the particular vegetable skins which are mentioned, respectively? In answer to this, the hypothesis put forward by Dr. Waller is, that the difference arises from the different natures of animal and vegetable protoplasms.^ ' * Vegetable protoplasm is in major degree an instrument of synthesis and accumulation, in minor degree the seat of analysis and emission. Animal RESPONSE OF ANIMAL AND VEGETAL SKINS 295 I shall, however, be able to adduce facts and considera- tions from which it will be possible to arrive at a simpler and more conclusive explanation of these phenomena, on the basis of the differential physiological excitability of the two surfaces. I shall show, moreover, that the difference between animal and vegetable protoplasm, thus assumed to exist, has nothing to do with the question. We have seen that when the physiological activity of a tissue is in any way impaired, its normal excitatory re- action of galvanometric negativity is depressed. This may even go sonfaF^s to^cause aTiT^aCttrah^reversal of the response, to I galvanometric positivity, as we found in the case of depressed tissues (p. 84). Taking a vegetable specimen, then, say a hollow petiole or peduncle, we find that the outer surface, which is habitually exposed to the manifold influences of the environment, becomes histologically modified, being much more cuticularised than the inner. Thus these outer and exposed cells generally become reduced in size, and thick- walled, with little protoplasmic contents. Hence, as regards [■Junctional activity, these epidermal cells are in a physio- logical sense very much degraded. We should then expect their excitability to be proportionately lowered in comparison with, say, the inner surface of the same tube, protected as that has been from outside influences. And the variation of physiological excitability thus induced may involve not only the surface, but also the subjacent layers to a certain extent. I^K Theoretically, then, the induced galvanometric negativity of the outer would be less than that of the inner surface,^ I and simultaneous excitation of both, by whatever means produced, should give rise to a resultant responsive current protoplasm is in major degree an instrument of analysis and emission, in minor degree the seat of synthesis and accumulation. The vegetable, in most immediate contact with inert things, combines, organises, and accumulates. The animal, in less immediate contact with inert matter, disrupts, utilises, and dissipates in their fragments organic compounds that it has received ready made from other animals and from plants.' — Waller, Signs of Life, p. 85. t' This refers to normal skin, and not to that in which the surface is typically 296 COMPARATIVE ELECTRO-PHYSIOLOGY from inner to outer. The degree of this diminution of excitatory negativity in the outer surface, moreover, cul- minating as this may in actual positivity, will depend upon the extent of its transformation. In connection with this it should be remembered that, in order to bring out the differential excitabilities of the two surfaces, it is necessary to apply localised stimuli of an intensity not too excessive. For, if the stimulus be very strong, there is always a possibility of its affecting deeper layers of the tissue and thus causing complications in the resultant excitatory changes. The intensity of stimulus which may be safely used without bringing about s uch complications will depend on the conductivity of the tissue. Epidermal cells are, generally speaking, feeble con- ductors, but in this matter it must be understood that the differences in this respect be- tween different tissues are not absolute, but a question of degree, and may to a certain extent be modified under dif- ferent circumstances. Thus a feebly conducting tissue, under a favourable condition of tem- perature and strong intensity of stimulation, will become to a certain extent conducting. Highly conducting tissues like nerve, on the other hand, under unfavourable circumstances may, as I shall show later, be converted into very feeble conductors. Returning now to the question of the responsive reactions of skin, we see the theoretical possibility of the following typical reactions. Let the scale of excitabilities be repre- sented by diagram to the left of fig. 177. Now, if the trans- formation of the outer epidermal surface, A, be maximum, the sign of its reaction will exhibit the greatest extent of deviation FtG. 177. Diagram Representing Different Levels of Excitability, Plus, Zero, and Minus Diagram to right of figure shows how resultant up response (inner to outer) may be obtained when induced change at A is plus, and at B minus, or when induced change at A is less negative than at B. \i I RESPONSE OF ANIMAL AND VEGETAL SKINS 297 from the normal negativity. That is to say, its response will become absolutely positive, as represented by a above the zero line. The response of the inner surface, B, may be normal and strongly negative, as represented by e below the zero line. When both these surfaces, then, are simultaneously excited, the excitatory positive variation, or ' outgoing ' current at A, will conspire with the 'ingoing' current at B and the resultant electro-motive difference will be B,, 4- A,,, the direction of the responsive current being thus from inner to outer. But the same resultant up-response will also be induced, even if the reaction of both surfaces be negative, provided only that that of the outer, A^, be less negative than that of the inner, B„ as explained by the diagram to the right of fig. 177. The responsive current will then be represented as Bg — A„, that is to say, as proceeding from the more negative B to the less negative A. We have thus examined the two extreme cases possible under the following formula, in which the arrows show the direction of the responsive current, from the more to the less induced negative : ^^K e-^d^c-^b-^a. ^^P" I shall next proceed to demonstrate the existence of these two extreme types, taking vegetable skins as the experimental specimen. It was supposed by Dr. Waller, as will be recalled here, that owing to characteristic differences between animal and vegetable protoplasm the response of vegetable skin was opposite to that of animal skin : that is to say, the former was ' ingoing ' and the latter ' outgoing.' That this generalisation is not, however, justified, will be seen from the experiments which I am about to describe, carried out on the skin of grape. These results, it should here be pointed out, are not dependent upon any one method of inquiry, for each problem was subjected to attack and analysis by four different modes of experiment. The first of these (i) was by the Rotary Method of Mechanical Stimulation. This method has the great advantage that by it the absolute I 298 COMPARATIVE ELECTRO-PHYSIOLOGY response of each surface is displayed separately, without either being affected by the other. There is here, moreover, no complication due to the polarisation factor, inevitable when uni-directioned induction shocks are employed for excitation. Thus, after the individual responses of each surface have been analysed, we are able to arrive at a definite conclusion as to what would be the character of the resultant response if both the surfaces were simultaneously excited. This conclusion is then submitted to three other tests. Thus, (2) the two surfaces of the specimen are subjected simul- taneously to the same thermal shocks, according to the method already described. Again, (3) the Method of the After Effect under equi-alternating shocks is employed. And, finally, (4) the Direct Effect of these equi-alternating shocks of moderate intensity is recorded. The results obtained by all these diverse methods are in complete concordance with each other, and fully support the theo- retical inferences which have already been made. I took the skin of a ripe muscatel grape, such as are available in Calcutta. On making the galvanometric con- nections with the outer and inner surfaces, a resting current, C, was found to flow in the skin from the outer to the inner, just as in the skin of the frog (right-hand diagram, "^g- ^78). The grape skin was now mounted in the rotary stimulating apparatus, first, for the stimulation of the outer or epidermal surface, with the outer layer placed upwards. The distance between the two electrodes was always the same — namely, 2 cm. On now stimulating one of the two contacts, response took place by the induced galvanometric positivity of that point. That is to say, the current was •outgoing,' into the galvanometer circuit, from the surface of the skin. When the second point was now stimulated the deflection previously obtained was reversed, the second contact thus also exhibiting galvanometric positivity on excitation. The position of the skin in the apparatus was now changed, the inner surface being placed upwards. In this way points diametrically opposite to those in the last case RESPONSE OF ANIMAL AND VEGETAL SKINS 299 were subjected to excitation. It was now found that the responses of the inner surface were normal — that is to say, of galvanometric negativity. I give here (fig. 178, a) records of two successive sets of responses obtained from the external and internal points A and B. These records clearly demon- strate that the resultant up-response, on simultaneous exci- tation of the outer and inner surfaces, is brought about by the induced galvanometric positivity of the outer added ■Hto the induced negativity of the inner surface. As the resist- ance of the circuit in the two successive experiments was maintained approxi- mately the same, the amplitude of these re- sponses gives a fairly accurate idea of the Ifcrelative electrical effects induced on the two sur- faces. The positive or * outgoing ' effect of the outer surface is here slightly greater than the ' ingoing ' effect. The jBdiagram in fig. 178, b, shows how the individual effects conspire to give rise to a responsive current from the inner to the outer. In order to test the reality of the correspondence between his response of the grape skin and that of the frog, I now repeated these experiments, employing the same apparatus for mechanical stimulation, on the skin of the frog. From Khe records given in fig. 179 it will be seen that the isolated espouse of the outer surface is positive, or ' outgoing,' that ►f the inner being negative, or ' ingoing.' The amplitude >f the former was, however, much greater than that of the latter. These responses disappeared altogether when the tissue had been killed by immersion in boiling water. From isolated responses obtained by means of induction shocks, Fig. 178. Electrical Response of Grape- skin to Rotary Mechanical Stimulation {a) A, positive response of outer surface ; B, negative response of inner surface ; (b) c, current of rest, from outer to inner ; R, excitatory response from inner to outer, consisting of summated results of positive response of outer with negative response of inner. I 300 COMPARATIVE ELECTRO-PHYStOLOGV skin as alone active, the inner being, to his thinking, in- effective. This particular result may possibly be accounted for by supposing that he used a stimulus intensity which was not sufficiently strong. In my own experiments I obtained clear demonstration of the effectiveness of both surfaces in opposite ways electrically, though the effect obtained from the outer was undoubtedly the more intense of the two. By comparing these two experiments, then, on the grape skin and skin of frog, it will be seen that the inference that the vegetable protoplasm reacts in any way Jji I Fig 179. Electrical Response of Frog's Skin to Rotary Mechanical Stimulation (a) A, positive response of outer ; B, negative response of inner ; {a') A and B exhibit abolition of response in skin on boiling ; {d) c, current of rest from outer to inner ; R, excitatory response from inner to outer, being summated effect of positive response of outer and negative response of inner. essentially different from that of the animal is quite unjus- tified. How widely applicable is the method of mechanical excitation by rotary stimulus will be seen in an attempt, successfully carried out, to determine the very difficult ques- tion of the characteristic response of the intact human skin. This will be seen in the following record of the results obtained with the skin of a forefinger. The responsive elec- trical changes represented by the down records, exhibit induced galvanometric positivity of the excited surface (fig. 180;. I shall next describe the results obtained by simultaneous excitation of the inner and outer surfaces of grape-skin. The responses now given, under stimulation by thermal i RESPONSE OF ANIMAL AND VEGETAL SKINS 301 shocks, are seen in fig. 181, the resultant current being seen to be ' up '—that is to say, from the inner to the outer. On observing the excitatory after-effect of equi-alternating shocks, the results were found to be the same, the responsive current being now once more from the inner to the outer. I next took a series of records of the direct effect of equi-alternating shocks, the results of which were precisely the same as before. On applying stimulation, by exactly equal and alternating shocks, we, as already explained, flG. 180. Photographic Record of Electrical Responses of Upper Surface of Intact Human Forefinger to Rotary Mechanical Stimulation. ■ )own responses here indicate induced galvanometric posi- tivily. Fig. 181. - Photographic Record of Electrical Responses of Grape-skin to Thermal Shocks at Intervals of a Minute Responsive current from inner to outer. btain a result which is due solely to the differential excita- tion of the two opposite surfaces. This is not complicated in any way by the factor of polarisation, although the latter could not have failed to be present if the exciting shocks I had been one-directioned. Under the conditions of these equi-alternating shocks, then, a certain effect is often seen, in the phasic variation of the base-line, which is ex- l^remely characteristic. We have already seen (p. 98) that Isvhen a tissue is subjected to repeated or continuous stimula- tion, its condition undergoes a phasic or periodic variation. Thus from a neutral or positive condition, it may pass into 302 COMPARATIVE ELECTRO-PHYSIOLOGY be subsequently reversed to positive once more. Such phasic changes, moreover, may be repeated. They find visible indications in appropriate shiftings of the base line of the record. Similar effects are also shown in the differential response as seen in the records given in fig. 182 ; this feature is very noticeable. We here ob- tain the resultant response of the two surfaces of grape-skin, from the inner to the oiiter. As the result of the series of stimuli applied, the existing current of Fig. 182. Photographic Re- cord of Electrical Responses of Grape-skin to Stimula- tion by Equi-alternat.ing Electrical Shocks at In- tervals of a Minute Responsive current from inner to outer. Note periodic variation of resting-current, causing shifting of base-line, down and up. mi^"^ Fig. 183. Photographic Record of Series of Electrical Responses of Frog's Skin to Equi-alternating Electrical Shocks applied at Intervals of One Minute Direction of responsive current from inner to outer. Note also variation of base-line. rest undergoes a periodic variation. If this had remained constant, the base-line would have been horizontal. In the present case, the original current of rest was from outside to inside. This, at first, underwent an increase ; then a decrease ; to be followed, later, by another increase. Thus, in the course of about ten minutes, it exhibited an alterna- tion of almost one whole cycle. In the next figure (fig. 1 83) I give a series of results obtained RESPONSE OF ANIMAL AND VEGETAL SKINS 303 with frog's skin in direct response to equi-alternating shocks. Here we find the usual ' up ' responses, showing that, as before, the direction of the responsive current is from within to without ; and here also we see the existing current of rest undergoing a periodic change. It has now been fully demonstrated that the response of skin is determined by the differential excitabilities of its two surfaces, upper and lower, that of the lower being the greater. That the resultant responsive current from lower to upper, is in such cases brought about by the greater excit- ability of the lower, has been fully shown, in a previous chapter, by experiments on e pulvinus of Mimosa, next made records of a long series of responses given by the last-named specimen, with the object of finding out whether or 1^ Fig. 184. Photographic Record of Trans- verse Response of Pulvinus of Mimosa to Equi-alternating Electrical Shocks The direction of the responsive current is from the more excitable lower to the less excitable upper. Note the cyclic variation of the current of rest. ot these also exhibited a periodic variation of the resting-current similar to those just observed in the I ^Anisotropic skins of grape and of frog. Electrical connections were made with diametrically opposite points on the upper and lower surfaces of this organ, and they were subjected to equi-alternating shocks. IBpwing to the conducting power of the tissues, it was not now the upper and lower skin surfaces merely, but ^he upper and lower halves of the organ that became [■ixcited. And the responsive current was from the lower to the upper, as already demonstrated. In the particular record seen in fig. 184, the general resemblance to the responses of skin is sufficiently obvious. The interesting feature of this record is the periodic changes in the resting- i 304 COMPARATIVE ELKCTRO-PIIYSIOLOGY I cu rrent, which exhibit a com plete cycle in the course of Jhirteen minutes. Thus, as a consequence of the after-effect of stimulus, a cyclic variation of relative conditions is induced, as between any two anisotropic surfaces, such as those of skin or pulvinus. This cyclic variation of relative conditions is indicated by the concomitant variations induced in the resting-current, shown in the shiftings of the base line. I have been able, further, to demonstrate the interesting fact that such phasic variations are capable of exhibition even through mechanical response. I have already ex- plained that autonomous pulsations, such as those of the lateral leaflets of Desniodium gyrans, may be regarded as the after-effect of stimuli previously absorbed and held latent by the tissue. In taking the record of a series of such pulsations, I have often found phasic variations to occur, similar to those obtained with long-continued response of skin or pulvinus. If, for example, the lower half of the pulvinus of the lateral leaflet of Desmodiitin undergoes an increase of turgidity above the average, that half will become more convex, and the base-line of the record will be correspondingly tilted. The converse will take place under the opposite change. Thus the phasic variations shown in the record (fig. 185) clearly indicate that the relative turgidities of the two surfaces of an anisotropic organ may undergo a periodic change. The corresponding electrical expression of this we have seen in the variation of the current of rest This variation may sometimes be so great as actually to reverse the normal current of rest. Thus, while under normal standard con- ditions the resting-current in the pulvinus of Mimosa is from the upper half to the lower, across the organ, this normal direction may sometimes be found to be reversed. It may now be asked. What is it, in the case of the skin, which determines the respective directions of the resting- current and the current of response ? We have seen that the current of rest in the frog's skin, from outer to inner, is generally attributed to the possession of glands by the outer, RESPONSE OF ANIMAL AND VEGETAL SKINS 305 a supposition seemingly supported by Rosenthal's discovery, already referred to, that an apparently similar 'ingoing' current was to be observed in the mucous coat of the frog's stomach. Against this may be urged the conclusion, to which Hermann drew attention, that the skin glands are nor- mally nearly closed to the external surface, and cannot there- fore have any external galvanic relation. There are, moreover, other arguments. First, a similar current is observed in the case of grape-skin, where there is no special glandular layer. Second, the specific response of a glandular surface is 5 P.M. 9 P.M. lWA**N«Jf«%r'**~*^ HA^V>" 12 IfFir,. , Th( efini 3 A. 6 A.M*. [85. Continuous Photographic Record of Autonomous Pulsation of Desmodium gyrans from 6 p.m. to 6 a.m. The lower record is in continuation of the upper. Note phasic variation. efinite, and is by galvanometric negativity, whereas the response of the outer surface of frog or grape skin on excitation is by galvanometric positivity. And, thirdly, we shall see that the current observed in the mucous membrane of stomach is most probably not the natural current of rest, but the- excitatory after-current. It will be remembered, however, that we have always found the natural current to flow in the tissue from the less to the more excitable, and the current of response in the opposite direction. In the skin, owing to physiological and I X 3o6 COMPARATIVE ELECTRO-PHYSIOLOGY histological modifications, the outer surface is reduced in excitability. The epidermal layers have little protoplasmic contents, and may be transformed in various ways, becoming corneated or cuticularised. The extent of such transforma- tion may be small or great, but the external layer will as a general rule become less excitable than the inner tissue. Hence, under normal conditions, we have a current of rest from without to within. If the inner layer be only moderately excitable, or if its power of recovery from excitation be great, then the dis- turbance caused by the prepara- tion of the specimen will be slight, or will pass off quickly. It is to be remembered that as the inner surface is the more excitable, the responsive current due to the mechanical stimulus of preparation will be from inner to outer ; and therefore its after-effect, proving in certain cases persistent, may give rise to a current apparently the re- verse of the true normal current. Thus, the direction of the current of rest, which we should have inferred theoretically to be from the less to the more excitable, may occasionally be found re- versed, owing to the excitatory after-effect of preparation. The current of rest, moreover, is liable to autonomous periodic variation, as we have seen. The most satisfactory method of determining the relative excitabilities of two surfaces, then, lies in subjecting them to simultaneous excitation, and observing the direction of the responsive current. In the skin, unless the tissue was fatigued, I have always found this to be from the more Fig. 186. Photographic Record of Electrical Responses in Skin of Neck of Tortoise to Stimulus of Equi-alternating Electrical Shocks at Intervals of One Minute The direction of the responsive current was from inner to outer. The so-called current of rest was also in this case, owing to the excitatory after- effect of preparation, from inner to outer. RESPONSE OF ANIMAL AND VEGETAL SKINS 307 li, c A . B — excitable inner to the less excitable outer, even in those cases where the normal direction of the resting-current had been reversed, as an excitatory after-effect of preparation. This fact is well illustrated in the following record, taken with the skin of the neck of tortoise. As an after-effect of preparation, the resting-current so called was here reversed, flowing from inner to outer. But the excitatory responsive current was nevertheless from the more excitable inner to the less excitable outer (fig. 186). It was stated at the beginning of this chapter that the esultant response from inner to outer merely expresses the general fact that the excitability of the inner is greater than that of the outer. And this will still remain true, even when the transformation of the external layer is not so great as actually to reverse its individual galvanometric response from negativity to posi- tivity. The experi- mental results obtained with the skin of ripe tomato form a case in point. The natural current of rest is here, as usual, from the outer to the inner, and the excitatory responsive current in the opposite direction. But from the analysis of individual responses, on the outer and inner surfaces, obtained by means of the rotary apparatus for mechanical stimulation, it will be seen (fig. 187) that both the surfaces alike give the normal excitatory response of galvanometric negativity. This responsive negativity of the inner, B, is, however, very much greater than that of the outer. The resultant tsponse, then, representing as this does the difference in Fig. 187. Isolated Responses of Upper and Lower Surfaces of Skin of Tonriato to Rotary Mechanical Stimulus (a) A, negative response of feeble intensity in outer surface ; B, negative response of much greater intensity in inner surface ; {b) c, cur- rent of rest from outer to inner. Resultant excitatory response from inner to outer, due to greater induced galvanometric negativity of inner. 3o8 COMPARATIVE ELECTRO-PHYSIOLOGY degree between two negativities, is still from inner to outer, owing to the greater excitatory reaction of the inner. That the direction of the resultant response is actually from the inner to the outer is seen in the series of records given in fig. i88. The stimulus consisted of equi-alter- nating electrical shocks, applied at intervals of one minute. The record shows negative responses, followed appa- rently by the positive after-effect. In order to observe the peculiarities of this response in greater detail, the record was taken on a faster-moving drum (fig. 189). From this figure it will be seen that there was a short latent period of no responsive reaction. Response then rose to a maximum, and again sub- sided. After now reaching the zero position, the record proceeded in the positive direction, and again reverted back to zero. In similar records, the occurrence of this latent period, and posi- tive after-variation, has been adduced by certain physio- logists as affording visible demonstration of the exist- ence of opposite processes of assimilation and dissimilation. It has been supposed that the various features of the response were the outcome of a sort of tug-of-war between the two opposed forces, the preliminary pause being the expression of a short-lived balance, while the subsequent negative and positive variations were to be regarded as indicating the predominance, now of the one process, and then of the other. That in the present case such an assumption is unwarranted will be evident when we observe the isolated responses of the upper and lower surfaces separately (fig. 187). In each of ■wm- Fig. 188. Photographic Record of Series of Responses in Skin of Tomato under Equi-alternating Electrical Shocks applied at In- tervals of One Minute Direction of resultant current from inner to outer followed by feeble opposite after-effect. RESPONSE OF ANIMAL AND VEGETAL SKINS 309 these we see the normal response of galvanometric negativity, followed by recovery, without evidence of any antagonistic process, such as might give rise to subsequent positivity. The difference between these two responses lies simply in their time-relations. On simultaneous excitation of the two, the predominant negativity of the inner gives the first half of the negative response. The persistence of the excitatory reaction of the outer, on the other hand, after the subsidence of the effect on the inner, gives rise to the apparently Fig. 189. A Single Response of Skin of Tomato to Equi-alternating Shock recorded on Faster Moving Drum positive after-effect. Thus, here the supposed tug-of-war between two opposite processes of assimilation and dis- similation is, in reality, between two normal responses having different time-relations. It is from a failure to recognise the fact that the excitatory reaction is not con- fined to one, but takes place on both surfaces, that such erroneous assumptions as that referred to have often been occasioned. I 3IO COMPARATIVE ELECTRO-PHYSIOLOGY The result which I have described — namely, a greater responsive negativity of the inner than of the outer, giving rise to a resultant responsive current from inner to outer — is that which occurs in the majority of cases with tomato. But, as establishing a continuity between this response and that of grape-skin, I may mention the interesting fact that in a few instances I obtained records in which, while the inner surface on excitation exhibited a strong negativity, the outer, under the same stimulus, exhibited a feeble positivity. The normal response of skin is sometimes, however, found to be reversed, and no explanations have yet been lii^vUl^" Fig. 190. Photographic Record of Series of Normal Responses in Skin of Gecko Responsive current from inner to outer surface. offered to account for this. But 1 have shown two definite conditions of universal application, which are liable to bring about the reversal of normal response. These are, on the one hand, sub-tonicity, and, on the other, fatigue. Should the condition, in a given case, be the former of these, then the impact of stimulus will of itself, by raising the tonic condition, restore the normal response. Thus in a case of abnormal positive response due to sub-tonicity, an inter- vening period of tetanisation will tend to convert the ab- normal response to normal. An abnormal positive will thus pass into diphasic, and thence into the normal negative. RESPONSE OF ANIMAL AND VEGETAL SKINS 311 P^or the following experiments, I took the skin of gecko, which can be detached from the body with very little injury. This animal offers remarkable facilities for many electro- physiological experiments. Its isolated tissues can be main- tained in a living condition for a very long time. Its sciatic nerve affords us a specimen about 15 cm. in length. Thus for electro-physiological investigation, it provides much greater advantages than the frog. Fig. 191 Photographic Record of Abnormal Diphasic Responses in Skin of Gecko, converted to Normal, after Tetanisation Taking a specimen of gecko skin, which was in a favour- able tonic condition, I obtained the series of normal responses to equi-alternating shocks, which is given in fig. 190. The responsive current here flowed from the inner to the outer surface. I next took another specimen, which was in a less favourable tonic condition, and obtained records of its responses, here seen (fig. 191) to be diphasic. An intervening period of tetanisation is seen, however, to restore the normal -esponse. CHAPTER XXIII RESPONSE OF EPITHELIUM AND GLANDS. Epidermal, epithelial, and secreting membranes in plant tissues— Natural resting-current from epidermal to epithelial or secretory surfaces — Current of response from epithelial or secretory to epidermal surfaces — Response of Z)///^;//a— Response of water-melon — Response of foot of snail — The so- called current of rest from glandular surface really due to injury- Misinterpretation arising from response by so-called ' positive variation ' — Natural current in intact foot of snail, and its variation on section — Response of intact human armpit — Response of intact human lip — Lingual response in man — Reversal of normal response under sub-minimal or super- maximal stimulation — Differential excitations of two surfaces under different intensities of stimulus, with consequent changes in direction of responsive currents, diagrammatically represented in characteristic curves — Records ex- hibiting responsive reversals. Having now seen how the responsive pecuh'arities of the epidermis may be elucidated by the responses of similar tissues in the plant, we shall next take up an inquiry as to the parallelism between the responses of epithelium and glands in animal and in vegetable tissues. And here, as in the last case, we shall find the obscurities of the one made clear by the study of the other. If we take the hollow peduncle of a Uriclis lily, and, cutting this into longitudinal halves, take a portion from the upper end of one, we shall observe noticeable differences between the investing membranes of the outer and inner surfaces. On the outer, as we have seen elsewhere, the cells are dry, thick-walled, and cuticularised. This surface then is naturally distinguished as epidermal. The internal mem- trane of the hollow tube, however, is very thin, and its cells J(V\ very little differentiated (fig. 192). The internal membrane ' \ may thus be distinguished as epithelial. \j0^^^ If now we examine this inner membrane continuously \ RESPONSE OF EPITHELIUM AND GLANDS 313 from the top to the bottom, at the point where the peduncle rises from the bulb, we shall find that the epithelial layer of the upper end passes imperceptibly into a markedly secret- ng (glandular?) layer at the lower. By this, secretion is constantly taking place, filling up the hollow tube with fluid. In one instance which I measured, the amount of this secretion was as much as 10 grammes in the course of the day. These secreting cells in this, which may be called the glanduloid layer, are very thin- walled and excessively turgid, and, from an evolutionary point of view, these gradual transitions from epidermal to epithelial, and from epithelial to secretory layers, observed under conditions of such great simplicity, are extremely interesting. When we come to test the electrical reactions of these tissues, IHbe find, on making electrical con- •"ections with the external epi- dermis and the internal epithelium, that a natural current flows in the tissue from the external surface to the internal. This would indi- cate that the internal was the more excitable of the two. This conclusion is confirmed on the application of simultaneous excitation to outer and inner ; for the direction of the responsive current is found to be from the internal surface to the external. If, next, electrical connections be made with the epidermal ' and secretory layers, a current of rest is once more observed from the external to the internal. On excitation, a very strong electrical response is given, its direction being from the highly excitable secreting layer to the less excitable Kidermal. From these experiments we see that the Fig. 192. Transverse Section of tissue of Hollow Peduncle of Uriclis Lily Cells of epidermis are small and thick-walled, those of inner surface large and thin-walled. 314 COMPARATIVE ?:lectro-physiology epidermal cells are, generally speaking, the least, and the secreting cells the most, excitable. I shall show, moreover, that all the responsive character- istics of these secretory cells are to be found repeated in those admittedly glandular layers which occur in the lining of the pitcher of Nepenthe, and cover the upper surface of the leaf of Drosera. Before, however, entering upon the consideration of these highly differentiated organs of Nepenthe and Drosera, which are further characterised by some of the digestive functions, I shall first discuss in detail the reactions of a simpler type of vegetable organ. This is exemplified by a single unripe carpel of Dillenia indica, already referred to. When this is carefully removed from the inside of the pseudocarp, and opened, the inside is found to be filled with a gelatinous secretion. This is gently removed, and electrical connections are made with the inner and outer surfaces. It must be borne in mind that these vegetable organs, being not highly excitable, admit of experimental preparations being made with little or no excitatory effect of injury. Allowing now for a period of rest after making the preparation, it will always be found that the current of rest is from the outer layer to the secreting i nner layer, which latter is thus,^ relatively speaking, galvanometrically positive. From the fact which we have generally observed, that the natural current of rest is from the less to the more excitable, it would appear, then, that the inner layer is here the more excitable. This conclusion, moreover, is in agreement with the inference already arrived at, in connection with the tissues of the Uriclis lily, that secreting cells as a rule are relatively the most excitable. This inference may, however, be subjected to the test of direct experiment. I first tested the response of the same specimen by means of thermal shocks, applied to both surfaces simultaneously. The definite direction of the responsive current, from the inner to the outer, across the tissue, proved conclusively that the inner surface was the more excitable, becoming as it did, galvanometrically negative in relation to the outer. A similar RESPONSE OF EPITHELIUM AND GLANDS 315 effect was obtained as an after-effect of equi-alternating shocks. I next took records of the direct effect of equi-alter- nating shocks. The direction of the responsive current was found, as before, from the inner to the outer. In the fruit of water-melon we obtain another specimen whose interior cavity is filled with secretion. On making a suitable preparation of this specimen, and arranging electrical connections with the outer epidermal and inner secretory- surfaces, I found the respon'sive current, under equi-alter- nating electrical shocks, to be from the inner secretory to the outer epidermal. Fig. 193 gives a photographic record of these responses. IBt From the typical responsive effects thus obtained with vegetable specimens under the simplest conditions, we are enabled to see that the effect of localised stimulus depends on the characteristic response of the surface layers of the organ. When dealing with this question of the electrical reaction of epi- thelium and glands in animal issues, Biedermann rightly Fig. 193. Photographic Record ot . ,t_ 1 . i.v . -i. Responses of Water-melon to ame to the conclusion that it Equi-alternating Electric Shocks f as the surface epithelial layer Responsive current from internal hich was, in an electro-motive ^^or^^^^g to external epidermal ' ^ surface. sense, most effective, the term, IB|n its widest sense, including the epithelium of glands and papillae. One complicating factor present in the electrical reactions f animal epithelia and glands, but relatively absent under the simpler conditions of the plant, is the effect of injury caused by the process of isolation. The very fact of making the neces- sary section involves a stimulus of great intensity, and unless the effect of this has thoroughly subsided, the after-effect of such stimulation may be so strong as to reverse the normal current of rest, and otherwise modify the excitability of the I— — 3l6 COMPARATIVE ELECTRO-PHYSIOLOGY out by Dr. Waller,^ on the isolated paw of a cat. The current of rest was found by him to flow from the surface of the pad to the section. From this he was led to the conclusion that this current could not have been due to injury, since in that case it would have flowed from the sectioned to the uninjured surface, and not in the opposite direction, as was found to be the case. This misconception arises from a failure to realise that the so-called * current of injury ' is, in fact, an after-effect of excitation. In the case under consideration, the section, acting as an intense stimulus, simply induces greater excita- tory reaction of the more excitable, which in this case happens, as we should have expected, to be the glandular surface. The injury-current here, then, is necessarily from the more excited glandular to the less excited non-glandular. A precisely similar result was obtained in the case of anisotropic plant-organs, where excitation caused by injury of the less excitable side, becoming internally diffused, induced greater galvanometric negativity of the more ex- citable distal point (p. 162). For a typical experiment on a glandular preparation, showing the principal effects, and the complications that may arise owing to injury, we may take the detached foot of the pond-snail, the lower surface of which, as is well known, secretes a slimy fluid. On allowing for the necessary period of rest, and then making electrical connections, we observe a current of rest, so-called, which flows from the glandular to the sectioned end. This is not to be mistaken for the true natural current of rest, being in fact the after-effect of a greater galvanometric negativity at the more excited glandular surface, consequent on section. An independent experiment, in support of this induction, will be described presently. On now simultaneously ex- citing the two surfaces, by equi-alternating shocks, the responsive current is found to flow from the gland inwards, the more excitable gland becoming thus galvanometrically negative. The responsive current is in this case in the same ' Waller, Signs of Life ^ p. loi. RESPONSE OF EPITHELIUM AND GLANDS 317 direction as the so-called current of rest, constituting a positive variation of it. From these experiments it is clear that the responsive current is due to the greater intensity of the induced gal- vanometric negativity at the more excitable glandular surface. It must, however, be noted here that this definite understanding of the phenomenon has been arrived at by fixing our attention on the relative excitatory reactions at the two contacts. If, instead of this, we had regarded it from the usual point of view, of variations of the resting-current only, we must have interpreted it as apparently an abnormal positive variation ; ^ for the so-called resting-current, in such a case, on account of the excitatory after-effect of injury, must also, as we have seen, flow from the more excited gland to the less excited muscle. Great confusion, and resultant misinterpretation of observations, have arisen from not sufficiently recognising these facts, that the resting-current may be originated in either of two distinct ways, and that the excitatory effect may consequently be summated with it in different manners. The resting-current in the primary condition is, as I have demonstrated elsewhere, the natural . L current. Th is originates in the natural differences of ex - ^ ^\a citability betw een different points, and is, in the intact ^ I specimen, through the tissue from the less to the more ex- i/^r^ citable. External stimulus now gives rise to a responsive current, which is in the opposite direction, and therefore constitutes a negative variation of this. This takes place because the more excitable point, which was naturally positive, has now become negative. But we may have a current of rest which is due to previous excitation, or injury, such as may be caused by the shock of the pre- tjaration. This current, though usually regarded as the I ^pesting- current, is not the true natural current of rest. It is really, as it were, the responsive current become persis- t;nt. Succeeding stimuli, inducing responsive galvanometric ' We shall find in Chapter XXVII. that similar misinterpretations have arisen : 3l8 COMPARATIVE ELECTRO-PHVSIOLOGY negativity of the more excitable, will now give rise to a current in the same direction as this resting-current, thus constituting a positive variation of it. It is only when fatigue has set in at the more excitable, and induced great depression of excitability there, that the response-current may undergo a reversal, its direction now being from what was originally the less excitable, to the originally more excitable (p. 177). An example of this I found in the sectioned foot of the large Indian garden-snail. Here the excitatory action on the glandular surface, due to the shock of preparation, was ex- tremely great, as evidenced by the profuse secretion which occurred immediately. Owing to this over-stimulation, fatigue was induced, with consequent great depression of ex- citability. Hence the responsive current was now found to be reversed, having, with reference to the glandular surface, become outgoing instead of ingoing. It has been stated above that the ingoing current of rest, observed at the glandular surface under preparation, was not the true natural current, but due to the excitatory after-effect of injury. This I was able to verify by observing the current of rest under natural conditions, without excitation. The snail was allowed to crawl on a glass surface, in the middle of which was a strip of linen moistened in normal saline. This brought the glandular surface into electrical connection with one of two non-polarisable electrodes. When the snail had of its own accord come to a temporary standstill on this piece of linen, the other electrode was quietly placed against the skin of the upper side of the protruding body. The natural current of rest was now found to be outgoing, as regards the glandular surface of the foot, the more excitable being thus galvanometrically positive. The absolute electro-motive difference was found to be -H •0013 volt The foot was now sectioned, and the difference of potential between the same points was found to have undergone a reversal. The supposed resting-current was now ingoing, through the glandular surface. Thus, owing to the excitation consequent on preparation, the more excitable surface, originally positive. RESPONSE OF EPITHELIUM AND GLANDS 319 had become negative. The induced variation from the original condition, in the present case, was from + -0013 volt to — '0020 volt. It will thus be seen that any irritation is liable to change the natural positivity of a highly excitable glandular surface to negativity. The supposed similarities between the ingoing responsive currents of frog's skin, and the glandular surface of the stomach, are therefore not real. That the two cases are quite different is proved indeed by the fact that local stimulation of the surface of the skin induces galvanometric positivity, whereas a similar stimulation If the glandular surface induces negativity. In experimenting on animal tissues, it is therefore ad- isable, wherever possible, to use intact specimens. The numerous experimental difficulties with which we are in that •case confronted, may be overcome by the method of simul- IBaneous and equi-alternating shocks which has been described. How practicable this method has been rendered will appear from the experi- ments which I have yet to describe on human subjects. ^^ We have seen that a protected sur- ^Lce is likely, other things being equal, ^Bd be more excitable than an exposed ^■ne. Partly owing to this fact, and ^^artly also to its richer possession of imbedded glands, it appeared to me |fc>robable that the inner surface of the armpit would prove electrically more fig, 194. Photographic excitable than a corresponding area, on. Record of Electrical 1^^ Responses of Intact IBay, the upper and outer surface of the Human Armpit same shoulder. In the records which I Responsive current from Iucceeded in obtaining (fig. 194), this ^^^^^ upposition was fully borne out. Equi-alternating shocks of »ne second's duration were applied at intervals of one minute, ind the direct effect photographically recorded. The re- sulting responses were found to be ingoing as regards the 320 COMPARATIVE ELECTRO-PHYSIOLOGY In order next to show that epithelial cells in the animal are relatively more excitable than epidermal, as we have already found to be the case in vegetable tissues, I performed the following experiment on the human lip. Here it was important that the electrical connections should be main- tained steady. A light spring- contact key was therefore made, as seen in the lower part of fig. 195. The lower contact of this spring-clip consisted of an amalgamated plate of zinc leading to the lower electrode. Over this were tied four thicknesses of blotting- paper soaked in zinc sulphate solution. On this again were placed three more thicknesses of blotting-paper, soaked in normal saline. The zinc plate which formed the upper limb of the clip, in connection with the second electrode, was similarly covered with separate layers of blotting-paper, soaked in zinc sulphate and normal saline re- spectively. The protruded lower lip was now placed in the clip, as shown in the upper figure, in such a way that the latter made a gentle but secure contact. A galvanometer and a source of equi-alternating currents were also placed in the circuit. Of the two electrodes, the upper was in connection with the epithelial, and the lower with the epidermal surfaces. The Jnatural current was now found to flow in the tissue, as in the /corresponding cases of plant specimens, from the epidermal { to the epithelial. The perfect steadiness of the contact was evidenced by the stillness of the deflected galvanometer spot of light. On now applying the alternating excitatory shock, the responsive current was found to be in the oppo- HuCL Fig. 195. Experiaoenlal Arrange- ment for Response of Human Lip. Lower figure gives an enlarged view of the spring-electrodes. RESPONSE OF EPITHELIUM AND GLANDS 32 1 site direction to the natural current, thus demonstrating the fact that the epitheHal layer was here, as in the plant, the more excitable of the two. The regularity of this effect will be seen from the series of photographic records given below (fig. 196), in which is exhibited a slight staircase effect. We next proceed to deal with the response of the glan- dular organ, the tongue. The tongue of the frog has formed the subject of a very extended series of researches, by Engelmann and Biedermann. On very careful isolation, entailing as little injury ILS possible, it was found by these workers hat the natural current was ' entering ' : hat is to say, it flowed across the tongue rom the upper surface to the lower. Both electrical and mechanical stimulation was I found by these observers to cause a negative variation of this natural current. I As isolation of such a highly excitable organ as the tongue may, however, give rise to unknown excitatory after-effects, it ap- jB|)eared to me very desirable that an investi- gation on this subject should be carried out '''^;,p^tRjoTo{ on the intact human tongue. In connection Electrical Response with this, I must point out that both the Lip ^^^ ""^^" surfaces of the tongue are excitable. Our Responsive current inquiry, therefore, is into the relative excit- ^^^^ epithelial to ^ •' ' ' epidermal surface. abilities of its upper and lower surfaces. Here Ilhe experimental difficulty lies in this very high excitability bf the organ, on account of which — except when in a quies- cent state and with a very steady contact — the galvanometer spot of light is apt to be erratic in its movements. Much (►f this difficulty is overcome, however, by holding the pro- ruded tongue lightly clamped between the teeth. The upper md lower surfaces may then easily be held in the clip-key ilready described. From this double support of the clip and he teeth it is, with" a little practice, possible to arrange natters in such a way that the galvanometer spot is 322 (Comparative electro-physiology practically stable. The current of rest in the intact human tongue is then found to be from the upper to the lower surface, as in the frog. This, according to our previous results, would indicate that the upper surface is the less excitable. This inference finds independent verification, when we subject the organ to the stimulus of equi-alternating shocks. A very strong responsive current is now found to flow through the tongue, from the lower to the upper surface. The tongue is so extremely sensitive that its characteristic response can be evoked even with very feeble stimulus. I have already explained that the alternating currents induced by speaking before a telephone are not exactly equal and opposite, the current being slightly stronger in one direction. Hence, if such currents be made to play upon an organ in which the excitability is only moderately differential, the preponderance of one of the two elements of the alternating shocks is then likely to mask the true excitatory effect. But the differential excitability of the tongue is so great that the responsive current is always from below to above, whether the exciting current be made to act in a favourable or unfavourable direction. Thus, if one speak, even in a very ordinary voice, into an exciting telephone, which is in series with the rest of the circuit, with its poles direct or reversed, a definite lingual current is induced in response. This, already said, is always in direction from the lower surface to the upper — surely a curious instance of the speech of one inducing lingual response in another, by direct, and not by provocative action ! The results which have been described are the normalj effects given in response to stimulus of moderate intensity, By moderate stimulus is here meant that intensity of current] which is obtained when the primary coil is slightly within] the secondary. By feeble, on the other hand, is meant thej intensity produced when the primary is at a distance froi the secondary. Excessively strong stimulus again occurs when the primary is pushed fully within the secondary. ll shall now proceed to describe occasional variations which, RESPONSE OF EPITHELIUM AND GLANDS 3:23 may be observed when the stimulus is either very feeble or excessively strong. We have seen (p. 83) that when the intensity of stimulus is below the critical degree which is sufficient to induce response, its effect is to increase the internal energy of the tissue. We have also seen that the sign of this increased internal energy is galvanometric positivity, being thus opposite to the excitatory effect. Hence, in a dif- ferentially excitable tissue, we may expect to find instances in which stimulus that falls below the threshold of true excitation will act by indiicing a greater galvanometric positivity of the more excitable, whereas, under normal intensity of stimulus, the more excitable would have become galvanometrically negative. We can thus see the possibility of response being reversed under very feeble stimulus. It must be remembered that the excitability of both the contacts is a factor in the response, which has hitherto been overlooked. A second very important factor, which has not yet been taken into consideration, is the difference between the characteristic curves of the tissues at the two different surfaces. By characteristic curve is here meant the curve which shows the relation between intensity of stimulus and response. This difference will be better understood from the diagram of the theoretical curves given below (fig. 197). This exhibits all the cases that can possibly exist. Let the curve K a a' a" represent the characteristic curve of the surface A. Let the curve J^ b b' b" similarly represent the characteristic curve of the surface B. Of these two surfaces, B is under moderate stimulation, normally the more excitable. In the middle portion of the curve, representing response under moderate intensity of stimulus, the induced galvanometric negativity of B is thus greater than that of A. Under moderate excitation, therefore, the current is b'-^a' through the tissue in the direction from B to A. But below the threshold of true excitation, B would be positive, and A 324 COMt>ARATtVE ELECTRO-PttYSlOLOGV relatively negative to it. Hence there would here be a reversal of response, the direction of the responsive current a-^b through the tissue being now from A to B. This current will be recorded by the galvanometer, provided the induced difference between A and B be sufficiently great. Having thus inferred the different effects possible under .sub-minimal and moderate stimuli, we shall next consider the differential effect which may sometimes be induced by excessively strong stimulus. In the middle part of the curve, •/ \ i i /I A ^ y / • \,./^ f H^ ■ riMULUS Fig. 197. Possible Variations of Responsive Current, as between Two Surfaces A and B, shown by Means of Diagrammatic Representations of Characteristic Curves A, a, a', a", characteristic curve of surface A ; B, b^ h\ b'\ that of B. Under moderate stimulation, B is the more excitable, its induced galvanometric negativity being greater, and the direction of current from b' to «', as in the middle part of the curve. Under subminimal and super-maximal stimulation the direction of the responsive current is reversed to a -> /5 and a" -> ^" respectively. B ^ 3' U' is seen to be very much steeper than A a a' a" \ that is to say, the excitatory effect increases very rapidly with the stimulus, in the more excitable of the two surfaces. But this increase may sooner or later reach a limit, that curve tending to become horizontal, aided in this process, possibly, by growing fatigue. The curve A a a' a'\ however, though not so steep, may yet continue to rise throughout a longer A abscissa, representing increasing intensity of stimulus. In " such a case, there would be a second crossing-point, and RESPONSE OF EPITHELIUM AND GLANDS 325 a second reversal of normal response into a"->b'\ under excessively strong stimulation. We are thus enabled to see the theoretical possibility of the reversal of normal response under the two conditions of sub-minimal and super-maximal stimulation. All these phases may not be displayed in the same specimen ; but it may be possible to find different specimens exhibiting one or the other. In some cases the difference a — b\s too small to allow of an appreciable galvanometric effect, and in their higher parts the curves do not cross. In such specimens, then, there is no response under sub-minimal stimulus, and only .^lormal response under increasing intensities, however strong. I^phe only exception to this will take place when fatigue supervenes, a case which will be dealt with presently. I find I^^at this type of response is the most common. ^P We have next to consider those cases in which in the sub-minimal region, the difference a—b is appreciable, the reversal to normal b'-^a' taking place under higher in- tensities of stimulus. There need not in such an instance, be any second reversal. Here, then, the normal response under moderate or strong stimulus is reversed when the stimulus is sub minimal. An example of this will be given presently. I^P Lastly, there may be a type of response in which in the sub-minimal region the difference a — b is slight, and the normal b'-^a' is reversed to a'^-^b" in the region of excessive stimulation. I shall be able to give an example of this also. «I have been able, by taking different specimens, to emonstrate the occurrence of these theoretical reversals of response, under sub-minimal and super-maximal stimulation. I have not yet been able to find a single specimen ex- hibiting both reversals, but it is not impossible that this exists. IB The type in which response remains normal throughout a wide range of stimulus-intensity is too numerous to require special illustration. But this normal response may be reversed under fatigue. Generally speaking, a highly II 326 COMPARATIVE ELECTRO-PHYSIOLOGY excitable tissue may be expected to show earlier or greater fatigue, than, other things being equal, a less excitable tissue.' Thus, under strong or long-continued stimulation, the excitability of the originally more excitable B may be depressed, so as to fall below that of A, with consequent reversal of response. This will be seen clearly in a typical experiment on the pulvinus of Mimosa. Electrical con- nections were here made with the upper and less excitable surface A and the more excitable lower surface B, records being then taken of normal responses to equi-alternating liu^liurJCi'i \ TftVi^i fiG. 198. Photographic Record showing Reversal of Normal Response in Pulvinus of Mimosa due to Fatigue (a) Series of normal responses, direction of current being from more excitable lower to less excitable upper ; {b) Reversed responses in same specimen, due to previous tetanisation, causing fatigue. electric shocks. The stimulus employed was of moderate intensity, the secondary being placed slightly overlapping the primary. The responses (fig. 198, a) are seen to be normal, the responsive current being from the lower to the upper. They also show signs of slight fatigue, their amplitude undergoing diminution. The secondary was then pushed over the primary, and the tissue subjected to the consequent intense .stimulus for two minutes continuously. • In this matter, the nature of the tissue must be taken into consideration, nerve, for example, being less subject to fatigue than muscle. RESPONSE OF EPITHELIUM AND GLANDS 327 The secondary was next brought back to its original position and a record once more taken of its successive responses to stimuli of the same intensity as before. It will be seen (fig. 197, b) that the response is completely reversed by the relative depression of the excitability of the lower surface B. Similar reversal under fatigue will be shown in the • glandular organ of Drosei-a in the next chapter (fig. 208). I have obtained other interesting variations of normal response induced by fatigue. Thus, taking the carpel of IK Dillenia indica, and making electrical contacts with its inner Inland outer surfaces, the responses under moderate stimulus IH-were found to be normal — that is to say, from inner to outer I^K — the secondary being here understood to be partially over- ly lapping the primary at a distance of six divisions of the (scale. The secondary was now pushed home, and the tissue Btsubjected for a short time to strong and continuous stimula- tion. Moderate fatigue was thus induced. When the I secondary was now brought back to the distance of six H divisions of the scale the response was found to be reversed. Thus, moderate fatigue had here been sufificient to bring about the reversal of the relative excitabilities of the two surfaces of the carpel of Dillenia indica^ when the testing stimulus was of original intensity. But when the intensity of stimulus was increased, by pushing in the secondary to a position marked three divisions on the scale, the response ecame once more normal. Thus, fatigue had in this case modified the excitability of the two surfaces in such a way that an intensity of stimulus, which was formerly effective to induce greater excitation of the originally more excitable, was now ineffective ; and its greater excitation, with the restoration of normal response, could now only be evoked under stronger I stimulus. We shall next describe the reversal ol normal response iinder sub-minimal stimulation. For this we shall once more select the carpel of Dillenia indica, A record of its normal response — the direction being from inner to outer — is 328 COMPARATIVE ELECTRO-PHYSIOLOGY this record the intensity of stimulus was reduced by pulh'ng out the secondary further away from the primary. The responses were now found reversed, as seen in the subsequent series. We have last to consider the reversal induced by intense stimulation. Such instances must not be confused with the effect of fatigue. The two can be distinguished by the fact that the fatigue-reversal takes place after a series of normal responses, whereas the true reversal, due to strong intensity of stimulus, which we are now discuss- ing, is exhibited at the very beginning. Such an effect I have observed in the response of the human Fig. 199. Photographic Record showing lip- The ^ direction of Reversal of Response in Carpel of the responsive current was normally, under moderate stimulus, from the epi- thelial to the epidermal surfaces. Under very strong stimulus, however, this normal direction was found to be reversed. But the employment of excessively strong stimulation introduces other complicating factors. The applied stimulus may be supposed to be localised only when it is of moderate intensity. With intense stimulus the subjacent tissues are liable to be involved in giving rise to excitatory response ; and it then becomes a difficult problem to discriminate how much of the observed effect is due to the superficial layer, and how much to others more deeply situated, DiUenia indica, under Sub-minimal Stimulation The first series show normal electrical responses under moderate stimulus, responsive current from internal to external surface ; the second series exhibit reversed response under sub- minimal stimulation, current from external to internal surface. CHAPTER XXIV RESPONSE OF DIGESTIVE ORGANS Consideration of the functional peculiarities of the digestive organs -Alternating phases of secretion and absorption — Relation between secretory and con- tractile responses. Illustrated by (a) preparation of Mimosa ; (d) glandular tentacle of Drosera - General occurrence of contractile response — True current of rest in digestive organs — Experiments on the pitcher of Nepenthe — Three definite types of response under different conditions — Negative and positive electrical responses, concomitant with secretion and absorption — Multiple responses due to strong stimulation — Response in glandular \e2S. oi Drosera — Normal negative response reversed to positive under continuous stimulation — Multiple response in Drosera — Response of frog's stomach to mechanical stimulation — Response of stomach of tortoise — Response of stomach of gecko — Multiple response of frog's stomach, showing three stages — negative, diphasic, and positive — Phasic variations. AVING now dealt with the responsive characteristics of the kin, epithelium, and glands, alike in plant and animal, the ext subject to be taken up is that of the response of the igestive mucosa. And here we have to determine, first, hether or not there is, broadly speaking, any continuity tween the responses of digestive organs and those which e have just been studying ; and, secondly, to what extent he functional specialisation of the tissue has acted in accentuating certain of its responsive peculiarities. Surveying the function of digestion as a whole, we see that it consists, briefly, of two different processes — those, namely, of a previous secretion, by which food is rendered soluble, and of a subsequent absorption, by which the dis- solved foodstuffs are absorbed. In the membrane of the simplest digestive organ, then, the epithelial lining, using that term in its most inclusive sense, must be endowed with Re two properties of secretion and absorption under different 330 COMPARATIVE ELECTRO-PHYSIOLOGY circumstances. The question then suggests itself, what are the circumstances which determine this outflow or inflow ? In the digestive processes, moreover, in plant and animal alike, each of the reactions referred to, whether of secretion or absorption, must be more or less long-continued. Thus these responsive actions, instead of being single and spas- modic, are likely to be multiple and long-sustained. The characteristic response to stimulus of a secreting organ is understood to be by secretion. Is this reaction essentially different from those fundamental processes which underlie the responses of contractile organs, or are we to regard con- traction and secretion as but different expressions of a single responsive phenomenon ? In order to test this question, of the connection between responsive secretion and contraction, it will be well here to draw attention to certain experiments of Sachs, on the response of Mimosa^ though our inferences will be somewhat different from those which their author intended. If we take a longitudinal slice of the lower half of the pulvinus of Mimosa and keep it in a moist chamber for some time till the tissue has recovered from the excitation due to section, and if we then subject it to fresh excitation, water will be found to ooze out, or undergo secretion from the excited tissue. We may explain this occurrence in either of two ways : first, that, in consequence of the molecular changes induced by stimulus, contraction and permeability-variations take place in the cells, the expulsion of water being an expression of the active process of contraction ; or, secondly, that the oozing-out of the water is a passive process, due to permeability- variation of the turgid cells alone, without con- traction. The two theories may be distinguished broadly as those of active contraction and passive secretion. In the thin section of the Mimosa pulvinus it is the exudation of water that is noticeable, and not any marked movement character- istic of contraction. But in the intact pulvinus, owing to its anisotropic structure, the greater contraction of the more RESPONSE OF DIGESTIVE ORGANS 331 excitable lower half is exhibited in a marked manner by downward mechanical movement, magnified as this is by the long petiolar index. The intact organ, moreover, is invested with an impervious skin, hence the excitatory exudation, or expulsion, of water, being internal, is not seen outwardly. Thus a single identical reaction may appear from different points of view, as either secretory or contractile. The occurrence of contraction is thus most easily demon- strable when it is accompanied by conspicuous movement. This, however, demands considerable physiological anise- tropy, the differential contraction then giving rise to a very marked lateral movement, as in Mimosa. In radial organs [of plants, on the other hand, owing to balanced contractions [of opposite sides, there is no marked responsive movement. Hence ordinary plant-organs have hitherto been regarded as non-contractile and insensitive. But I have shown that all [these radial organs exhibit longitudinal contraction, to be detected and recorded by means of suitable magnifying idevices. All motile responses are brought about, it must be [remembered, by transference or redistribution of fluids. Now, [in organs invested with impervious membranes the effect ot [fluid-transference is manifested by mechanical movement ; [whereas, in naked tissues, the fluid-transference is directly [visible as secretion. From the very important series of researches carried out ^by Darwin, on the excitatory reactions in the tentacles of \Drosera^ we know that the pedicel, carrying the gland on its summit, is somewhat flattened, and that it is this anisotropic lower part which is alone capable of movement. The gland- :ells on the head of the tentacle have been shown by Gardiner to be provided with delicate uncuticularised cell- walls, which are curiously pitted on their upper or free ^surfaces. The terminal organ, or head, which is radial, ^ould thus seem to be peculiarly fitted for the exudation of liquid on excitation. In the anisotropic motile portion of the pedicel, on the other hand, the responsive reaction mani- fests itself by bending. It would thus appear that the same 332 COMPARATIVE ELECTRO-PHYSIOLOGY excitatory reaction may exhibit itself in different parts, even of the same organ, by mechanical movement and secretion respectively, according to the facilities which one or the other portion offers. That the phenomenon of contraction is behind the ex- citatory expulsion of water in a vegetable organ, would appear highly probable from certain results obtained in the electrical response. The stimulation of an ordinary vegetable tissue gives rise to two distinct electrical effects at a distance. The first of these is the arrival of the hydro-positive effect of galvanometric positivity, with positive turgidity-variation. The second is the wave of true excitation, with its character- istic of negative turgidity-variation and concomitant gal- vanometric negativity. The first, consisting, as this does, of a hydrostatic blow delivered at a distance, can only, it appears to me, be ascribed to an active process of contrac- tion, causing the squeezing- out of water in the excited region. A passive escape of fluid, due to mere permeability-variation, could not, as I think, originate that impulsive hydrostatic shock which is transmitted to a distance. For such a result to take place an active expulsion would seem to be requisite. In view of these facts, is it necessary to hold the doctrine of discontinuity, or, when there is evidence in its favour, are we to believe in the continuity of these apparently different reactions ? The excitatory reactions of different classes of tissues have hitherto been regarded as different, chiefly because some were looked upon as motile, and others as non-motile ; muscle, for example, was held to be typical of the first, and nerve of the second, of these classes. In this case of the nerve, it has been believed that there was no visible manifestation of the excitatory change. I shall, however, be able to show that even this supposition is incorrect, since the excitatory reaction in the nerve is in fact attended by contraction. The electrical indication of gal- vanometric negativity which is concomitant with contraction in contractile tissues, is also obtained in the case of excited li: RESPONSE OF DIGESTIVE ORGANS 333 glands. The visible changes which occur under stimulation in these three types of tissues would thus appear to differ only in degree. We may now turn more especially to the question of the ■ electrical reactions of the digestive mucosa. As regards the natural current of rest, we have seen that Rosenthal and others found that this current was ingoing — that is to say, the mucous layer was negative, as compared to the muscular coat of the stomach. Biedermann had also noticed a strong current of rest between the glandular surface of Drosera and he stalk.- But it will be shown that the glandular coat of e stomach is more excitable than the muscular layer, ence we should have expected that the natural current of rest would have been from the less excitable to the more excitable, the mucous layer in a state of rest being thus relatively galvanometrically positive. The opposite direc- tion of the current which has been observed, would rather ppear to be ascribable to the excitatory after-effect of pre- paration. I have already described how the glandular foot f the snail, under conditions of perfect rest, is galvano- metrically positive. But the excitation caused by prepara- tion renders this highly excitable glandular surface negative That the fact of cutting open the stomach, to make the Ixperimental preparation, similarly, would cause intense xcitation with galvanometric negativity, was to have been xpected. I shall be able, indeed, to show, by means of experiments to be described presently, that the shock con- Kequent on this preparation is to give, not one, but a pro- Dnged series of multiple electrical responses. I have almost invariably found, in making electrical ontacts after section, with the inner and outer surfaces of irog's stomach, that the multiple responses caused by section, |)ersisted for more than an hour ; and until these had ubsided no fresh experiment could be undertaken, to >btain records of the response of the stomach to external timulus. I have also found that many of these frogs were ["■"'■■"■■"■'"""" 334 COMPARATIVE ELECTRO-PHYSIOLOGY mechanical irritation of which would contribute to the negativity of the mucous lining. Finding, then, that it would be impossible to obtain the natural current, in a stomach which had to be cut open, I next turned my attention to stomachs which are naturally open. These are seen in the upper concave surface of the leaf of DroserOy for instance, which is provided with glandular tentacles. I here made electrical connections with the upper and lower surfaces respectively. But the tentacles excited by the contact of the electrode bent and clasped it round, an excitation which was seen in the galvanometer as negativity of that surface. From this may be gauged the difficulties which attend the observation of the true natural current of rest in such an excitable organ as the stomach. The demonstration, however, of the galvanometric positivity of the snail's foot, and of the inner glandular surface of the carpel of Dillenia indica^ lead to a strong presumption in favour of the true resting-current in the stomach being from the non-mucous layers to the mucous. Having thus seen the difficulties imposed by the high motile excitability of the tentacles of Drosera^ I next turned my attention to other specimens. We have seen that there is a secretion of fluid at the lower end of the hollow interior of the peduncle of Uriclis lily, and that the secreting inner layer is here galvanometrically positive in a state of rest. As this tube, however, is closed, it cannot be regarded as subserving the absorption of food-material. But the same limitation does not apply to those modified foliar structures, the pitchers of Nepenthe ^ (fig. 200). These, as is well known, are open. They have a histological differentiation, moreover, of their lining membrane, actual glands being present (figs. 201, 202), which are admitted to be comparable to those of the animal digestive organ, though of a much simpler type. A fluid is secreted by these glands, and insects entrapped in the pitcher are in it dissolved or decomposed. The products • I have to thank the authorities of the Botanical Garden, Sibpur, for supplying me with these valuable specimens. RESPONSE OF DIGESTIVE ORGANS 335 are subsequently absorbed by the tissue, as in corresponding cases, by the stomach of animals. From the study of the responsive peculiarities of so primitive a type of stomach, we might then expect to gain much light on the action of more complex and highly specialised digestive organs. In order first to obtain the true current of rest, 1 took a young pitcher which had previously been kept free from all disturbance. I next made electrical connections, by means of non-polaris- able electrodes, with the inner (glan- dular) and outer surfaces of this pitcher respectively. As some excita- tory reaction may be induced in a highly excitable organ, even by the contact of normal saline, the cotton threads in connection with these non- polarisable electrodes were moistened with the natural secretion of the Ditcher itself On carrying out the (Experiment under these ideal condi- tions, I found, as I had expected, :hat the current of rest flowed from •;he outer non-glandular to the inner glandular surface, the Jatter thus being galvanometrically positive. In investigating next the excitatory reaction, I obtained three different types of responses — negative, diphasic, and positive — characteristic of certain definite conditions. Before entering upon the details of these experiments, it is advisable to discuss here the probable significance of the negative and jjositive electrical reactions observed. We have seen that a slice of tissue from the pulvinus of Mimosa excretes water under excitation. The electrical reaction under these circumstances is one of galvanometric negativity. But during the process of recovery, when the tissue is absorbing water, this negativity diminishes, a change ,g.that is tantamount to that increase of positivity with which Fig. 200. Pitcher of Ne- penthe^ with lid removed. I 336 COMPARATIVE ELECTRO-PHYSIOLOGY we are already familiar, as the invariable accompaniment of a positive turgidity-variation. Thus, if galvanometric negativity is to be taken as the concomitant of the expulsion or secretion of fluid, it would appear that the opposite process of absorption would be indicated by the respon- sive galvanometric posi- tivity. Again the fresh pul- vinus of Mimosa responds, when excited, by a me- chanical fall, a negative turgidity-variation, and by galvanometric negativity. But after long-continued stimulation, these normal responses are found to undergo reversal. The pulvinus expands; water must be re-absorbed, and the leaf is re-erected. The normal galvanometric nega- tivity is now reversed to positivity. It will thus be seen that while, in a fresh tissue, stimulus gives rise to expulsion of Fig. 20I. Glandular Surface of a Portion of the Living Membrane of the Pitcher of Nepenthe. W fluid — the electrical indi- cation of this process being galvanometric nega- tivity — in a tissue which has already, on the other hand, been under con- tinuous stimulation there will be a tendency towards the phasic reversal of re- sponse to galvanometric positivity, indicative of the process of absorption. In the case of motile tissues, these excitatory reactions of the outflow and inflow of fluids appear to us of little consequence, except in the form of those appropriate Fig. 202. Transverse Section of Tissue of Pitcher of Nepenthe. «, outer surface ; L, inner surface ; g^ glands present in the internal surface. I RESPONSE OF DIGESTIVE ORGANS 337 motile responses which they occasion. But in glandular organs, they become possessed of much greater significance, since they constitute the main function of such structures. Electrical responses, in every way analogous to those which have been described, are obtained from the glandular surfaces of digestive organs. That is to say, the glandular surface when fresh exhibits responsive galvanometric nega- tivity on excitation. In Drosera^ for example, under these conditions secretion is seen to take place. Thus the negative phase of response, in this as in the case of Mimosa^ is asso- ciated with expulsion of fluid or secretion. After continuous stimulation, again, the responsive phase here, as in Mimosa^ is found to be reversed to positive, indicative, as there is every reason to believe, of absorption. From a consideration of the functions of the digestive organ, we should be prepared, as already pointed out, to expect the occurrence of two alternating processes. In the fresh state, ingestion of food, acting as a stimulus, would naturally induce excitatory secretion ; and this excitatory secretion must be followed later by the absorption of dissolved food. These alternating phases of secretion and absorption indubitably occur. We shall also find, in the electrical response-records, a phasic alter- nation of negative and positive under appropriate conditions. We have seen that there is a continuity between the different reactions of non-glandular and glandular tissues. We have also seen that, as in the one case, so too in the other, a phasic change takes place from negative to positive. In the digestive organ, however, we have to deal mainly with the fluctuations of fluids — secretion and absorption— and the attendant electrical variations, which consist of two opposite phases, positive and negative. As far as I have found it possible to test the matter experimentally, it has invariably been the case that the negative electrical phase was associated with secretion ; and everything points to the probability that the converse of this— the association, namely, of the positive electrical phase with the process of absorption — holds equally good. I 33^ COMPARATIVE ELECTRO-PHYSIOLOGY We now return to the question of the normal response of the pitcher in its three different conditions. Of these, in the youngest, no flies are present ; such a specimen will be known as 'fresh.' In others, somewhat older, are a few insects. These pitchers may be regarded as moderately excited, owing either to the struggles of the insects or to the supply of food, or to both. Still another class is found, in which the glandular part of the inner surface of the pitcher is practically coated with captured insects, and has thus already been subjected to long- continued stimulation. The responses of these three classes of specimens are in each case, as I shall show, very characteristic. I shall first describe experiments carried out on fresh specimens. Records were made of their responses to equi- alternating shocks of moderate intensity, at intervals of two minutes. The responsive current was here found to flow from the internal glan- dular to the external non- glandular surface. Fig. 203 gives a series of such responses. It was said at the beginning that the responses of digestive organs were likely to be multiple. This is seen to be true even under the moderate stimulus applied in the present case. But under the action of stronger stimulus, such as that of a thermal shock, the response is found to consist of a long and multiple series, records of which will be seen later. Another peculiarity to be noticed, in the series of re- sponses given in fig. 203, is that the base-line of the record Fig. 203. Photographic Record of Series of Normal Negative Responses of Glan- dular Surface of Nepenthe in Fresh Con- dition to Equi -alternating Electric Shocks given at Intervals of Two Minutes Responsive current from internal glandular to external non-glandular surface. Note occurrence of multiple response and trend of base-line upwards. RESPONSE OF DIGESTIVE ORGANS 339 trends upwards. This indicates that the glandular surface, by the residual effect of stimulation, is being rendered more and more galvanometrically negative. This explains why the internal surface of the stomach has been found by different observers to be negative, a condition of more or less persistent negativity being thus clearly due to the ex- citatory after-effect of preparation. Had it not been for the exceptional opportunity afforded by the open pitcher of Nepenthe^ it would have been impossible to make galvano- metric connections with the intact inner glandular surface and thus to ascertain that such a surface is naturally gal- vanometrically positive. I may here point out the very interesting modifica- tion of response which occurs in the same specimen under a long-continued series of stimulations. This modification, due to fatigue so-called, makes its appearance first in dimi- nution of the height of the responses. Some of the con- stituent multiple responses due to a single stimulus are then found to be reversed to positive, and after this they show a tendency to become more or less completely reversed. It is also interesting to find that the same modifications make their appearance, in the same order, in those pitchers which have been subjected to continuous stimulation, to a greater or less extent, by the supply of insects. That is to say, a pitcher containing a few insects is found to give responses, the multiple constituents of which are sometimes positive and sometimes negative. This intermediate phase is seen well illustrated in the record given in fig. 204. But in the pitcher whose inner surface is already thickly coated with insects, and which has long been exposed to the continuous action of such stimulation, the characteristic response is found to be the reverse of that of the fresh specimen. It will be seen from the record in fig. 205 that in such a case the individual effect of a single stimulus is a series of mul- tiple responses which are positive. In this record a curious [effect is again seen, that of the shifting of the base-line, now Z2 340 COMPARATIVE ELECTRO-PHYSIOLOGY downwards. This indicates an increasing positivity of the glandular surface. The results which have thus been described, in the case of the fresh pitcher, and of one subjected for a long period to the stimulus of food, are fully compatible, it will be observed, with the theory of digestion as a diphasic process, in which galvanometric negativity is associated with a predominant secretion, and the subsequent galvanometric positivity with a predominant absorption by the glandular membrane. - NIia/ m KW M Fig. 204. Photographic Record of Responses of Pitcher in Intermediate Stage, having Attracted a Few Insects Note here the occurrence of two phases in constituent responses, positive being predominant. Fig. 205. Photographic Record of Re- sponses of Pitcher in Third Stage, the whole Glandular Surface thickly Coated with Insects. Stimuli applied at Intervals of two Minutes The response here is in the positive phase, direction of current being from non- glandular to glandular. Note also the multiple character of responses to single stimuli. It is also important to notice that while in the fresh condition the glandular surface is positive, and in the moderately stimulated condition negative, yet positivity of the glandular surface is not always to be taken as a sign of its fresh condition. For we have here seen that under long- continued stimulation, the electrical condition is apt tc^ be reversed to one of positivity. RESPONSE OF DIGESTIVE ORGANS 341 It has been stated that on account of the highly excitable nature of the digestive organ, a single stimulus, if strong, i •^ W ' J\ A • \ ^ iiA/UAyv^. \ I' 'a« 4 wmammjMaHTM i. w Z ; l\ * ^ V 'vv \ 'i^ M L , . . J 1/ V U^b ifOkim 1 1 1 1 1 II 1 T j ,r^ 5' 10' I Fig. 206. Multiple Response of Pitcher of Nepenthe, in First or Fresh Stage, to Single Strong Thermal Shock The constituent responses are both negative and positive, the former being stronger. would give rise in it to a multiple series of responses. The two following records (figs. 206 and 207) illustrate this fact in two different speci- mens which were in somewhat different con- ditions. The stimulus employed in each case was a single strong thermal shock, and the multiple responses were found to persist, in both, for quite an hour. In the first of these figures, the constituent responses of the series were both negative and positive, the former being pre- dominant. In the second record (fig. 207) the positive phase is pre- dominant in the responses, and the trend of the base-line down wards shows increasing positivity of the glandular surface. Fig. 207. Multiple Response of Pitcher of Nepenthe, in Third Stage, to Single Strong Thermal Shock The constituent responses are here pre- dominantly positive. 342 COMPARATIVE ELECTRO-PHVSIOLOGY In taking up this investigation on the pitcher of Nepenthe it appeared to me that much light would be thrown, by the study of this simple organ, on the many difficulties connected with the response of the more complex digestive organs of the animal. This surmise has proved to be fully justified, for in the experiments which I have carried out in the latter field, the results are a mere repetition of these typical effects seen in Nepenthe under corresponding circumstances. Before passing from Nepenthe to the study of digestive tissues in animals, it will be well to deal here with the more complex type of vegetal digestive organ seen in the plant Drosera. I took for my experiment a specimen of the Indian Drosera longifolia^ the upper surfaces of whose leaves are covered, as is well known, with glandular Fig. 208. Photographic Record of Responses , , t tj^^^ „^ • in Fresh Leaf of /;;wra to Equi-alternating tentacles. Here, as m Electrical Shocks t^g case of the pitcher The first series show normal responses. Current r at- ; 4-U from upper glandular to lower non-glandular of Nepenthe, the re- surface. In the second series normal response gponse of leaveS which IS reversed to positive, after tetanisation, T. ^ are fresh and have not been subjected to previous excitation, is by induced negativity of the glandular surface, and this is reversed to positive under long-continued stimulation. These two phases are seen in figure 208, in which the first series is a record of normal responses of galvanometric negativity, to equi-alternating shocks applied at intervals of one minute ; and the second, the reversed responses exhibited by the same leaf, to the same .stimulus, when it has, in the meantime, been subjected to tetanising .shocks for three minutes continuously. It is RESPONSE OF DIGESTIVE ORGANS 343 curious and interesting to note here, as in the case of Nepenthe, the trend of the base-line up, when the response is the normal negative, and down when it is the reversed positive, indicating in the one case increasing negativity, and in the other increasing positivity. As in the Nepenthe^ so also in the leaf of Drosera^ specimens which arc not fresh — that is to say, previously unexcited — are apt to exhibit the positive phase of response. I give below a series of multiple responses (fig. 209) induced in such a leaf by a strong stimulation. The stimulus was in this case given by sectioning the leaf, and the response therefore illustrates the fact that preparation itself acts as a stimulus. In the present case, electrical con- nections with the galvanometer, were made with the upper and lower surfaces of the leaf on the plant, intact. On now cutting the petiole across, a long series of multiple responses, lasting for about 45 minutes, was found to be set up. These pulsations were at first rapid, and then slowed down gradually, the average period of a single pulsation being about 30 seconds. Only a portion of the record is shown in fig. 209. Having thus seen the typical responses exhibited by the digestive organs of plants, we shall- now pass to the consideration of the reactions induced in animal stomachs. Here, again, two different subjects of inquiry arise, the direction, namely, of the natural current of rest, and that of the action or responsive current. As regards the first of these, it will be remembered that Rosenthal found it to be strongly ' ingoing ' — that is to say, from the mucous to the Fig. 209. Photographic Record of Multiple Response of Leaf q\ Drosera in Positive Phase Stimulus was caused here by section of the petiole. 344 COMPARATIVE ELECTRO-PHYSIOLOGY muscular coats of the stomach. From this it was supposed, as we have seen, that the mucous coat of the stomach of the frog had the same electro-motive reaction as its outer skin. We shall find, however, that there is in reality no such similarity between the two, inasmuch as, while the excitatory reaction makes the outer skin galvanometrically positive, its effect on the mucous surface under normal conditions is to induce galvanometric negativity. In the case of Nepenthe, further, we have seen that the natural current of rest is from the non-glandular outer to the glandular inner surface, and that this is liable to reversal, as an excitatory after-effect of preparation. The ingoing current, therefore, observed in the preparation of frog's stomach, is to be regarded, not as the natural current of rest, but as the excitatory after-effect due to isolation. With regard, next, to the current of action, Biedermann states that direct electrical excitation, by rapidly alternating shocks, induces a negative variation usually preceded by a positive swing. Since the so-called current of rest is ingoing, a * negative variation ' of it evidently means an outgoing current — that is to say, galvanometric positivity of the mucous coat. Hence the responsive action of the mucous coat, as described by Biedermann, is a transient negativity followed by positivity. In dealing with this question of the electrical response of the digestive organ, we must be prepared, as the result of previous experiments on plants, to meet with variations of the excitatory effect, due to the phasic condition of the tissue. And first, for the clear demonstration of the effect of ex-' citation on the mucous surface, uncomplicated by changes induced at the second contact, I employed the Rotary Method of Mechanical Stimulation of the given area. The rotating electrodes were applied to the inside of a properly mounted frog's stomach, and experiment commenced some time after the cessation of the multiple response due to preparation. The following record (fig. 210) exhibits the first four of these responses to individual mechanical stimuli, RESPONSE OF DIGESTIVE ORGANS 345 applied at intervals of one minute. The responsive variation took place by the induced galvanometric negativity of the excited area. Under long-continued stimulation fatigue was found to be induced, the responses becoming diminished and even tending to a reversal from the normal to positive. In order to show that the inner mucous surface is relatively more excitable than the muscular coat, I next subjected the two to simultaneous excitation by equi-alter- nating electrical shocks. And for the sake of establishing a Fig. 2IO. Photographic Record of Normal Negative Responses of Frog's Stomach to Mechanical Stimula- tion \JWv^U\M Fig. 211. Photographic Record of Normal Negative Re- sponses of Stomach of Tor- toise to Stimulus of Equi- alternating Electric Shocks applied at Intervals of One Minute generalisation as to the reaction of the stomach, I now took a different specimen — namely, the stomach of tortoise. The responses in the figure (fig. 211) showed relative galvano- metric negativity of the inside of the stomach. We have seen that fresh vegetable stomach responds by normal negativity, but that, under continuous stimulation, a phasic change is induced, by which response is reversed to positive (cf fig. 208). I shall next demonstrate the corre- sponding effect in the animal stomach. Taking a preparation of the stomach of gecko, I obtained normal responses, whose direction was from the glandular internal to the muscular 346 (OMI'AKATlVr, K.I.lKTUO-niVSlOI.OC.V external sinracc. After an iiilci vcMiiiij; period of lelanisjilion, however, the responses are seen to In- reversed (fig. 212). The next record has heen selected for the purpose of showin(^ the (gradual process of transition from the normal negative to the reversed positive response. The specimen taken was frog's stomach. At the commencement of the experiment the galvanometer spot was quiescent, but when the specimen was subjected t«» a single strong thermal shock, a prolonged scries of multiple responses was initiated, per- sisting for more than an hour. Of this series 1 here re- produce four difTerenl portions (fig. 213). The first of these {a) con- sists of pulses of gal- vanomctric negativity of the internal surface. The recoveries are here incomplete, and the base-line shifts upwards, showing an increasing negativity of that sur- face. The negative pulses arc then reversed Fm. 2X2. riu„n«,„,,l,i.. Uccml „f Nomu.l »" posi'ivc, throiigh an Ros|X)nsc in Stonmoh of Gecko to v.k\\\\' intermediate di-j)hasic rtUcrimlinu Shocks, seen to he reversed nlttr /» . 1 .% r . *. Tctanisution (^> I" ^^C first part of this pronouncedly positive response (c) the base line is horizontal. It then begins to shift downwards (r/), thus exhibiting a decreasing negativity— or increasing positivity— of the internal surface. In this periodic variation of the electrical condition we have a significant parallel to the records which we have already seen in Nepenthe and in Drosera (figs. 203, 205, 208, and 209). It has already been pointed out that, in view of the functional peculiarities of the digestive organ, it might be ex- pected that the alternate reactions of secretion and absorption would neither of them be single and .spasmodic, but each long- R£SF019SE or DtGESIlVE ORGiLXS 347 :Mjtst»niedL Intliibccmiiectnn kksiii^gestivcthiitthe df^ans sliQukl sbov so stroc^^ iniilfeqpie rehouse. It mtmid iImb roffhamicail stjomlition dhMin^ iogc^lioo oC food gives the responsive l eacti o n of s c u e Uo n, e v i d cD c ed response of gdhw m ome uk ncgalivily of IlieintenHi Tliere then sets in the opposHie phw^ jfisociated with the to liar levcirsal of eiectikai lespons^ ipnktMy indicrtiiig the ab- sofptive piooess. Tliis le^pecaJ of r ey on s e^^ to gd l i nmum C U ic posdbhpilf , may be the vorit of three dB0CR»l fiMJbtxs, whidi wuay or may not be mutnally dependent In the fint place; itsdC Other things beiag eqaal^to grce itse to arevctsal of response. Secondly, after isgu e ti on bns leached its maxi- mnn^ the emp^ mnoons odk in contad widi And woild natMaDy tend to teabsoibL And» lasdy, we have of internal cneigy, in 348 COMPARATIVE ELECTRO-PHYSIOLOGY tends to give rise to a responsive reaction, whose sign is opposite to that of excitation — expansion instead of con- traction. Now such an increase of internal energy could not fail to be the result of the absorption of the chemically- dissolved food. Another interesting consideration to be remembered in connection with digestive organs is that periodically-acting forces give rise to an induced periodicity, which persists for a time, even in the absence of the periodically-exciting cause. A well-known illustration of this is met with in the nycti- tropic movements, so-called, of plants, induced as these are by the periodic variation of night and day. These move- ments persist for a certain length of time, even when the plant is kept in continuous darkness. Similarly, animals accustomed to the supply of food at regular intervals would undoubtedly exhibit alternating phasic changes apparently spontaneous, in the condition of the digestive organ in consequence of the original periodicity of the exciting cause. Such an organ, therefore, must necessarily exhibit periodic electrical variations. CHAPTER XXV ABSORPTION OF FOOD BY PLANT AND ASCENT OF SAP Parallelism between responsive reactions of root and digestive organ— Alternating phases of secretion and absorption — Association of absorptive process with ascent of sap — Electrical response of young and old roots— Different phasic reactions, as in pitcher of Nepenthe — Response to chemical stimulation — Different theories of ascent of saip — Physical versus excitatory theories — Objections to excitatory theory — Assumption that wood dead unjustified — Demonstration of excitatory electrical response of sap-wood— Strasburger's experiments on effect of poisons on ascent of sap — Current inference unjus- tified. We have seen in the last chapter that in the digestive pro- cess as a whole there must be alternating phases of secretion and absorption. The secretion of dissolving fluids, by which insoluble substances are rendered soluble, we found to take place under stimulation, and to be succeeded by a process of absorption, by means of which the now dissolved food- material found access into the organism. These functions, though seen characteristically in the digestive organs of animals, are also to be observed in some plants, such as the pitcher of Nepenthe^ or the leaf of Drosera. Here, situated externally, we find what are practically open stomachs, digesting, as do those of animals, solid organic food. But plants in general have to depend on the supply of inorganic food-material, often presented in solid or insoluble forms, for their nourishment. In this case also it is obvious that the same sequence (jf solution by dissolving fluids, and subsc- quent absorption, must be gone through. And the ors^an b y which this takes place must evidently be the root. In this regard the well-known experiments on the corrosion of marble by the root of a growing plant are sufficient to show 350 COMPARATIVE ELECTRO-PHYSIOLOGY (that these organs secrete acids, by means of which insoluble substances are made soluble. It is equally clear, further, that the inorganic solids so dissolved are afterwards absorbed by the plant. Thus it will be seen that these alternating pro- cesses of secretion and absorption of food-material, as they take place in the vegetable organism, are not very different in their essential features from the ordinary phenomenon of digestion as known to us. The chief distinction between the two would now seem to lie in the fact that in the animal the supply of foo d is in_^ the main organic, and in the plant inor- ganic. Even here, however, we meet with connecting links IrTthe form of insectivorous plants, in whose case the organic supply is obtained by means of the digesting leaf, and the inorganic through the roots. We may regard digestion, therefore, in its widest sense, as a process of abs orption of insoluble food rendered soluble, whetKer^^ch food be organic or inorganic. ApparentlyTthen, in the case of the plant the root functions as a digestive organ. But whether or not this analogy is merely superficial can only be determined by an experimental inquiry into the parallelism which may or may not exist between the various excitatory reactions of the root on the one hand and a typical digestive organ on the other. In order to obtain the large quantity of inorganic material which is necessary to the nutrition of a tree, for instance, it is clear that fresh quantities of charged fluid must be con- stantly taken up. In order further that this process may be maintained continuously it must be possible to get rid of the useless water, which is accordingly passed off, chiefly from the transpiring leaves, in the form of vapour. The absorption of food and the ascent of sap, or transpiration-current, would appear therefore to be related phenomena. I shall, in the course of the present and following chapters, then, take up in detail the consideration of these two aspects of the problem, which will thus constitute two main lines of inquiry : (i) Whether or not the excitatory reaction of the root has any similarity to that of digestive organs in general ; and ABSORPTION OF FOOD BY PLANT 35 1 (2) whether or not the ascent of sap is fundamentally due to similar excitatory reactions. With regard to the latter of these questions it may be stated here that the nature of the efficient cause of the ascent of sap is universally regarded, in plant physiology, as constituting a problem of the greatest obscurity. The various non-physiological theories which have hitherto been advanced are admitted to be inadequate, as we shall see later. We are thus confronted either with an insoluble problem or with the necessity of finding physiological reactions which' will account for the ascent of sap. As regards the latter of these alternatives, however, ob- jections apparently very serious have been brought forward. Against the physiological character of the action it has been urged {a) that wood, being supposed to be dead, could take n o_ part^ in the._aacent^ of sap. It is known moreover {b) that^kjlHng the roots with boiling water does not prevent^ the ascent j)f_sa£. And, lastly (c), in the well- known experiments of Strasburger it was found that strongly poisonous solutions can be carried to the tops of trees. From these facts it has been held to be proved that the ascent of sap cannot be dependent on the livingness of the tissues concerned. But if, on the other hand, it could be shown that these 1 objections were not valid, and if, further, some crucial experi- ' ment were devised to demonstrate that excitatory action was attended by a concomitant responsive movement of water in the tissue, it might then be claimed that the physiological theory of the ascent of sap had been established on a firm basis. The attempt to do this will form the subject of the next chapter. The first question that falls within the scope of our investigation, then, is as to whether the reactions of the root are or are not similar to those of digestive organs in general. We have seen, in the case of the latter, that as there are two opposite activities, of secretion and absorption, so also there are two opposite responsive phases, negative and positive, the 35^' COMPARATIVE ELECTRO-PHYSIOLOGY former being the more characteristic of the fresh condition, and the latter of a specimen which has been previously sub- jected to continuous stimulation. Before giving any account, however, of these electrical responses, it will be interesting to demonstrate here the occurrence of secretion in young or fresh specimens, when subjected to excitation. The fact that young rootlets secrete, on excitation by contact, has already been seen in the well-known experiment on the corrosion of marble, mentioned above. But I shall now describe a new experiment, in which this fact is even more convincingly demonstrated. I took a specimen of Colocasia, growing in marshy soil. The plant was lifted bodily, with earth adhering, and placed in water, so as to expose the roots gradually, without causing injury. It was then kept overnight, with the roots in normal saline solution, which was slowly absorbed by the tissues. Next morning, again, it was carefully washed till there was no trace of salt ad- hering. One of the very young roots was now immersed in very dilute solution of silver nitrate. If the previous washing had oeen effective, there ought now to be no white precipitate, or only the merest trace, formed in the silver solution. The two electrodes of a Ruhmkorff's coil were next connected, one with the silver solution, and the other with the stem of the plant. On now passing tetanising shocks, the immersed root became excited, and secreted its contained salt solution, this being seen in the silver nitrate as streams of white precipitate. Turning next to the electrical mode of investigation, we have found that in the digestive organs, the galvanometric negativity, which is the characteristic response of a specimen in the fresh condition, becomes reversed to positivity under continuous stimulation. In the case of Nepenthe, very young pitchers exhibited this normal response of negativity, which was converted, under continuous stimulation, into diphasic, tending towards positivity. Older specimens, again, pre- viously stimulated by the presence of excitatory food-material, ABSORPTION OF FOOD BY PLANT 353 were found to be in the positive phase, giving rise to response by galvanometric positivity. In the case of the root, it is interesting to find that there is a similar alternation of responsive phases. For this demonstration I again took the root of Colocasia, and recorded its responses to equi-alternating electric shocks. Among the mass of roots there are naturally some which are dead and decaying. One of these was selected for one electrical contact, while the other was made with a young and vigorous root. The responsive reaction, under these conditions, was found to take place by galvanometric negativity (fig. 214). Under long-continued stimulation, however, 1 have often found this normal response by galvanometric negativity to be reversed to its opposite, positivity. From this we may pass to the consideration of response in older roots, where the phasic reaction is typically positive. TVT , . . Fig. 214. Photographic Now we have seen m previous Record of Normal Nega- chapters, as will be remembered, that ^ive Response of Young Root of Coiocasia there are two different conditions under which the positive may be substituted for the normal negative response. The first is that of reversal under long- continued stimulation, which we have just seen. And the second occurs when the stimulus falls below the critical level which is necessary to the evoking of true excitation. In this latter case, as we saw further, the incident stimulus increases the internal energy, and causes expansion, positive turgidity-variation, and galvanometric positivity. It would thus appear that one identical stimulus may induce one effect, that of galvanometric negativity, in a highly excitable tissue, and the opposite, or galvanometric positivity, in a tissue that is less excitable. In connection with this question the ex- perimental results which I am about to describe are very significant. A A 1 354 COMPARATIVE ELECTRO-PHYSIOLOGY We have seen that a young root of Colocasia, when fresh, gives the normal response of galvanometric negativity. Taking next an older root of the same plant, and employ- ing the same intensity of stimulus as before, I found the responses to take place, generally speaking, by galvanometric positivity (fig. 215). This would appear to suggest a ten- dency towards specialisation of function, galvanometric nega- tivity being associated, as we have seen, with secretion, and positivity, in all probability, with the opposite— namely, absorption. A similar specialisation of certain cells for secretion and others for absorption is manifested more unmistakably in the digestive organs of the higher animals. Thus in the young roots the pre- dominant reaction would seem to be secretion, reversed under continuous stimulation to absorption. In the older roots, on the other hand, the pre- dominant reaction must be supposed to be absorptive. Here, then, judging from the electrical indications, we Fig. 215. Photographic would seem to have proof of that s"in ofcTer'Roofrf Physiological activity i,. virtue of whicii Colocasia water is taken up by the root, thus giving rise to the so-called * root- pressure.' We can also see how, by the summated activities of numerous roots, this ' root-pressure ' is kept approximately constant for a certain length of time. Taking longer periods into account, further, we can see that this physiological activity is likely to undergo periodic change, a fact which is evidenced by the known periodic variation of root-pressure. The question, however, of the actual influence of excitation on the process of the ascent of sap will be dealt with in the next chapter. One form of stimulus to whose action the roots must often be subjected is that of the chemical substances present in the soil, and I undertook to test the electrical variations ABSORPTION OF FOOD BY PLANT 355 induced by these. The results obtained, at least with the specimens which I have tried, are in general parallel to those obtained by the electrical form of stimulation. Thus, in young roots, in the majority of cases, when subjected to the action of so dilute a solution as 5 per cent, of sodium car- bonate, an electrical change of galvanometric negativity was induced. The continued action of this solution, however, tended to induce a reversal to galvanometric positivity. But the reactions of older roots were different — that is to say, in the greater number of the latter cases, a solution of '5 per cent, induced galvanometric positivity ; and it required a much stronger solution, of from 5 to 10 per cent, to bring about the reaction of galvanometric negativity. We have thus seen that in the root, as in the digestiv e organ, there ar e alternating phases of secretion and absorp - tion, and that it is by means of the secreted fluid that soli d inorganic substances are rendered solu ble^ for subseque nt aHsorption as food. We have seen moreover that the elec- trical reactions in the two cases are similar ; that in the \oung root, as in the young glandular organ of Nepenthe^ the characteristic response is by galvanometric negativity ; and that long-continued stimulation induces diphasic variation, with a tendency towards the reversal to positive. We saw further that older roots, like the glands in the older pitchers of Nepenthe^ have a phasic reaction which is predominantly positive. And now, having thus completed our first line of inquiry, we shall turn to the second — the question, namely, as to whether the ascent of sap is or is not essentially due to physiological reaction. The possible explanations of the ascent of sap may be grouped broadly under two different h eads^ as either physical or ph ysiological . Under the former of these must be named such theories as those of _atmospheric pressure, capillarity, osmosis, and evaporation from leaves. Under the latter, e physiological, the movement of water is regarded as ainly due, in some hitherto undefined way, to excitatory actions by which the sap is propelled in a uni-directioned ^pn; I 356 COMPARATIVE ELECTRO-PHYSIOLOGY manner, this primary movement being aided by accessory factors. Among the physical theories which have been pro- pounded for the explanation of the ascent of sap those of atmospheric pressure and of capillarity are admitted to be inadequate. But that of osmosis and transpiration, put forward by Dixon, Joly, and Askenasy, is of much greater weight. According to this, the ascent is brought about by transpiration from leaves. The fluid in the mesophyll cells of the leaves becomes concentrated by evaporation ; thus osmotic attraction is set up by the leaves, and the suction thereby exerted is supposed to be transmitted backwards as far as the roots, through cohering columns of water. The difficulties in the way of this theory lie (i) in explaining how a slow osmot ic action could produce so rapid a water-curren t ; and ( 2), in the absence of any conclusive proof that, under actual conditions within the plant, the water-column could have sufficient tensile strength. Even apart from these objections, however, the fact remains that energetic water- movements take place in the plant in the entire absence of transpiration. For example, sap exudes from the cut end of a tree which may exert a pressure as great as that of a column of liquid 1 3 metres in height. It is thus seen that there is an independent activity or some kind which maintains the movement of water through the plant. That this activity, moreover, is not resident in the root merely is seen from the fact that exudation of water takes place from the tips of grass-blades when their cut stems have been placed in water. Criticising the theory of trans- piration, Strasburger rightly remarks that transpiration only makes a place for inflowing water, but cannot furnish the force necessary to convey a large volume of fluid rapidly for a considerable distance through wood. From the considera- tion of these and other facts, Pfeffer, in his summary, was led to the conclusion which he states as follows : ' A satis- factory explanation of the means by which the transpiration - current is maintained has not yet been brought forward. If ABSORPTION OF FOOD BY PLANT 357 no vital actions take part in it, then it is obvious that we have only an incomplete knowledge of the causes at work and of the relationship of the different factors concerned.' ^ I The inadequacy of these theories to explain the ascent of sap has, then, been freely admitted. With special refer- ence, further, to that of osmosis, I shall myself be able to show that the movement of water often takes place in the plant, in a direction contrary to what it would be if osmosis alone were involved. The fact that the absorption of water is not a merely passive process, but a phenomenon connected with irritability, will be further shown in the depression of the rate of water-movement by such conditions as depress irritability, whereas the opposite circumstance will be found to enhance it. We are thus driven to examine the possibility of a physiological explanation of the ascent of sap. On such a theory it must be supposed to be brought about by the action of stimulus, inducing reactions expressed in the re- sponsive movement of water. The objections made to the physiological explanation have already been recapitulated. They are ( i ) that the movement of water is known to tak e place rapidly, and by preference^ through woody ^issues which are supposed to be dead ; and (2 ) that when the roots have been killed by hot water, or when poison is supplied, the transport of sap continues to take place,. I shall now proceed to examine these arguments, and to show that the objections raised, though apparently so strong, are not really valid. We shall first refer to the argument which has been based upon the fact that in trees conduction takes place very rapidly through woody portions which are regarded as dead. It is not, it must be noted, implied by this that the presence of wood is essential to the ascent of sap, in- asmuch as even in trees there are tracts of living cortical tissue in the roots which have to be traversed before the water can reach the woody tissues. In seedlings of Gramince t' Pfeffer, Physiology of Plants (English translation ), 1903, p. 224. 358 COMPARATIVE ELECTRO-PHYSIOLOGY only one or two days old, again, water ascends, and is ex- creted at the tip of the yet unopened leaf. Transpiration is here at its minimum, and the fibro-vascular elements at this early stage cannot be regarded as * dead wood.' Finally, I in herbaceous plants, where woody elements are insignificant, the ascent of sap is seen to take place. Assuming, for the sake of argument, that in the case of the tree, the mass of wood in the interior were dead, it might still conceivably be of use in the irrigating system as a central reservoir. This would certainly be advantageous to rapidity of transit. At the lower end of the tree, the wood abuts upon the delicate parenchymatous tissues of the root, and at the upper upon those of the leaves. According to the physiological theory, then, it might be supposed that it was by the multiple activity of the cells of the root that water was pumped into the wood ; and that at the other end the central reservoir was able to furnish a supply to make up for the constant loss by transpiration. Laterally also, in the stem itself the cortical tissues could draw upon this central supply. Under such an arrangement no part of the plant could be very far away from the reservoir. As a matter of fact, this sketch corresponds roughly to the working of the tree as an hydraulic machine. The system is, however, somewhat more complex than has been indicated. Besides the central, we have also to remember the presence of lateral reservoirs, in the parenchymatous tissues of the cortex. But tne transport of water through these is not, of course, so rapid as through the central, more specifically conducting, system. In the case of herbaceous plants, where the quantity of wood is insignificant, we may regard the central channels as abolished. Here we have soft cortical tissues extending continuously from root to leaves through the stem, and it is obviously through these that the ascent of water takes place. In woody trees, then, there is no reason to suppose that the cortical tissues could not play a similar part in the conveyance of water. The difference is, that in this case there is also an added and ABSORPTION OF FOOD BY PLANT 359 better channel available, which will naturally come into requisition where quick transit is required. In a woody trunk, then, we have (i) the ou ter cortical cylinder of water-conducting tissues^ by which the ascent of sap takes plac e slowly. We have (2) the highly-conductin g central woody tissue, which not only allows of water ascend- ing rapidly through it, but is also (3) in lateral communi- cation with the outer cylinder. The hydraulic system thus consists of a large central canal, as it were, connected with innumerable lateral reservoirs, which are the cells of the cortex. When a demand arises for rapidity of water-supply on account of transpiration, we can now see that no less than three different factors are brought into requisition. First there is the rapid upward transit through the wood ; secondly, the slow ascent through the cortex ; and thirdly, the lateral supply from the cortex by way of the nearest wood. As regards the last of these, the cortical tissues in contact ^ with the wood act in a manner not very unlike that of the v. ^*^ roots towards the soil. That is to say, under different cir-V V ' ^ cumstances, they absorb water from it, and excrete^jwater 1 ^ into it, these alternating processes being by no means ^^ accidental, but guided by appropriate excitatory reactions. Turning our attention for a moment to the movements of Mimosa leaf, we find that on excitation the expelled water makes its way to the fibro-vascular tissue. There is here, in the excitable tissue, unlike the case of secretory organs, no external vent, and we see the necessity of a central reservoir to which water excitatorily expelled may find access. On the subsidence of excitation, the water is re-absorbed by the organ, causing expansion and re-erection of the leaf • Such movements of inflow and outflow evidently take place in the trunk of the tree itself Under the stimulus of sunlight, the excited cortical tissue will squeeze water inwards into the central reservoir. If this takes place, the effect will be seen in a diametric contraction. At the time when transpiration is most rapid, under the action of sunlight, there is thus 360 COMPARATIVE ELECTRO-PHVSIOLOGY besides the water coming from the roots, an additional supply available from the lateral reservoirs. The loss of water thus sustained by the cortex during the day is made up again at night, when it will suck water outwards from the central reservoir. We have here a case analogous to the action of the excitatory tissue of the pulvinus of Mimosa expelling water into the wood on excitation, and re- absorbing it on the cessation of excitation. The occurrence of these reactions in the cortex explains the observation made by Kraus that the organs of the plant diminish in bulk from morning to afternoon, the reverse process taking place from afternoon to morning. We have thus seen how important a factor is excitatory reaction in the observed movements of water, even on the supposition that the woody tissue, being dead, is a merely passive agent. The question has still to be attacked, however, whether this assumption, so generally made, is correct, that the wood used for conduction of water is dead. This supposition has arisen from the chemical transformation undergone by the protoplasm in woody vessels. We have seen, however, in the case of the epidermal cells of the skin^ that it is possible for chemical transformation to occur, without necessarily being accompanied by the death-change. Before proceeding to inquire whether the conducting woody channels are really dead, it is desirable to say a few words as to the particular tissues in the wood, which are most effective for this purpose. Many experiments have been carried out to determine this. Among other things various staining fluids have been employed. But an objec- tion raised in the case of some of these has been that the water of such solutions travels faster than the dye dissolved in it. For my own part, I have found the employment of dilute solution of phenolpthaline to be exceedingly delicate and useful for the purpose of this investigation. It is perfectly colourless, and the staining appears only after appropriate development. The cut end of the stem is placed in this dilute solutiop and l^ft for som? time, Transverse ABSORPTION OF FOOD BY PLANT 361 sections of the stem at different heights are then made and placed in the field of a microscope. There is now nothing distinguishing to be seen, as the solution sucked up by the stem was colourless. Dilute solution of potash, however, will act on this as a developer. The particular tissues, therefore, through which the solution has been conducted, on now being subjected to the action of this agent, become of a rich crimson colour. By this means it is easy to see, as already determined by various observers, that it is the younger, or * sap-wood,' which is concerned in the work of the rapid conduction of water, the older, or ' heart- wood,' being ineffective for this purpose. If, however, the wood taking part in the ascent of sap had been dead, and acted as a passive agent merely, it is difficult to understand the reason of this selective action of the younger, and presumably more living, woody tissues. It occurred to me, finally, that as electrical response is an indubitable concomitant of the excitatory reaction of living tissues, the question as to whether sap-wood was alive or dead could be subjected to a decisive test. For this purpose I took various strips of sap-wood from different woody plants. The cortical tissue was in each case carefully removed, and the specimens were placed in water, allowing them a period of rest. The first experiment was to observe whether local stimulation by the Rotary Mechanical Stimulator did or did not evoke electrical response. I found from this, that mecha- nical excitation of the sap-wood induced considerable excita- tory response of galvanometric negativity. I then subjected the same tissue to the action of boiling water for a length of time, and again tested its electrical reaction by the same method. The wood was found to be very resistant to the action of heat, and if was only after long immersion that the responses were entirely abolished. Drying was in fact found, significantly enough, to be an easier method than the application of heat, to kill, and therefore to abolish the responsiveness of, the wood. If the wood be first dried, and_ then soaked in water, it entirely ceases to manifest electrical 362 COMPARATIVE ELECTRO-PHYSIOLOGY response. The ordinary wood of commerce exhibits no response. The familiar fact that the cut end of a woody- stem, when not placed in water immediately, ceases to suck up water, has been supposed to be due solely to the intervention of air-bubbles. From the experiment which I have described, however, it would appear that the death of the exposed tissue by drying must be included here as a factor in this abolition of suction. With sap-wood I was also able to obtain the in- dication of galvanometric negativity in response to thermal stimulation. Hot platinum wire was applied at a distance of 5 mm. from the proximal contact. Response was thus due to the transmitted effect of excitation. I was next desirous to obtain photographic records of the normal response of living wood and its variations under chemical agents. For this purpose I employed both the electrical and vibrational modes of stimulation. For the first of these, the strip of sap-wood was cut in the form of a two-pronged fork, of which one prong was killed by exposure to the drying influence of the air, while the other was kept alive by immersion in water. The specimen was now placed in water as a whole, in order to moisten the dried half After this, electrical connections were made in the usual manner with the killed and unkilled ends of the speci- men. On next subjecting it to equi-alternating shocks' response was obtained as induced galvanometric negativity of the living prong. These responses are seen in fig. 216 in the first series of records to the left. Chloroform was next Fig. 216. Photographic Record of Electrical Response of Sap-wood The normal negative responses seen in the first series are depressed after application of chloroform in the second. ABSORPTION OF FOOD BY PLANT 363 applied, and we observe the consequent depression of re- sponse ; when the chloroform was blown off the responses were found to undergo revival. In the next experiment, a specimen of living wood was mounted in the vibrational apparatus, and its normal re- sponses taken. I next applied copper sulphate, and the record shows the consequent abolition of response (fig. 217). I have thus been able to establish the fact that the woodv vessels of t he sap-wood are not dead but living, and hence fully susceptible of physiological rea ctiorT This will, I think, be found to dispose of one of the difficulties raised in regard to the physiological theory of the ascent of sap. We come, secondly, to the objections that have been based on the ground of the ascent of sap through a tree whose roots have been killed by boilinjT •^ '^ Fig. 217. Photographic Record water, and, further, on the ex- showing Normal Responses ot periments of Hartig and Stras- I'Sus.'^r .h: A^i^riM burger. These observers set cut Response by a Toxic Dose of ends of trees in tubs of poisonous "^^'^"^ u p a e solutions, such as copper sulphate, which were found, in spite of their toxic character, to ascend to the leaves. It is clear that if such violent protoplasmic poisons ascend the trunk, they must kill all the cells lying in their path. And from this it was inferred that the living cells in the stem could not be necessary to the rise of sap. Strasburger was thus led to the conclusion that 'the supposition that the living elements in any way co-operate in the ascent of the transpiration current is absolutely precluded.'^ It does not, however, appear that this inference on the part of Strasburger was justified, for we must remember the fact that any cut piece of stem when placed in water is found to exhibit suctional activity. Hence the active cells con- cerned — if the process is to be regarded as due to such — ' Strasburger, 7^^;i;/-^(7<7/^ > 12-8 „ I -8 mm. per second 1-9 mm. ,, 2-1 mm. ,, Specimen II. — Centrifugal Transmission The distance traversed by stimulus was 38 mm. Stimulus Time Velocity 1 •01 Microfarad charged to 8 volts ,, ,, 16 ,, , „ . „ 24 „ 1 „ M 32 „ 1 1 -6 seconds 10-2 „ lO-I ,, 9 9 M 3-27 mm, per second ^ 372 mm. ,, 376 mm. 3-83 mm. V G G 450 COMPARATIVE ELECTRO-PHYSIOLOGY With regard to the effect of temperature, I found that cold reduced the velocity of transmission. Thus, in one experiment, slight cooling reduced it to one-third, and when carried still further, it abolished the conductivity altogether. A rise of temperature, on the other hand, had the effect of enhancing velocity of transmission. The following table shows that a rise of temperature from 30° C. to 35° C. doubled the velocity, and that at 37° C. the rate was almost three times that at the first temperature. The velocity was in this case determined in the centrifugal direction. Table Showing the Effect of Rise of Temperature on Velocity. Distance traversed by stimulus 41 mm. Temperature Time Velocity 30° c. 35° C. 37° C. II seconds 5-5 „ 4-5 ,, 37 mm. per second 7-4 mm. ,, 9-1 mm. ,, Transmission of excitation, as I have shown elsewhere, and shall show again, is depressed or abolished by the action of anaesthetics. We shall also see, further, that the polar effects of currents on the velocity of transmission are the same in the plant as in the animal, being opposite, accord- ing as it is the anode or kathode. In the case of a so-called * sensitive ' plant, by taking advantage of the motile indica- tions afforded by the leaf or leaflet, it is possible to determine the velocity of transmission of excitation and its modifica- tions. With ordinary plants, however, no such indications being available, it is obvious that we must find some other means of detecting and observing the excitatory wave during transit. One such I have described elsewhere as the Electro- tactile Method. It is found that the passage of the excitatory wave, even through an ordinary tissue, brings about minute form-changes. These give rise to pressure-variations as between two enclosing contacts. And this variation of pressure, in turn, can be recorded by means of a sensitive electrical device. VELOCITY OF TRANSMISSION OF EXCITATION 45 1 There is, however, a more direct way of detecting the excitatory wave during its passage through a vegetable tissue. In this — the Electro-motive Method — the galvano- meter takes the place of the motile leaflet. It has been shown that when the plant tissue is directly excited, the state of excitation is invariably accompanied by an electro- motive variation, the excited point becoming galvano- metrically negative. Hence any excitatory wave which is transmitted through the tissue will always have an electro- motive wave as its strict concomitant. The moment, there- fore, at which excitation reaches any given point, may always be determined by observing the arrival at that point of the excitatory electrical disturbance of galvanometric negativity. In order to prove that the arrival of excitation at the given point is attended by this .specific electrical response, we may perform an experiment on a plant such as Biophytum, which is provided with motile leaflets. One of the indicating leaflets is attached to the optic lever, its base being connected with one of the electrodes of the galvanometer, while the second is attached to a distant point on the leaf The two spots of light, one from the optic lever indicating the mechanical response, and the other from the galvanometer, indicating the electrical, are so adjusted as to lie one above the other, on the same revolving-drum. On now applying a stimulus, say thermal, at a distant point, it will be found, after the lapse of a definite interval, that both spots of light are deflected at the same time, showing that both alike give an outward indication of that state of molecular disturbance which is synonymous with excitation. These manifestations, of both kinds, would therefore take place at an identical moment, if only the inertia of the two indicators were absolutely the same. But, just as the same impulse would be indicated at slightly different times, if one indicating-lever were light, and the other heavy, so here also there may be a slight difference as regards time etween the appearance of the mechanical and electrical i 452 COMPARATIVE ELECTRO- PHYSIOLOGY responses, according as the virtual inertia of the one indicator exceeds that of the other. Jj\ determining velocity of transmission by the Electro- motive Method, a previous experiment gives us the loss of time due to the inertia of the galvanometer. This, deducted from the observed interval between the application of stimulus and response, gives the time required for trans- mission through the given distance. In this manner I have been able to determine the rate of transmission of excitation in ordinary plants. I give below a table which shows these velocities as determined by me in the case of sensitive plants, and of ordinary plants, and for the purpose of comparison, those obtained by other observers, in the nerves of some of the lower animals, from which it will be seen that all these are more or less of the same order. Tables giving Velocities of Transmission of Excitatory Wave (a) Animal. Subject Velocity Nerve of Anodon .... Nerve of Eledone (observed by Uexkiill) ID mm. per second •5 to I mm. ,, (3) Sensitive Plants. Subject Velocity Mimosa pudica : petiole .... Neptiinia oleracea : petiole .... Biophytum sensitivum : Petiole of, direction centripetal . Petiole of, direction centrifugal . Peduncle of 14 ram. per second I'l mm. ,, 2*1 mm. ,, 3-8 mm. „ 37 mm. {c) Ordinary Plants. Subject Fern : isolated nerve of Fictis religiosa : stem . Cticurbita : tendril Jute : stem Artocarpus : petiole . Velocity 50 mm. per second 9-4 mm. ,, 5 mm. ,, 3*5 mm. ,, •54 mm. ,, VELOCITY OF TRANSMISSION OF EXCITATION 453 Since the conduction of excitation takes place by the transmission of protoplasmic changes, it is evident that it must occur most easily along those paths in which there is greatest protoplasmic continuity. It is clear, then, that certain elements in the fibro-vascular bundles will furnish the best conducting medium. Cells of indifferent tissue, on the other hand, like the parenchyma of the leaf, are divided from each other by more or less complete septa, the fine filaments, by which neighbouring cells may be protoplasmically connected, being so minute that the conduction of stimulus through such imperfect channels must be comparatively feeble. Such tissues are, therefore, indifferent conductors of excitation, the stimulus remaining more or less localised in them. Plant-organs, then, which contain fibro-vascular elements, such as the stem, peduncle, and petiole, are for that reason relatively good conductors. Conductivity in such an organ, again, is, as we should expect, much greater along the length than across. I shall now describe an important method by which the relative conductivity of a tissue in different directions may be experimentally determined, verifying by its means the difference in the power of a tissue to transmit stimulus longitudinally and transversely. For this purpose I took a thick peduncle of Musa, and made two electrical con- nections, of which one was at a fixed point B, transversely situated as regards c, the point of application of stimulus. The second point, A, was longitudinally above C, and its distance from it could be varied in successive experiments (fig. 273). If we now take a point. A, in such a position that CA is equal to CB, then, on account of the better conductivity along CA, the excitation will reach the A contact earlier than that at B, making that point galvanometrically negative. The direction of the first responsive current, therefore, will be from A->B in the tissue. If, next, the longitudinal contact be moved to A", that is to say, so far that the excitation reaches the B contact first, then the responsive current will I 454 COMPARATIVE ELECTRO-niVSIOLOGY be reversed, flowing now from b->a". A point of transition, or of balance, a', may now be found by searching, at which the movement of the exploring contact, nearer or further, will give rise to opposite responsive currents. The con- ductivity along the longitudinal direction will then be, to that in the transverse direction, as the balancing- distance CA' is to CB. With a given specimen of the peduncle of Musa the transverse distance CB was 37 cm., and the longitudinal balancing-distance CA" was determined at 10-4 cm. Hence- the longitudinal velocity was 28 times that in the transverse direction. Fig. 273. Experimental Arrangement for Comparing the Relative Conductivities in Transverse and Longitudinal Directions c, point of application of stimulus ; b, permanent transverse contact ; A, a', a", exploring points of longitudinal contact for obtaining balance. It has been shown that different tissues in the plant may possess extremely different powers of conducting stimulus. In animals there are specialised channels of conduction known as nerves, and in plants also I have been able to discover similar conducting tissues, which can be isolated for the study of their responsive peculiarities. Experiments on this subject will be related in detail in Chapter XXXII. It may be said here, however, in anticipation, that the velocity of transmission of true excitation through these nervous channels is, generally speaking, fairly high, being at the rate of about 50 mm. per second in the case of isolated VELOCITY OF TRANSMISSION OF EXCITATION 455 nerve of fern. This, for the relatively sluggish vegetable tissue, is undoubtedly very high. In connection with this question of velocity of trans- mission, a fact not hitherto taken into account is, that there are two distinct kinds of nervous impulses, travelling with different velocities — namely, the hydro-positive and the true excitatory negative. Of these the velocity of the former is greater. In the nerves of higher animals, where the velocity of transmission of true excitation is also great, it is not generally easy to distinguish one from the other, so rapid is their succession. But their occurrence as distinct waves, even in animal tissues, I shall be able to demonstrate in a subsequent chapter. In plants, however, where the velocity of transmission of true excitation is not very high, it generally lags perceptibly behind the positive wave (p. 59). Burdon Sanderson, in his determination of the velocity of trans- mission of excitation in Dioncea^ arrived at the exceptionally high result of 200 mm. per second. I have shown, however, that the wave whose velocity he measured was not of true excitation, but of hydro-positive disturbance (p. 231). In the present chapter it has been my object to demon- strate the reality of true excitatory propagation in plants similar to that in the animal. The examples given will be found more fully described on referring to my book on ' Plant Response.' I shall, however, in the course of the present work describe new and extremely delicate means by which the modifications of conductivity may be studied in plants unde • varying physiological conditions. CHAPTER XXXI ON A NEW METHOD FOR THE QUANTITATIVE STIMULATION OF NERVE Drawbacks to use of electrical stimulus in recording electrical response— Response to equi-alternating electrical shocks — Modification of response by decline of injury — Positive after-effect — Stimulation of nerve by thermal shocks — Enhancement of normal response after tetanisation — Untenability of theory of evolution of carbonic acid — Abnormal positive response converted into normal negative after tetanisation— Gradual transition from positive to negative, through intermediate diphasic — Effect of depression of tonicity on excitability and conductivity — Conversion of abnormal into normal response by increase of stimulus-intensity — Cyclic variation of response under molecular modifica- tion. In the study of the electrical effects of excitation on the nerve, the chief experfmental difficulty lies in the selection of a form of stimulus which can be made quantitative. In such investigations it is usual to employ the electrical form of stimulus, because of the great facilities which it offers. A marked drawback to its use, however, lies in the fact that unless extraordinary precautions are taken it is liable to lead to serious error. It must be remembered that for the detec- tion of responsive variations in the nerve an extremely sensitive galvanometer has to be employed. The excitatory effect which is to be detected being indicated by the relatively feeble electrical response, and the form of stimulus being also electrical and being of high intensity, the results are liable to be disturbed in an unknown manner by leakage of the stimu- lating current. In some cases it is possible to take the bold step of including the experimental nerve itself in a circuit in which the exciting coil and the galvanometer are in series. Under these circumstances, and employing strictly equr-alternating QUANTITATIVE STIMULATION OF NERVE 457 -WM. shocks, we have seen that the resultant response is due to the differential excitabilities of the two nerve- contacts A and B. If, for instance, we wish to obtain the responsive reaction of one point only, say A, uncomplicated by that of B, it is only necessary to abolish the excitability of the latter. This can be done to a greater or less extent by injury, as, say, by making a transverse section, or by scalding. Response will then take place by the induction of relative galvanometric negativity at A. In fig. 274 is seen a series of records obtained in this manner. The responses here apparently indicate growing fatigue of the nerve. They also exhibit the positive after- effect With reference to the method of obtaining response by injuring one con- tact, commonly employed, it may be said that the assumption that the ex- citability of the injured point is totally abolished is not justified ; for I have found that though recent injur}'- causes a great depression of excitability, yet after a lapse of time the injured point tends to recover its excitability to a greater or less extent. In such a case we may expect two different effects to be exhibited in the responses. The re- sultant response being due, as we have seen, to the differential excitability of A and B, the gradual restoration of the excitability of B will progressively diminish the amplitude of the resultant response, thus giving it the appearance of fatigue. Under these conditions, and after a sufficiently long interval, response may almost disappear. This appears to me to be the true explanation of the gradual fall in the amplitude of response, when the specimen is a nerve, having one contact at the transverse section. It also explains why, in such a nerve, a fresh section, causing Fig. 274. Response of Frog's Nerve under Simultaneous Excita- tion of both Contacts, by Equi - alternating Electrical Shocks, one Contact being Injured Note the positive after- effect. 458 COMPARATIVE ELECTRO-PHYSIOLOGY renewed depression of excitability, is necessary in order to obtain renewed amplitude of response. The second effect due to this depression, without abolition of the excitability of B, is seen in the diphasic character of the responses. The positive after-effect observed in the record shown in fig. 274 may thus be ascribed to the later induction of negativity at the depressed point B. The electrical re- sponse of the nerve is apparently liable in this way to great variations, when the method of record employed is differential. But it must be remembered that true characteristic variations of the response as determined by physiological modification can only be obtained by finding some means which shall be strictly independent of this differential factor. With this object, I have succeeded in devising a new mode of observing and recording the direct effect of stimulus on the nerve, uncomplicated by the differential factor. In a subsequent chapter we shall, using this method, be able to determine the conditions which induce the characteristic variations in the response of nerve, from the staircase increase to the fatigue- decline, or even reversal, through the intermediate phase of uniform reponses. The method which has just been described, of exciting the nerve at both contacts by equi-alternating shocks, is not applicable, however, where the object of investigation is the conductivity of an intervening tract of nerve between the exciting and the led-off circuits. Here the employment of electrical shocks as exciting stimulus gives rise to disturbing unipolar effects, which persist even when the physiological conductivity of the intervening tract is destroyed as, say, by ligature or by crushing. Thus — ' If the nerve of a frog's leg is laid across two electrodes connected with the poles of a secondary coil, so as to close the induction circuit, a ligature being then applied to the myopolar tract, tetanus may still be observed in the isolated leg, on making the lead off from it at a certain distance of coil. . . . These unipolar effects QUANTITATIVE STIMULATION OF NERVE 459 may obviously be very disturbing, and are indeed pro- ductive of fallacies in vivisection and also in experi- ments with the galvanometer, if not avoided by due precautions. Hering has pointed out that in experiments such as the investigation of the negative variation of nerve-currents, in which galvanometers and exciting circuits are separated by a long tract of nerve, the most complete insulation of the two circuits is no guarantee against the overflow of induced electricity through the interpolar part of the nerve into the galvanometer circuit. . . . This kind of unipolar stimulation is an obvious danger in all experiments on action-currents and negative variation in nerve, while it shows what narrow bounds restrict the intensities of current that may be safely used in these experiments.' ^ From this it will be seen how important it is to have at our command some non-electrical form of stimulation, when the response to be recorded is electrical. Heidenhain em- ployed a mechanical form of stimulation, by which the nerve ■ was subjected to blows from an ivory hammer, which was kept vibrating by means of an electro-magnetic arrangement. The employment of this mode of stimulation would there- fore eliminate all that uncertainty — arising from the possible escape of current — which is inseparable from the use of electrical stimulus. Though this method must be regarded as one of great value, yet it is impossible to say how far the excitability of a given point in a structure so delicate as nerve will remain unmodified under the repeated action of such blows. In any case, it appeared desirable to inquire whether there was no other non-electrical form of stimulus that could be rendered practicable. Besides the mechanical, the only remaining non-electrical forms of stimulus are the chemical and the thermal. Of these, the former is obviously incapable either of repetition or ' Biederma.nn, £/earo-r/iysio/o^y {EngMsh translation), 1898, vol. ii. pp. 222- 223. I 460 COMPARATIVE ELECTRO-PHYSIOLOGY of being rendered quantitative. As regards the latter, 1 have already shown its practicability for experiments on excitatory phenomena in vegetable tissues. Thus a single loop of platinum wire may be made closely to surround the experi- mental tissue, A definite current sent through the platinum loop for a given length of time will now subject the encircled area to a sudden thermal variation, which acts as a stimulus. Successive closures of the circuit for a definite length of time are ensured by means of a key actuated by a metro- nome. The intensity of stimulus may be graduated in a pre- determined manner by the adjustment of the heating-current. Excitation may then be caused either by one or by a summated series of thermal shocks. I was now desirous of determining whether this form of stimulation would prove advantageous to experiments on the nerve, and in the course of the investigation I found it to be extremely convenient and appropriate. With good speci- mens of nerve I have been able, using thermal stimulus, to obtain long-sustained records of perfectly regular ' responses. As regards its pliability and facility of application this form of stimulus is quite unique. How many difficult problems are made possible of attack by its means will be realised in the course of the two following chapters, where the responsive variations of different conducting tissues under changing conditions are subjected to investigation. In order to obtain the electrical responses of animal nerve — that of frog, for example — the distal contact is killed and appropriate electrical connections made with the galvano- meter. The heating current is then adjusted for the desired amount of excitation. The thermal variation, it must be remembered, should not be so great as to injure the tissue in any way. The platinum loop is not in this case in contact with the specimen, and this is the mode generally employed. Should a more intense stimulation be desired, however, the nerve may be allowed to rest on the platinum loop. In such a case care must be taken to see that the rise of temperature is not so great as in any way to injure the QUANTITATIVE STIMULATION OF NERVE 461 tissue. The nerve, as usual, must be enclosed in a moist chamber, a convenient form of which, as employed in practice, will be seen in fig. 291. I shall next give a few records in illustration of the ease and efficiency with which this mode of stimulus may be applied. These records will show the characteristic varia- tions of response given by the nerve under different con- ditions. When making records of electrical responses with frog's nerve, under electrical stimulus, Dr. Waller obtained re.sponses of three different types. The first of these was the normal, and consisted of negative responses ; the second was diphasic ; and the third was the abnormal positive. This last he regarded as characteristic of stale nerve. These normal negative responses of the first of the three classes were found by him to undergo enhancement after a period of tetanisation ; while the third, that of the abnormal response of stale nerve, underwent a change into diphasic, or a reversal to normal, after tetanisation. From the fact that carbonic acid enhances the normai negative response of nerve, Dr. Waller has suggested that the enhancement of response in normal nerve after tetanisa- tion, and the tendency of the modified nerve to revert to the normal, are results of the hypothetical evolution 01 carbonic acid in the nervous substance, due to metabolism accompany- ing excitatory reactions. It must be said, however, that no ^^ trace of the presence of carbonic acid has yet been detected ^Kn such cases. I shall be able to show, moreover, that these ^Kefifects are in no way due to the evolutions of carbonic acid, ^^Bbut take place in consequence of molecular changes induced I^Kin the responding tissue, which find concomitant expression ^^Bin changes of conductivity and excitability. I shall now give records of responses of these various types obtained under the action of thermal stimulus. In ^Border to exhibit the effect of tetanisation I give, in fig. 275, ^Ka series of normal responses by induced galvanometric " negativity, given by nerve of frog in its normal excitatory condition. This nerve was then subjected to tetanic thermal 462 COMPARATIVE ELECTRO-PHYSIOLOGY shocks, after which its responses to individual stimuli of the former intensity were recorded once more. The subsequent responses show, as is seen in the record, an enhancement of amplitude. The next series of responses, in fig. 276, exhibits abnormal galvanometric positivity. It may be mentioned here that these abnormal responses are not, as supposed by Dr. Waller, exclusively character- istic of the stale condition of the nerve. For employing other and more delicate methods of record I have found even fresh nerves, under certain conditions, to exhibit this effect. Neither is this positive response due in general to any chemical degradation. Instead of this, as we shall see in the present and succeeding chapters, it may be attributed to the run-down of the latent energy of the specimen, a process which becomes accelerated in isolation. When such a de- pressed specimen is supplied again with the requisite energy, it becomes normally, or even supernormally, ex- citable. The first part of the following record (fig. 276) gives a series of abnormal positive responses obtained from a specimen of frog's nerve, which was in a somewhat sub-tonic condition. After the appli- cation of tetanic thermal shocks it will be noticed that the responses in the second part of the figure have become converted into normal. Fig. 275. Enhancement of Amplitude of Response, as After-effect of Thermal Tetanisation, in Frog's Nerve The first three responses are normal. Brief thermal tetanisation is here applied, and the responses subse- quently obtained under original stimulation are seen to be en- hanced. QUANTITATIVE STIMULATION OF NERVE 463 Between these two extremes of normal negative and abnormal positive responses there lies the intermediate diphasic. All these — positive, diphasic, and negative — may be exhibited in the same specimen, in the course of a sus- tained record of responses to single stimuli, without tetani- sation. This fact is illustrated in fig. 277, where the first series shows the unmixed abnormal positive. The response then passes by a gradual transition into diphasic — positive followed by negative — and this phase, lastly, is succeeded by a series of purely negative responses. We come next to the ex- planation of these phenomena. We have seen that on account of isolation the tonic condition of a highly excitable tissue will undergo a gradual decline. On account of this its ex- citability and conductivity will fall below par. We have also seen that in this de- pressed condition the normal response by negativity tends to be reversed to positivity. With regard to the con- duction of excitation it may be said that this condition of depression will lower the power of the tissue to conduct true excitation. Thus a stimulus of given intensity, capable under normal conditions of transmission to a certain distance, will, when the tissue is thus depressed, fail of conduction to the same distance. It will now, therefore, be the hydro-positive effect of stimulus which will make its appearance alone at the distant responding point. And the electrical expression of this will be galvanometric positivity. Fig. 276. Conversion of Abnormal Positive into Normal Negative Re- sponse after Thermal Tetanisation Abnormal positive response to left converted into normal negative on right, after intervening tetanisa- tion. I 464 COMiPARATIVE ELECTRO-PHYSIOLOGY We have also seen that a tissue which is not in the highest tonic condition may have its tonicity increased by the action of impinging stimukis, with consequent enhance- ment of its excitability. I shall also demonstrate, in Chapter XXXIV, that the effect of an impinging stimulus on a sub-tonic tissue is a similar enhancement of con- ductivity. The result of this will be either (i) that a tissue which has already conducted a moderate intensity of Fig. 277. Gradual Transition from Abnormal Positive, through Diphasic, to Normal Negative Responses in Frog's Nerve Cf. similar effect in response of skin of gecko, fig. 191. stimulus to a distant point will show, after continuous stimu- lation, an enhanced power of conduction ; or (2) that in a very sub-tonic tissue, in which true excitation has at first failed to reach the responding point, the true excitatory negative is subsequently transmitted instead of the hydro- positive alone. Under actual experimental conditions, where the stimulus is applied at a distant point, the twofold effects of exaltation of excitability and conductivity under tetanisation both come into play. In normally responding nerve, the increased con- QUANTITATIVE STIMULATION OF NERVE 465 duction of excitation, and the enhanced excitability of the responding-point, give rise to an increased amplitude of response after tetanisation, as already seen in fig. 275. In a depressed nerve, as the transmitted effect is positive, and the tendency of the responding point itself, owing to sub-tonicity, is to the abnormal positive, the record will exhibit the abnormal positive alone, as in fig. 276. But under a series of successive stimuli, the conductivity and excitability of the tissue are both gradually raised, and the effect of this is seen in the consequent gradual restoration of the normal negative response, through the intermediate diphasic (fig. 277). Or, if we do not wish to trace out the intermediate steps of transi- tion, we may tetanise the depressed nerve for a certain length of time, and record only the terminal change to the restored normal negative, as is seen in fig. 276. Taking one of the extreme cases — say that in which the response to transmitted stimulus is positive, and is converted into normal negative after tetanisation —we see that the first result is due to inefficient conductivity, allowing only the hydro-positive effect to cause response. After this, increasing conductivity, making an increasing transmission of true exci- tation possible, gives rise to a diphasic, and ultimately to the normal negative response. This result is analogous to the three types of responses — positive, diphasic, and negative — which we have already obtained with the imperfectly con- ducting tissue of the petiole of cauliflower and the tuber of potato (figs. 47, 48). We there saw that where excitatory efficiency of transmitted stimulus was sufficiently great, it gave rise to the normal negative response. When this, how- ever, was not so great, we obtained the diphasic. Finally, when the true excitatory effect could not be transmitted, only the abnormal positive response appeared. That gradation by which the transmitted stimulus was made fully, partially, or non-effective, to induce true excitation, was simply and most conclusively carried out in the case of the potato, by removing the point of stimulation to an increasing distance from the responding point. In the cases H H ■ 466 COMPARATIVE ELECTRO-PHYSIOLOGY described, then, the three types of response are exhibited by the same tissue, in indubitable relation to the variation of its effective conductivity. .If, then, results exactly parallel can be demonstrated to occur in the case of nerve also, it follows that there is no necessity there to make any such hypothetical assumption as that of the evolution of carbonic acid, suggested by Dr. Waller, in explanation of the conver- sion of abnormal response to normal. In order to show how a varying conduction will give rise to these three types of responses, I shall now describe an ^AAA, M Fig. 278. Abnormal Positive Response converted through Diphasic to Normal Negative under the increasingly Effective Intensity of Stimulus, brought about by Lessening the Distance between the Responding and Stimulated Points experiment which I carried out with a frog's nerve in some- what subtonic condition. Here, when the stimulator was placed at some distance from the responding point, the response was the abnormal positive (fig. 278). When the effective intensity of transmitted stimulus was now slightly increased by moving the point of application a little nearer, the response became diphasic ; and finally, when the stimu- lator was placed still nearer, the response became normal negative. Thus with an identical specimen we may obtain at will either negative, diphasic, or positive response, by making changes only in the effective intensity of stimulus employed. We have also seen, moreover, that if we kept QUANTITATIVE STIMULATION OF NERVE 467 the stimulator at a certain distance from the responding point, such as at first to cause only positive response, succes- sive stimulations would then act to enhance conductivity gradually, and thus give rise to the appropriate changes, diphasic and negative in the response. The ultimate cause of these variations must therefore lie in the molecular condition of the tissue. Under varying cir- cumstances, this undergoes a cyclic change, the responsive reaction at any given moment constituting an indication of the particular molecular condition of the tissue. A more complete demonstration of this, carried out by an altogether different method, will be given in a subsequent chapter. My principal object in this chapter has been to prove the efficiency of the thermal shock as a mode of stimulation of nerve. Its wider applicability, in the case of other related investigations, will be treated in the two succeeding chapters. H II 2 CHAPTER XXXII ELECTRICAL RESPONSE OF ISOLATED VEGETAL NERVE Specialised conducting tissues— Isolated vegetal nerve — Method of obtaining electrical response in vegetal nerve — Similarity of responses of plant and animal nerve : {a) action of ether — (6) action of carbonic acid— (^■) action of vapour of alcohol— (<^ action of ammonia — (e) exhibition of three types of response, negative, diphasic and positive — (/) effects of tetanisation of normal and modified specimens — Effect of increasing stimulus on response of modified tissue. It has been shown in the previous chapter that the state of excitation is transmitted to a distance in vegetable tissues. It has also been proved that such transmission is not due to the propagation of hydrostatic disturbance but to that of protoplasmic changes, precisely as in the case of animal tissues. It is obvious, further, that such transmission will be the more perfect the less the interruption of protoplasmic con- tinuity. Hence tissues like stems and petioles, which contain fibro-vascular elements, are found to be good conductors of excitation, whereas indifferent tissues, such as those of leaves and tubers, are relatively feeble as regards this power, excitation in their case remaining somewhat localised. Even with regard to stems and petioles themselves, a contrast is found to exist in this respect between the fibro- vascular elements and the ground tissue. Thus, in the case of a petiole of cauliflower, I made two experimental prepara- tions. In the first, the ground tissue was cut away, leaving the fibro-vascular elements ; and in the second, a column of ground tissue was left outstanding, denuded of fibro-vascular elements. The former of these was found to transmit excitation to a certain distance, whereas in the latter the transmission was practically absent. In the case of a third kESPONSE OF ISOLATED VEGETAL NERVE 469 preparation I bifurcated the specimen, stripping away from one of the two limbs the fibro-vascular elements, and from the other most of the ground tissue. Galvanometric connec- tions were now made with the free ends of the fibro-vascular and ground tissues respectively, and stimulus was applied by means of transverse cut, or by application of a hot plate across the area of union. The transmitted effect was now perceived as galvanometric negativity, at the end of that strip which was composed of fibro-vascular elements. In studying this subject of conduction, I found the transmitted effect of excitation to be universally well ex- hibited in the petioles of ferns, successive re- sponses, obtained at a distance from the point of stimulation, being in their case singularly perfect and uniform. From this I was led to the conclusion that the disposition of the conductors must here be particularly well adapted to their purpose. Fig. 279. Frond of Fern with Conducting Nerves n exposed in Enlarged Figure to Right m I had long been desirous of isolating .whatever elements in the vegetable tissue were to be regarded as performing the function of nerves, and it appeared to me that I had here found a good subject for this investigation ; nd accordingly, on carefully breaking the hard casing of the petiole, and pulling it away in both directions, I was able to isolate the conducting fibro-vascular threads, which were long, soft, and white in colour, remarkably similar in their appearance to animal nerves (fig. 279). These threads vary in number with different species of ferns, and resemble 470 COMPARATIVE ELECTRO-PHYSIOLOGY possible to detach one of them having a length of 20 cm. or more. Now the essential feature of a nerve is its protoplasmic continuity, which is ensured by its fibrous structure. And in what I have called the vegetable nerve we find the same characteristic to hold good. On viewing this structure, as it appears on making a transverse section of the petiole, we find it enclosed within sheath-like sclerenchyma. It mainly consists in itself of a bundle of fine fibres with a few vessels in the centre. But however remarkable these external resemblances may seem, they are by no means so startling as the more fundamental similarities which are demonstrated so soon as we proceed to subject this vegetable structure to those tests of electrical response which are characteristic of animal nerve. It may be said that for the following investigation the nerves of the common maiden- hair fern {Adiantum) and Nephrodium inolle were found most suitable. In obtaining a plant nerve for purposes of experiment it is possible to dissect it out and at the same time to avoid injury. It is then placed in normal saline solution for about half an hour, so as to remove all traces of excitation due to handling. When the external temperature is not high, the excitability of the isolated plant nerve is found to remain relatively unaffected for a considerable period, but in the hot weather it undergoes rapid decline ; and the only way in which I could overcome this difficulty was by placing the specimen in normal saline solution which was ice-cold. The experimental precautions to be taken are precisely the same as those observed in corresponding experiments with animal nerve ; that is to say, the specimen should be placed in a moist chamber. For the process of drying is found to induce a transient increase of excitability followed by a permanent abolition of responsiveness, in the one case as in the other. In order to obtain responses, one end of the specimen may be killed by the local application of hot salt .solution. The galvanometric connections are then made, one with the killed, RESPONSE OF ISOLATED VEGETAL NERVE 47 1 and the other with the unkilled portions of the specimen higher up. In order to ensure that the electrical indication be a true responsive reaction, it is well to use a non-electrical form of stimulus. One of the most perfect forms — as we have seen in the previous chapter, on excitation of animal nerve— is the thermal, and this may be applied in precisely the same manner, that is to say, by means of a platinum wire, surrounding, but not necessarily in contact with, the given area of the specimen, this wire being heated periodically in the manner previously described, by means of a metro- nome closing an electric circuit. With a good specimen, a single thermal shock, lasting for less than a second, will be found sufficient to induce a considerable electrical response, or a response of still greater amplitude may be obtained by the summated effects of several such stimuli. One of the most noticeable differences between this plant nerve and other vegetable tissues lies in its greater excitability. For example, while a single thermal shock of less than one second's duration is sufficient, as has been said, to evoke immediate and considerable response from the isolated nerve, we find that, in order to ev^oke similar response from the petiole of the fern as a whole, it is necessary to submit it to the same stimulus some twenty times in succession, the response even after this taking place with relative sluggishness. A still further characteristic is its indefatigability. A long series of responses to uniform stimuli, such as would in the case of ordinary tissues bring about marked fatigue, will in that of nerve induce little or none. Rapidly succeeding tetanising shocks, moreover, such as in other tissues induce rapid decline, induce, generally speaking, but little of such an effect on the response of nerve. In the case of this vege- table nerve also the same statements hold good. A long continued series of responses shows little fatigue. After tetanisation, moreover, we find that the responses of nerve, whether animal or vegetable, become enhanced. In the matter of the effects induced by chemical re- agents on animal and vegetable nerves, a further remarkable 472 COMPARATIVE ELECTRO-PHYSIOLOGY parallelism is to be observed. The completeness of this may be seen in greater detail in the next chapter. I shall, at the present point, confine myself to giving a few typical cases. Ether, for example, when acting on animal nerve, induces a preliminary exaltation of excitability, which is followed under its long continued action by depression. On blowing off the ether vapour again the original state of excitability is restored. In fig. 280 are seen the similar effects of this reagent on vegetable nerve, where {a) exhibits the normal response, {b) the immediate exaltation due to ether, {c) the Fig. 280. Photographic Record of effect of Ether on the Electrical Response of Plant-nerve {a) Normal response : application of ether at point marked with t 5 {h) Enhanced response in first stage of action of ether ; [c) Subse- quent depression ; {d) Restoration of normal response after blowing-oft of ether. subsequent effect of depression, which becomes marked after continuous action during twenty-five minutes, and {d) the restoration of the original condition on the blowing-off of the ether. Carbonic acid is known, in the case of animal nerve, to have the effect, in the first stage, or in small quantities, of inducing exaltation, which passes under its prolonged action, or, in the case of a stronger application, into depression. A similar effect is seen in fig. 281, where {ii) shows the normal response of a vegetable nerve, and {b) the preliminary exalta- tion due to carbonic acid introduced into the vegetable nerve- RESPONSE OF ISOLATED VEGETAL NERVE 473 chamber. This is seen to increase continuously for some twenty minutes in (c). But after the expiration of half an hour depression makes its appearance (d). This becomes still more marked, after the fortieth minute, in (e\ -y^/^iimmii Fig. 281. Photographic Record of Effect of CO.^ on Electrical Response of Plant-nerve a, normal responses ; d and c, enhanced response during first stage of action ; d and e, subsequent growing depression. Alcohol vapour in strong, or long-continued applications, induces marked decline of response in animal nerve. Parallel effects are seen in the case of vegetable nerve in fig. 282. The effect of ammonia on animal tissues is character- istically different, according as the subject of experiment is 'AJWuuyu Fig. 282. Photographic Record of Abolition of Response by Strong Application of Alcohol nervous or ordinary tissue. While the excitability of the muscle, for example, is but little affected by its application, that of nerve is quickly abolished. In order to see v^hether the same characteristic difference would be exhibited, as 474 COMPARATIVE ELECTRO-PHYSIOLOGY between ordinary vegetable tissues and vegetable nerve, I first studied its efifcct on the ordinary tissue of the petiole of Fig. 283. Photographic Record of Eflfect of Ammonia on Ordinary Tissue of Petiole of Walnut Note that the effect of ammonia here is practically negligible. walnut It will be seen from fig. 283 that ammonia here induced practically no change in the excitability. But when the same reagent was applied to the isolated nerve of fern the response underwent depression, followed by total abolition, in the course of five minutes (fig. 284). One very curious charac- teristic of the electrical response of frog's nerve is the occurrence, as referred to in the last chapter, of three distinct types of re- sponses, according to its condition. Thus, as has already been said, while highly-excitable nerve ex- hibits the normal negative response, the same nerve, when it has become sub-tonic, will give a mixed or diphasic response ; and a nerve which is FiG. 284. Photographic Record of Effect of Similar Application of Ammonia on Plant-nerve The response here is rapidly diminished and finally abolished. RESPONSE OF ISOLATED VEGETAL NERVE 475 modified to a still greater extent will show a purely abnormal or positive electrical response. In the case of vegetable nerve, I find exactly the same three types of response repeated, under the same conditions. This will be seen in the three sets of records given in fig. 285. The normal responses, which are negative, are here represented as 'up,' while the abnormal positive is represented as ' down.' Still more remarkable is the parallelism observed between the effects of tetanisation, on animal and vegetable nerve, both normal and modified. In the case of fresh fros-'s nerve the responses are, as we have seen, enhanced, after uu hhhh ^^i^i. Fu;. 285. Photographic Record of Exhibition of Three Types of Response, Normal Negative, Diphasic, and Abnormal Positive, in Nerve of Fern under Difterent Conditions a period of tetanisation. The effect of tetanisation on vegetable nerve is precisely similar, as is seen in fig. 286. In the case of the modified frog's nerve, moreover, it is found that the abnormal positive response tends, after tetanisation, to become normal. This is seen in the abnormal response, whether positive or diphasic, being converted to the normal negative type. I have obtained exactly parallel effects in the case of modified vegetable nerve. In fig. 287 we see the abnormal diphasic response of vegetable nerve converted, after tetanisation, into normal negative. Thus, as in the response of animal nerve, so also in that of the vegetable, tetanisation is found to have the effect of enhancing the normal, or converting the abnormal into A76 COMPARATIVE ELECTRO-PHYSIOLOGY 'normal response. The abnormal response of nerve we found to be due to the joint depression of conductivity and excita- bility, on account of which the positive alone, instead of the true excitatory negative, was exhibited. In experimenting with frog's nerve we saw that abnormal response might, at will, be converted into normal through the intermediate diphasic by appropriately increasing the effective intensity of stimulation. A simple means of effecting this was to bring the stimulator gradually nearer the responding point. Fig. 286. Photographic Record of Effect of Telanisation in Inducing Enhancement of Normal Negative Response in Nerve of Fern The first series of responses seen to be enhanced after intervening tetani- sation, T. In the response of vegetable nerve effects exactly parallel are to be observed. With a given specimen of vegetable nerve, the stimulator had at first been placed at a distance of 2 cm. from the proximal galvanometric contact, and the responses then taken were found to be of the abnormal positive type. The stimulator was now brought nearer, the distance being reduced to i cm., and the next pair of responses is seen to be diphasic, consisting of a positive twitch followed by the RESPONSE OF ISOLATED VEGETAL NERVE 477 normal negative response. The distance was next reduced still further, namely, to "5 cm., with the result that the Fig. 287. Photographic Record of Conversion of the Abnormal Diphasic into Normal Negative, after Tetanisation, T, in Nerve of Fern Fig. 288. Photographic Record showing how the Abnormal Positive Response is converted through Diphasic into Normal Negative, by the Increasing Effective Intensity of Stimulus, due to Lessening the Distance between the Responding and Stimulated Points responses now became normal negative (fig. 288). It is thus seen that there is a continuity of response in the same I 478 COMPARATIVE ELECTRO-PHYSIOLOGY tissue, as between the abnormal and normal, through the intermediate diphasic. From the various experiments, then, which have been given in this chapter, it will be seen that the response of the isolated vegetable nerve is in every respect similar to the corresponding responses of animal nerve. And we shall also see how, by means of the study of this vegetable nerve, we are enabled to elucidate many obscurities in the responses of the corresponding animal tissue. We shall in the next chapter enter in detail into the question of the modifications induced in the conductivity and excitability of vegetable nerve under the action of various external agencies, and these will be found to exhibit the strictest parallel with corresponding variations induced in the animal. CHAPTER XXXIII THE CONDUCTIVITY BALANCE Receptivity, conductivity, and responsivity — Necessity for distinguishing these — Advantages of the Method of Balance— Simultaneous comparison of variations of receptivity, conductivity, and responsivity — The Conductivity Balance— Effect of Na^COg on frog's nerve — Effect of CuSO^ — Effect of chemical reagents on plant nerve — Effect of CaCL on responsivity — Responsivity variation under KCl — Comparison of simultaneous effects of NaCl and NaBr on responsivity — Effects of Na^COj in different dilutions on conductivity— Demonstration of two different elements in conductivity, velocity, and intensity — Conductivity versus responsivity — [a) effect of KI — {b) Effect of NaT — Effect of alcohol on receptivity, conductivity, and responsivity — Comparison of simultaneous effects of alcohol — [a) on receptivity vo'sus conductivity — (b) on receptivity versus responsivity. We know that when any point in a tissue is acted on by external stimulus, it receives the stimulation and is thrown into a state of excitation. This excitation is then conducted along the length of the tissue, and may be made outwardly manifest at some distant point by means of a suitable in- dication such as motile or galvanometric response. There are thus three different aspects of the excitatory effect to be distinguished from each other, namely, first the excita- tory effect at the point of reception of stimulus, which I have elsewhere designated receptive excitability, or simply Receptivity : secondly, the power of transmission of excita- tion, or Conductivity : and thirdly, the excitatory effect evolved at the distant responding region, which I shall henceforth term Responsivity. Though these three aspects of the excitatory reaction are all alike dependent upon the molecular derangement caused by stimulus, it is nevertheless important to consider them separately, since their variation is not always the same under the same circumstances. We I 480 COMPARATIVE ELECTRO-PHYSIOLOGY have seen, for example, that a rise of temperature, by in- creasing molecular mobility, enhances conductivity. But this increase of molecular mobility and internal energy also goes to augment the force of recovery, and, owing to this, the amplitude of excitatory response may be decreased. Thus, while a rise of temperature increases conductivity, it may appear to decrease responsive excitability. So much for the necessity of a distinction between conductivity and responsivity. The term * excitability ' is commonly used for receptivity and responsivity indifferently. But I shall show in the course of the present chapter that it is important to make a distinction between these, since the same external agent may effect the two differently.^ In the following investigation, receptive excitability, or receptivity, will be represented by R, conductivity by C, and responsive ex- citability, or responsivity, by E. In determining the effect of any external condition such as the application of a chemical reagent on responsive ex- citability, in the case of animal nerve, it is usual to take a series of normal responses, and then to record the modified responses after the application of the reagent. By com- paring a number of such series of records, representing the action of various reagents on different specimens, the relative effect of each chemical may be inferred. The drawback to this method lies, first, in the fact that by the addition of the chemical reagent the resistance of the electrical circuit undergoes an unknown change, thus inducing a variation in the amplitude of response, which is not necessarily due to the excitatory electromotive change per se. It is true that this difficulty may to a greater or less extent be obviated by interposing a high external resistance in the circuit, but this, by reducing the deflection, necessarily reduces the sensitive- ness of the method also. Different specimens again cannot but be characterised by slight individual peculiarities, and the experimental arrangements therefore can only be considered to be perfect when we are able to compare the effects of two ' See also Bose, Plant Response^ pp. 215-230. I THE CONDUCTIVITY BALANCE 48 1 agents on an identical specimen. Again, in a series of chemical compounds which differ but slightly in effect from one another, an arrangement has to be devised by which the most minute excitatory variations will be conspicuously displayed. The same delicacy of experimental adjustment also becomes necessary when we wish to investigate the varying effects of time and quantity in the application. Similar considerations are involved when we attempt to observe the effects of various agents on conductivity and receptivity ; and still more complicated are the difficulties to be overcome when we have to study the property of con- ductivity versus responsivity or receptivity, or of receptivity versus responsivity, under the action of the same external agent. The methods hitherto available are neither perfect nor delicate enough for a complete and satisfactory determina- tion by their means of the various problems which arise in this connection. I shall now, however, describe a very perfect and delicate method carried out by an experimental arrangement which I have devised, and shall designate as the Conductivity-Balance, by which the variation of an affected region may be continuously compared with a normal area as regards each of the three different aspects of the ex- citatory reaction, namely, receptivity, conductivity, and responsivity. In this method, moreover, the result is un- affected by any variation of resistance in the circuit that may be induced by changed conditions. It also enables us to solve the various difficulties encountered in comparing the relative changes induced in conductivity with those induced in receptivity or responsivity, or in the two last in respect to each other, under the influence of a given reagent. In fig. 289 is given a diagrammatic representation of the principal parts of this Conductivity- Balance. The thermal stimulator produces stimulation of the enclosed area of the specimen. The excitatory wave travels along both arms of the balance, through the conducting region C and C', and induces excitatory electromotive effects at the two responsive points E and E'. The excitatory electrical effects at E and E' I I 482 COMPARATIVE ELECTRO-PHYSIOLOGY are opposed, and when these are equal, and balance each other, the galvanometer indication is then reduced to zero. E and E' are usually at a distance of about 4 cm. from each other. When the stimulator is brought too near to the left contact E', the excitatory effect of galvanometric negativity which is induced there is relatively greater than at E. The balance is thus dis- turbed, and the resultant responsive deflection is then, say, downwards. When the stimulator is placed, on the other hand, too near the contact E, to the right, the resultant galvano- metric deflection will be up.^ By suitable movement of the stimulator, to and fro between these two ex- tremes, a point may be found where the excitatory effects at E and E' will exactly balance each other. I give here (fig. 290) a record taken Fig. 290, Photographic Re- cord made during Pre- liminary Adjustment for Balance of Nerve of Fern The first two down -responses show over- balance, when S is too near the left, e' being relatively more excited. The up-responses indicate over - balance caused by S being too much to the right. The horizontal record shows attainment of exact balance. s, Fig. 289. Diagrammatic Representa- tion of the Conductivity Balance thermal stimulator ; C and c', the conducting arms of the balance ; E and e', responding points. Dif- ferential excitatory electrical effects at E and e' recorded by galvano- meter, G. during this preliminary stage of adjustment. The first two down-responses were obtained when the stimulator was too far away from the balancing-point to the left. The next two * It is to be understood that what is said here refers to nerve in a normal condition of conductivity. THE COXDUCTIVITV BALANCE 483 ui>-responses were obtained when it was contrariwise too far to the right More careful adjustment reduced this up- movement, as seen in the next two responses, and finally^ when the exact balandng-potnt was reached, tiie effect was null, as seen in the horizontal record. In studpng the question of the variation of responsive excitability- induced by any given reagent, the agent is applied at the point E to the right Any variation of excit- ability win then upset the balance. If die reagent be of a stimulator\' character we shall obtain a resultant up-response, but if it be of a depressii^ natore, E will be rendered rda- tively the less excitable of die two points, and die response will consequently be down. It will thus- be seen that diat upsetting of the balance by whidi eidierop- or down-re^iooses are induced is due sim{^ to die relatively excitatofy or depres^g effect of the reagent, and is completdy inde- pendent of any variation of resistance ndiich might be broug h t about by its application. In die coarse of die foDowing investigation, it is to be miderstood that die dec- trical GonnectioDS are so made that the greater excitation of the r^^t-hand contact is always represented by up-reqionse, and vice versd. If it be desired to make a comparison between die exdtatofy reactions of two reagents, dien the two are applied simultaneoosly, one at E and the other at E'. The resoltii^ record dien afibrds as a coudmi oas graphic iUnstration of tibe rdathreand vaiyii^ effects of the two. If, again, it is the infinence of any agent on ooodoctivity that is to be studied, we first take a balanced leoord and then apply the given reagent on an area of about i cm. at C on tfaeooiMliicliii^arm. In this case, the r esponsi v e exdtabilities of the two points E and E' are die same, but if the effect of the agent have been to induce increased conductivity of C, dien tlie excitation transmittBd to die right-hand side, E, will be greater, and the response caused by the upsrtt t i^ of the balance will be upwards. Conv e r s e ly , a ckywn-response will indicate that the efiect of the a^^t las been to depress the conductivity. Again, we can oom p are die rdative effects in 112 484 COxMPARATIVE ELECTRO-PHYSIOLOGY conductivity-variation induced by two different agents which are applied simultaneously, one on the arm C and the other on C'. It is possible again to compare the variation of con- ductivity with that of responsivity, by applying one agent at a responding region, say E, and the other on the opposite arm of the balance at c'. The mode of investigation of receptivity changes will be described presently. In fig. 291 we have the complete apparatus. The animal or vegetal nerve, N N, rests on non-polarisable electrodes of Fig. 291. Complete Apparatus of Conductivity Balance The nerve N supported on electrodes Eg, E3. The two other electrodes E, E4 are not used in this experiment, but are employed for experiments on electrotonus ; T, thermal stimulator, the relative lengths of the arms of the balance being adjusted by the slide s. a U -shape. For the present experiments, two electrodes, E^ and E3, are sufficient, their mutual distance being capable of any variation by movement along a sliding-bar. The same apparatus might be used for experiments on electrotonus, in which two additional electrodes would be required. The position of the electrothermic stimulator T is capable of very careful adjustment for purposes of balance, by means of the sliding-rod S. A glass cover, not shown in the figure, fits into the groove which is represented by a double dotted line sur- rounding the apparatus, and thus enables the chamber con- taining the nerve to be kept in a properly humid condition. THE CONDUCTIVITY BALANCE 48s In all these experiments by balance, it is to be borne in mind that adjustment is always made for perfect balance at the beginning of the record, and represented by a short, more or less horizontal, line. In order to show the typical effects of induced variations of excitability, in upsetting the balance, I shall first give records of experiments carried out on the nerve of frog. Dilute sodium carbonate is known to be an agent which enhances excitability. A long-continued application, or the application of a stronger dose, may, however, bring about a depression. When a dilute solution of NaaCOg was ap- FiG. 292. Eft'ect of Na^COg Solution on Responsive Excitability of Frog's Nerve In this and following records the hori- zontal line at the beginning indicates exact balance. The upsetting of the balance in the up-direction repre- sents either the enhanced respon- sivity of the right-hand responding point E, or the increased con- ductivity of the right-hand arm c. Down -curves represent correspond- ing absolute or relative depressions. NaXOg applied to e is seen to exalt the responsivity of that point. Fig. 293. Effect of CUSO4 on Frog's Nerve The down record shows depression of excitability. plied at the responsive point E on the right side, the up- setting of the balance upwards immediately indicated the greater excitability induced by the reagent. The long- continued action of this reagent, however, showed that the enhanced excitability was undergoing a gradual decline (fig. 292). In order to exhibit the characteristic upset caused by a depressing agent, I employed on another .specimen a toxic solution of copper sulphate, applying it at E on the right. The previous state of equilibrium is seen by the horizontal line at the beginning of the record, and the -4^ COMPARATIVE ELECTRO-PHYSIOLOGY subsequent depression of excitability at E is shown by the upsetting of the balance downwards (fig. 293). I shall next take up the determination of the changes induced by chemical agents on the excitability of plant nerve and shall begin by describing the different effects which occur on the application of ca/dum and potassium salts. For this purpose, deci-molecular solutions were employed. Fig. 294 shows the effect of CaClj on vegetable nerve, the solution being applied at E on the right-hand side. It will be noticed that this caused an upset of the balance, showing an increase of excitability that becomes considerable after the expiration of five minutes. In the case of KCl, however, this effect was re- versed, that is to say, a de- pression was induced. This is seen in fig. 295, where the balanced record gives way, first to diphasic, and after- wards to a down-response, indicating an effect of de- pression at E. These two of the basic moiety in in- FiG. 294. Photographic Record show- ing Enhancement of Responsivity by Application of CaCl^ CaClj applied to E is seen to exalt the responsivity of that point. experiments show thfe effect ducing changes of responsive excitability. I shall next describe experiments by which the simul- taneous effects of two different reagents on the responsivity of a given tissue may be compared. For this purpose, one agent is applied at one end of the balance E, the other being administered at e'. In the case of animal nerve, it was shown by Griitzner, that both NaCl and NaBr induce ex- citatory effects, that induced by NaBr being relatively the greater. But the continued action of either of these reagents THE CONDUCTIVITY BALANCE 487 induces depression, which sets in earlier in the case of NaBr. The effect of these two reagents on vegetable nerve is pre- Photographic Record showing Depression of Responsive Excitability by Application of KCl cisely the same, as will be seen from an inspection of the record given in fig. 296. The NaBr was applied on the Fig. 296. Photographic Record exhibiting Comparative Effects of NaCl and NaBr on Responsivity NaCl was applied on e' and NaBr on E, the formula being E'NaCiENaBr- The record shows the greater and earlier effect of NaBr at E in causing relative excitation followed by relative depression. right-hand side E, and NaCl on the left-hand E', a process which is expressed, for the sake of brevity, by the formula 'E'NaciI^jTaBr- Thc greater and earlier excitatory effect of 488 COMPARATIVE ELECTRO-PHYSIOLOGY NaBr, applied on the right-hand side, is shown by the resultant up-responses. But after a time, E being now depressed by the continued action of NaBr, the effect of NaCl, applied on the left, becomes relatively predominant, a fact demonstrated by the upset of the balance in the oppo- site direction, with concomitant down-responses. We shall next take up the subject of variations induced in conductivity. We have seen that dilute solutions of Na^COa have the effect of exalting responsive excitability. Fig. 297. Photographic Record of Effect of Dilute ( -5 per cent. ) Solution of Na2COj on Variation of Conductivity Reagent applied on right arm c. Record shows immediate enhancement of conductivity giving rise to up-curves, followed by depression, seen in down-curves. Note the appearance of a down-twitch at the be- ginning of the sixth response due to the later arrival of excitation at E. Note further the replacement of up- by increasing down-responses. Long-continued applications, or strong solutions, however, have the effect of inducing a depression. Similarly, I find that this reagent has the effect of enhancing conductivity, provided the solution is sufficiently dilute. In the case of the petioles of ferns, a 2 per cent, solution was found to induce a preliminary exaltation of excitability, followed by a depression (p. 136). In dealing with the conductivity- variation in certain isolated vegetable nerves, however, a 2 per cent, solution was found to induce a depression of con- ductivity, but a -5 per cent, solution caused an enhancement THE CONDUCTIVITY BALANCE 489 of conductivity, followed, after long-continued action, by- depression. These facts are illustrated in an extremely interesting manner in the records given in figs. 297 and 298. In both these cases the solution was applied on the right arm of the balance at C, the difference being only that in the first experi- ment the strength of solution was '5, and in the second 2 per cent. An inspection of fig. 297 shows that the application of the first induced a great and immediate enhancement of conductivity, causing resultant up-responses, which were par- ticularly marked during the first four minutes. This increased Fig. 298. Photographic Record of Effect of Stronger Dose (2 per cent.) of NagCOj Solution on Conductivity. The solution was applied on the right arm of the balance c. Note grow- ing depression and appearance of diphasic effect. conductivity is then seen to undergo a continuous decrease and reversal into growing depression, as seen in the substi- tution of increasing down-responses. This record deserves special attention, inasmuch as it affords us an insight into a phenomenon which could not otherwise have been suspected. Greater conductivity is usually associated with increased velocity of transmission. It would appear, however, that the term conductivity really covers two different phenomena which may not always be concomitant. That is to say, an increase of conductivity may mean either a greater speed of transmission of excitation or a greater intensity of the I 49<>. COMPARATIVE ELECTRO-PHYSIOLOGY excitation transmitted. In the first four records of the present series the induced enhancement of conductivity is shown by the occurrence of up-responses only. The fifth record, how- ever, shows a marked preliminary twitch in the negative direction, followed by an up-response of some amplitude. This shows that the excitatory effect reached the right end E. later than the left, though the intensity still remained greater. The continued action of the reagent subsequently reduced the intensity also, so that this diphasic ultimately became converted into a purely monophasic down-response, gradually increasing to a maximum. In fig. 298 we observe the depression of conductivity by a stronger dose of 2 per cent, solution of NagCOg, applied on the right-hand side at C. Here, again, we can see the separated effects of the two elements of conductivity — that is to say, the intensity of the effect transmitted and the speed of transmission. In the first few responses of this series we see the diminished intensity of transmission to the right giving rise to resultant responses which are entirely downwards. Later, this transmission of enfeebled excitation becomes delayed also, and by the phase- difference thus induced we obtain the growing diphasic effects which have already been fully explained on p. 144, fig. 100. Owing now to this growing difference of phase, the two opposed effects no longer neutralise each other to the same extent as before, and we obtain increasing amplitude of both the constituent phases. The down-curve in the diphasic response represents the earlier arrival, and relatively greater intensity, of effect at the left contact E'. And the up-curve shows the later arrival of the less intense effect at the right- hand contact E. It is thus clearly seen that conductivity includes two different elements of speed and intensity which may not in all cases be coincident. I shall next describe experiments which will demonstrate the variation of conductivity versus that of responsive excit- ability under the action of the same reagent. In animal nerve responsive excitability is diminished by the action of strong solutions of neutral salts, and potassium salts induce greater THE CONDUCTIVITY BALANCE 491 depression than corresponding sodium salts. But neutral salts, generally speaking, affect conductivity to a much slighter extent than responsivity. There is, however, a very curious exception to this rule in the case. of animal nerve, where 6'i per cent, of Nal is found to affect the conductivity to a much greater extent than the responsive excitability. I find a remarkable parallelism to these effects in the case of vegetable nerve, which is capable of striking demonstration by the comparative method of simultaneous variations of conductivity and excitability already de- scribed. In order to demonstrate these con- trasted effects of KI and Nal on conductivity and excitability, I shall here give an account of two different experi- ments. In the first, after obtaining the pre- liminary balance, KI was applied at c, on the right arm, the same reagent being also ap- plied at the end E' of the left arm, this pro- cess being represented by the formula E'^jC^i. The record seen in fig. 299 shows, by its resultant up-responses, that a greater depression of responsivity at e' than of conductivity at c has been induced- In the next experiment (fig. 300) Nal was applied instead of KI, on C to the right, and E' to the left, the formula thus being E'^.^,Cp,^i The resultant responses were now down- wards, showing that there was a relatively greater depression Fig. 299. Responsivity versus Conductivity under KI This photographic record shows the effect of KI on responsivity and conductivity when reagent applied at e' and C simultaneously. The formula is E'kiCki- Record shows greater depression of responsivity than of conductivity. 492 COMPARATIVE ELECTRO-PHYSIOLOGY at the responding point E. of conductivity than of responsivity. In respect of conduc- tivity and responsivity, therefore, the effects of these two drugs, KI and Nal, are seen to be opposite. In order to observe the effect of alcohol on nervous tissue, by means of the conductivity balance, I first experimented on the nerve of frog. A 5 per cent, solution was applied This is seen (fig. 301) to induce a depression of responsivity. A more dilute solution generally induces a preliminary exaltation followed by depression. We said in the previous chapter that when alcohol vapour was passed into the chamber of the vegetable nerve the responses underwent a rapid abolition. This result, however, Fig. 3CX). Responsivity versus Conductivity under Nal The formula in this case is E'NaiCNai* Photographic record shows an effect opposite to that of KI as previously described, there being now a relatively greater de- pression of conductivity than of responsivity. Fig. 301. Effect of Alcohol on the Responsivity of Frog's Nerve Upsetting of the balance in the downward direction shows depression. was due to the joint action of the variations of receptivity, conductivity, and responsivity, some of which may possibly have been in the positive and others in the negative direction. In order to determine the effect of each of these we must, then, perform separate experiments. Such a deter- mination I have made, using the method of the so-called * negative variation,' in which the proximal galvanometric THE CONDUCTIVITY BALANCE 495 contact was on an unkilled, and the distal on a killed area. The first of these experiments was on variation of receptivity. The thermal stimulator was provided with mica shields, so that the receptive area was strictly circumscribed at the centre of the thermal platinum loop. Normal responses were first taken ; the receptive area was next touched with I per cent, solution of alcohol, and the modified responses were recorded. The results are seen in fig. 302, which gives Fig. 302. Photographic Record of Effect of Alcohol Vapour on Receptivity The three normal responses to the left are seen to be exalted after applica- tion of ether on the receptive point. a striking demonstration of the increased receptivity induced by dilute alcohol. The effect on conductivity, however, is in curious con- trast to this. On applying i per cent, solution in the conducting region between the stimulator and the proximal contact, a very great diminution of the conducting power is observed, as seen in fig. 303. It may be stated here that a similar enhancement of receptive excitability, and depression of conductivity, are found to be the result of the action of alcohol in animal nerve also. In the next ex- periment, it is the variation of responsivity under the action of dilute alcohol which is tested. After taking the normal I 494 COMPARATIVE ELECTRO-PHYSIOLOGY responses as usual, a i per cent, solution of alcohol was applied at the proximal contact. It will be seen from the record in fig. 304 that the immediate effect was a depression Fig. 303. Photographic Record of Effect of Alcohol on Conductivity The three large responses to the left show the normal effect of transmitted excitation. Responses almost abolished, as seen on the right, by depression of conductivity. of the amplitude of response. This subsequently becomes converted into a diphasic response, consisting of a preliminary positive followed by the normal negative ; and finally the f^#,,/yfim Fig. 304. Photographic Record showing Effect of Alcohol on Responsivity a, normal responses, depressed, after application of alcohol, to d ; and converted later to abnormal positive responses c. response was totally reversed to positive, by the abolition of the true excitatory effect. It is thus seen that while dilute alcohol exalts the recep- tive excitability, it induces a depression of both conductivity THE CONDUCTIVITY BALANCE 495 and responsivity. I shall now describe further experiments by which the relative effects of alcohol are compared, as between conductivity and receptivity, and as between recep- tivity and responsivity. For the purposes of such a comparison, a new balancing arrangement has to be employed (fig. 305). Here, two electro-thermic stimulators are in series, so that ex- citations may be produced at two different points simultaneously. The gal- vanometer contacts E' and Fig. 305. Diagrammatic Repre- sentation of Experimental Ar- rangement for Demonstration of Receptivity versus Con- ductivity, or of Receptivity versus Responsivity S and s' are exciting thermal loops in series ; R and r', the enclosed receptive points ; c and c', con- ducting arms ; E and e', the responsive points. Fig. 306. Receptivity versus Respon- sivity under Alcohol Alcohol was applied at the receptive point to the left r', and the responsive point to the right E. The formula was R'aic.Eaic. The photographic record shows the relative enhancement of receptivity. E are made with two points intermediate between the stimu- lators. The distance of one of the two stimulators is kept constant, at, say, 2 cm. to the left of E', while the other is moved nearer to, or further from, E, until a balance is obtained. A I per cent, solution of alcohol is then applied to the left receptive point, R', and the right conducting area, C, the formula now being R'aicQic.. The fact that the receptive excitability is heightened by this reagent, and conductivity depressed, receives independent confirmation from the upset of the balance, giving rise to a downward response. I 496 COMPARATIVE ELECTRO-PHYSIOLOGY The next experiment consists of a comparison of the simultaneous variations of receptivity and responsivity. Alcohol is applied at R' and E, the formula thus being R'aic.Eaic.. And we find here in confirmation of our previous results that, on account of the opposite effects of this agent on the receptive and responsive excitabilities, the resultant response is downwards (fig. 306), showing that the receptivity has been relatively exalted. Thus the experi- ments which I have here described show that the same agent may have different effects on receptive and responsive excitability, and thus accentuate the necessity of clearly distinguishing between the two. ^M' EFI CHAPTER XXXIV EFFECT OF TEMPERATURE AND AFTER-EFFECTS OF STIMULUS ON CONDUCTIVITY Effect of temperature in inducing variations of conductivity : {a) by Method of Mechanical Response ; {d) by Method of Electric Balance^Effect of cold — Effect of rising temperature — The Thermal Cell — After-effect of stimulation on conductivity — The Avalanche Theory — Determination of the after-effect of stimulus on conductivity by the Electrical Balance-^ After-effect of moder- ate stimulation — After-effect of excessive stimulation. In studying the effect of temperature in inducing variations of conductivity, we may use either of two different methods — in the first place the method of mechanical, or in the second that of electrical response. For the first of these it is neces- sary to have what is generally known as a ' sensitive ' plant, the leaves or leaflets of which afford conspicuous motile indications of the arrival of the excitatory wave from a distance. In such a case the time-interval between the application of stimulus and the response of a leaflet at a known distance gives us a measure of the velocity of con- duction ; and if we carry out successive experiments at different temperatures we have a means of determining the effect of temperature on conductivity. Employing this method, I have elsewhere given a determination of the effect of temperature on the velocity of transmission in Biophytu7n sensitivum. It was there shown that lower- ing of temperature reduced the velocity of transrhission even to the extent of abolition, when the cooling was suf- ficiently intense. With moderate cooling the velocity was found to be decreased to about one-third. The effect of rise of temperature was, on the contrary, an increase of K K 498 COMPARATIVE ELECTRO-PHYSIOLOGY velocity. When it rose from 30° C. to 35° C, for example, the velocity was doubled. By employing the electrical method of response, however, we are rendered independent of the use of sensitive plants, and by means of the Conductivity Balance we are enabled to demonstrate the slightest variation of conductivity, as between the left arm of the balance, which is maintained at standard temperature, and the right, which is subjected to the given change. Thus in a definite experiment on a nerve of fern the temperature of the room was 30° C. After first obtaining the balanced record, the temperature of a portion of the right arm of the balance was lowered. This one-sided cooling was effected by supporting the right arm of the nerve, through a certain length, in the concavity of a U-tube through which cold water at 15° C. was passed. Stimuli were now applied at intervals of one minute. Previously, as will be understood, such stimuli, owing to balance, had induced no resultant effect. But now, on account of the depression of conductivity on the right side, brought about by cooling, the balance was disturbed, and the resultant down-response seen in fig. 307 shows the diminished con- ductivity of the right arm. On the cessation of the flow of cold water the balance was gradually restored, in concomit- ance with the return to the original temperature. I next investigated the results of a rise of temperature, and here I specially desired to observe the conductivity variations, not at any one degree, but throughout a graduated and con- tinuous rise. I was confronted at the outset of this investi- gation by the difficulty arising from the fact that there was no convenient and satisfactory means for the local variation of the temperature of the nerve, in definite and known degrees. In connection with this there was also the further difficulty that a sudden variation of temperature will, in itself, act as a stimulus. Hence, in studying the effects of temperature per se, it is essential that there should be no such sudden variation. These difficulties were overcome by EFFECT OF TEMPERATURE ON CONDUCTIVITY 499 the employment of an electrical arrangement to bring about the graduated and continuous rise of temperature. A certain length of the vegetable nerve on the right arm of the Conductivity Balance was thus raised continuously in temperature, and its conductivity compared with that of the left arm of the balance, the latter being maintained at the temperature of the room, which happened at the time to be 33° C. The device by means of which this was accomplished Fig. 307. Photographic Record showing Effect of Cooling on Con- ductivity of Plant-nerve Balance v^^as obtained at starting, when temperature of both arms was 30° C. On cold being applied on right arm, the balance was dis- turbed, showing diminished conductivity on that side. On restoration of normal temperature, the balance is seen at the end of the record to be again restored. will be understood from fig. 308. A piece of cork has a small chamber cut into it measuring i cm. each way. In this is placed moist blotting-paper, which keeps it damp, and across it passes a length of i cm. of the right arm of the vegetable nerve in the Conductivity Balance. This cork- chamber has inlet and outlet tubes t and t\ The first of these contains a spiral, H, of platinum, which can be heated to a suitable degree by means of an electrical current, the 500 COMPARATIVE ELECTRO-PHYSIOLOGY intensity of which is capable of careful adjustment. The cork chamber is closed with a cover, through which passes a thermometer, T, for the indication of the temperature within. The tube /' is connected with an aspirator, and air is thus sucked in by /, and, passing through the platinum spiral, is warmed, and raises the temperature of the nerve in the chamber. This rise of tem^ perature is adjusted (i) by regu- lating the electrical current which heats the spiral, and (2) by con- trolling the inflow of air. As regards the first of these two processes, the electrical heating- circuit has a carbon rheostat interposed, by which the rate of rise of temperature may be regulated. The movement of the current of air, on the other hand, is controlled by adjusting the stopcock of the aspirator. By the joint manipulation of both these the rate of rise of temperature inside the chamber may be made perfectly uniform, and in my experiments this rate was approxi- mately 1° C. per minute. As already said, I selected a piece of vegetable nerve and took a balanced record. After this the temperature of the thermal cell > on the right-hand side was raised continuously, the response-record being taken at each degree of the rise, till a temperature of 50° C. had beenat tained. From the record given in fig. 309 it will be seen that the conductivity was always greater at temperatures up to 47° C. than it was on the left-hand side, which was all the time maintained at the constant temperature of 33° C. At 48° C, Fig. 308. The Cork Chamber for Gradual Raising of the Temperature of one Arm of the Balance A and B, the two halves of the chamber ; T, thermometer ; / and /', inlet and outlet tubes ; H, thermal coil for heating. EFFECT OF TEMPERATURE ON CONDUCTIVITY $01 however, the reversal of response showed that the conduc- tivity was now being depressed. And at still higher tem- peratures it was found to undergo a very great depression, as is seen by the abrupt downward movement of the curve. It is thus seen that, by means of the Method of Balance, this very difficult problem of the variation of conductivity under variation of temperature is made^capable of exact study. I shall next describe the results of an investigation into the after-effects of stimulus on conductivity and excitability, >viLaM Fig. 309. Photographic Record Showing Effect of Rising Temperature on Conductivity Balance obtained at starting at 33° C. Successive responses recorded at each degree C. of rise of temperature. Record shows increasing conductivity up to 47'^ C. A depression of conductivity is seen by reversal of curve to set in at 48° C. , and this becomes extremely pro- nounced at 50° C. a subject of much difficulty and of considerable theoretical importance. It has been found in Animal Physiology that the sciatic nerve of a frog, for instance, is not equally excitable throughout its length. When such a nerve, with its attached terminal muscle, is cut off from the spinal cord, it is seen to be more excitable the further from the muscle is the point on the nerve that is subjected to stimulation. From this fact that excitation increased with the distance of the point excited from the motor organ, Pfliiger was led to 502 COMPARATIVE ELECTRO-PHYSIOLOGY the ' Avalanche Theory,' namely, that during the passage of excitation down the nerve it actually gathers strength. But it is clear that this cannot be true, since we have seen that, other things being equal, excitation is always greater the nearer the point of stimulation to the responding region, and on this fact have depended all those experiments already described, which involved a delicate balance of equal excitations. It follows that the observed enhancement of excitability of a point on the nerve which is distal from the muscle, and in the neighbour- hood of a section, must be ascribed to some other cause. In reference to this Heidenhain, indeed, explained the greater excitability of higher tracts of divided nerve by the proximity of the artificial section. For the lower end of the nerve at once exhibits the same marked activity as the upper end if a section be made lower down. Excitability is, in fact, raised near the section, wherever the section may be. The distance travelled by the excitation could not, therefore, be the determining factor in the magnitude of effect. For so far from increasing it, this, as a matter of fact, causes diminution. It is to be remembered that though the excitability is increased near the point of section, yet at the section itself it is almost abolished, otherwise there could not have been any response by so-called negative variation. The question now arises. Why should the excitability be raised near the point of section ? It has been supposed that this was due to certain electri- cal changes induced by section, which in turn gave rise to electro-tonic variations of excitability. We shall see, in Chapter XL, that the passage of an electrical current through a living tissue induces changes of excitability. And this phenomenon is known as the electro-tonic effect. Now any ' injury,' such as a mechanical or thermal section, is known to induce galvanometric negativity, or anodic change, at or near the point of section. But it is the kathode-effect which is excitatory. And the observed greater excitability of the nerve near a point of section is supposed to be due to kat- electrotonus, produced within a certain tract from the cross- AFTER-EFFECT OF STIMULUS ON CONDUCTIVITY 503 section by internal short-circuiting of the nerve-current. That this explanation, however, does not meet all the requirements of the case will appear from certain experi- ments which I shall describe, where, under exactly similar electrotonic changes due to section, a result directly the opposite of this, that is to say, of depression, is seen to be induced. All these various facts will be found fully reconcilable, however, on the basis of a proposition which I shall establish, namely that in a nerve ^ moderate stimulation enhances ex- citability and conductivity y while excessive stimulation has the opposite effect of bringing about the depression of both. It is indeed natural to expect that while moderate stimulation, by increasing molecular mobility, would bring about one effect, excessive stimulus, by inducing overstrain, would result in exactly the opposite. Before proceeding to give an experi- mental demonstration of this hypothesis, we shall first consider the explanation which it affords of the peculiar excitatory changes observed in the case of cross-sectioned nerve. In the first place we know that a cut acts as. a stimulus. And since we found that the effect of stimulus decreases with the distance from the point of stimulation, it would appear that at the section itself the stimulation would be excessive ; moderately strong at a certain distance from it ; and practically negligible when very far away. In complete accordance with this is the resulting increase of excitability which has been observed near the point of section, while at the point itself the nerve is relatively inexcitable. The fact that stimulation, when not excessive, increases the conductivity and excitability, we found illustrated in the staircase increase of electrical response, and in the enhance- ment of amplitude after tetanisation, in vegetable and animal nerves (figs. 275 and 286). The same fact will be demonstrated later by means of the mechanical response of nerve. I shall now describe certain experiments which demonstrate it once more in a new and interesting manner. 504 COMPARATIVE ELECTRO-PHYSIOLOGY A vegetable nerve was adjusted for balance, with the ends projecting some distance beyond the electrodes. In order to show that the effect of injury is due to stimulus as such, and not to any particular form of it, I now made a thermal instead of mechanical section, by applying salt solution heated to about 60''. C. in the region A, at a distance of I cm. to the right of E (fig. 310). The effect of this stimulation was to induce a moderate excitation of the right arm of the balance, relatively to the left. If this moderate stimulation were to induce any increase of excitability and conductivity, that fact would be demonstrated by OJU^ Fig. 310. Experimental Arrange- ment for Studying After-effect of Stimulus on Conductivity and Excitability The stimulator adjusted to obtain balance between E and e'. Stimulus of moderate or strong intensity is applied to a point on the right of E. Upsetting of the balance in an upward direction shows an en- hancement, and in a downward direction, depression, of con- ductivity and excitability. Fig. 311. Photographic Record Showing Effect of Moderate Stimulation in Enhancing Conductivity and Excita- bility otted line at beginning shows the resting-current, as a per- sistent effect of stimulation. The upsetting of the balance upwards constitutes a positive variation of the resting- current, and indicates en- hanced conductivity and excitability. the upsetting of the balance, the resultant response being upwards. That this is what actually occurs will be seen from the records in fig. 311. It will be noticed that in consequence of stimulation to the right of E, that point became, more or less permanently, galvanometrically negative. This is represented by the dotted line upwards at the beginning of the record. It must be remembered that before the application of the thermal section, the right and left hand excitations, proceeding from the electro-thermically stimu- AFTER-EFFECT OF STIMULUS ON CONDUCTIVITY $0$ lated point in the middle, were exactly equal and balanced. The fact that after this application, however, there are resultant responses which are upwards, shows, as already- said, that by the moderate stimulation of the right-hand side, both excitability and conductivity have been increased. The resultant upward response here is, then, in the same direc- tion as the so-called * current of injury,' and forms, as it were, a positive variation of it. In another experiment, in which I wished to try the effect of excessive stimulation, instead of applying a hot solution at 60° C, I produced greater injury and consequent excessive stimulation, by scorch- ing the nerve at the same point as before, with a red-hot platinum wire. In this case resultant response was downwards, show- ing that the excitability and conductivity of the right-hand side of the balance had been depressed by over-stimulation. I was next desirous of demonstrating that the excita- bility of the over-stimulated or excited point undergoes depres- sion. For this purpose I took a fresh specimen and first ob- tained a state of balance. Similar produced a balanced or null effect, injured by touching it with a hot platinum wire. On now proceeding to take records, it is seen that the responses were downwards, showing the depression of excitability at the injured E (fig. 312). The fact that galvanometric negativity had been induced ^K at E, by reason of injury, is demonstrated at the beginning ^Kof the record as an up-line. The subsequent resultant Fig. 312. Photographic Record showing Eflfect of Excessive Stimulation in Depressing Ex- citability and Conductivity Up-line at starting shows the rest- ing-current due to after-effect of stimulation. The upsetting of balance in a downward direction constitutes a negative variation of the resting-current, and shows depression of conductivity and excitability. excitation of e' and E The point E was then 506 COMPARATIVE ELECTRO-PHYSIOLOGY seen to be a negative variation of the resting-current due to injury. It is thus seen that while simultaneous excita- tions of two normally excitable points E and e' are prevented by balance from giving rise to any response, the excitatory response becomes manifest when the balance is disturbed by the injury of either point of galvanometric contact ; and that, under these circumstances, the response is a negative variation of the current of injury. This experiment is important as giving a theoretical insight into the so-called response by negative variation. It also shows how limited is the applicability of the assumption that response is always by negative variation. For, in the similar experiment, previously described, under moderate injury, the response was by positive variation of the resting-current. It is further seen from these experiments that the enhancement of excitability, under the stimulation due to moderate injury, could not be caused by the suggested electrotonic effect. For the same anodic change induced by injury at E causes, in the case of moderate injury, an enhancement, and under greater injury a depression, of excitability. It is thus clear that the modifying influence is the effective intensity of stimulation. This fact, that moderate stimulation enhances, and excessive stimulation depresses excitability, will be further demonstrated in a future chapter, by the independent method in which the effects of electrotonus are completely eliminated. CHAPTER XXXV MECHANICAL RESPONSE OF NERVE Current assumption of non-motility of nerve— Shortcomings of galvanometric modes of detecting excitation — Mechanical response to continuous electric shocks — Optical Kunchangraph — Effect of ammonia on the mechanical response of nerve— Effect of morphia— Action of alcohol— Of chloroform- Abnormal positive or expansive response converted into normal contractile through diphasic, after tetanisation— Similar effects in mechanical response of vegetable nerve— Mechanical response due to transmitted effects of stimulation — Determination of velocity of transmission — Indeterminateness of velocity in isolated nerve — Kunchangraphic records on smoked glass — Oscillating recorder — Mechanical response of afferent nerve — Record of mechanical response of nerve due to transmitted stimulation, in gecko— Fatigue of conductivity — Conversion of normal contractile response into abnormal expansive, through diphasic, due to fatigue. I HAVE already referred to the distinctions which are com- monly insisted on, as between the reactions of different animal tissues. Certain of these are regarded as motile and others as non- motile. From an evolutionary point of view, however, it is difficult to conceive of such a hard-and- fast distinction. It would be easier, believing in continuity to suppose that a certain responsive reaction, characteristic of the simplest living substance, had become accentuated in some tissues, and not so accentuated in others, according to their different functional requirements. Thus the belief held so implicitly by physiologists that nerves exhibit no motile response whatsoever ^ becomes questionable, and is seen to require investigation. After submitting it to this, moreover, one finds it difficult to understand how such an * ' Nerves are irritable ; when they are stimulated, a change is produced in them ; this change is propagated along the nerve, and is called a nervous impulse ; there is no change of form in the nerve visible to the highest powers of the microscope.' (Kirke's Handbook of Physiology^ 15th edition, p. 105.) 508 COMPARATIVE ELECTRO-PHYSIOLOGY idea ever gained currency, unless, indeed, it was due to the tyranny imposed on our thought by these arbitrary classifi- cations themselves. Before entering, however, on the question whether the excitatory reaction in nerve finds motile expression or not, we shall first examine the only method at present available for the detection of the condition of excitation. Since ex- cited nerve has hitherto been supposed to exhibit no visible change, it followed that the only method possible for the detection of the excitatory change was the electrical. In- vestigations on nerve, therefore, had perforce to be carried out by this means, through the medium either of the capillary electrometer or of the sensitive galvanometer. But the elec- trical method labours under certain inherent disadvantages, and first of these is the objection which it raises to the free employment of the most convenient form of stimulus, that, namely, by induction shocks. For we have seen that unless extraordinary precautions are taken, we have here, owing to the possible escape of current, an element of error and un- certainty in the results. If, on the other hand, it should become possible to obtain mechanical response from the nerve, this particular form of stimulation might be employed without misgiving. The second limitation which the electrical mode of detection imposes upon us is that arising from the differ- ential character of the response which it indicates. For stimulus induces electrical changes at both the contacts — proximal and distal — the record made being finally due to the algebraical summation of the two. It is true that the excitability of one contact is artificially depressed by injury. But it is often difficult to say how far this injury has been effective in completely abolishing the excitability of this point. The depression of excitability, due to partial injury, will sometimes disappear to a certain extent, with lapse of time, and much uncertainty sometimes occurs as to whether a certain curious variation in the response of the nerve — negative followed by positive — is due to this or some other MECHANICAL RESPONSE OF NERVE 509 cause. With mechanical response, however, provided this could be rendered practicable, no such difficulty need arise. For in that case it would be the direct effect of the exci- tatory change, uncomplicated by any other disturbance, which would be recorded. Finally, as regards the detection of the excitatory change itself, the galvanometer is unable to indicate any change below a certain high intensity of excitation. Thus it gives no indication when excitation is due to one or to a few shocks : it can only detect an excitatory effect which is much stronger than this, having been brought about by the super- posed effects of tetanic shocks of a certain duration. In order to obtain even such effects, a galvanometer of very high sensitiveness is necessary. That of a fairly delicate instrument, detecting a current of about -ooi ampere, will have to be exalted some ten millions of times before it can give efficient indications of excitatory effects in nerves ; and in such a degree of galvanometric sensitiveness we approach a limit which cannot be very much exceeded. Returning now to our original question, we have first to determine whether excitation causes any motile effect in nerve. Under observation, it is easily seen that when the nerve is excited by tetanic electrical shocks it increases in thickness and at the same time shortens in length. We have here a phenomenon in every way analogous to the thickening and shortening of muscle under excitation. The contraction which occurs in nerve, moreover, is of an order by no means microscopic. I give here a record (fig. 313) of the contractile response of nerve under continuous stimulation by fairly strong tetanising electric shocks. This record was obtained by means of the ordinary lever-recorder, the magnification employed being only three times. The induced contraction in this particular case was about 14 per cent. It will also be seen that this contraction reached a limit, at which state of maximum contraction the nerve remained for a considerable time. After this we observe a tendency I 5IO COMPARATIVE ELECTRO-PHYSIOLOGY to decline, owing to fatigue. In some other cases, moreover, I have obtained a contraction of as much as 20 per cent. If we wish to obtain a series of successive responses, however, it is desirable to avoid over-stimulation of the tissue. In order, then, to obtain a response-record under moderate stimulation, we have to employ a higher magni- fication. This magnification, if made about 200 times, is more than sufficient for all practical purposes, and the photo- graphic records given in the course of the present chapter are of this order. With long specimens of nerve, however, a magnification of fifty times would be enough, and in the Fig. 313. Record of Contractile Response in Frog's I^erve under Continuous Electric Tetanisation. Magnification, three times. course of the next chapters, I shall give certain records on this scale, obtained directly on a smoked glass surface. The apparatus used for the purpose was the Kunchangraph (Sanskrit, kunchan^ contraction), which I had already devised and employed in recording the contractile responses of plant- tissues. This apparatus, as adapted for the purpose of recording mechanical response in nerves, consists of, first, a nerve-chamber, N ; secondly, a modified Optical Lever, O ; and thirdly, a photographic recorder, D (fig. 314). Of these, the nerve-chamber consists of a small rectangular ebonite box, the front of which is closed by a semi-cylindrical MECHANICAL RESPONSE OF NERVE 511 Fig. 314. Optical Kunchangraph for Record of Mechanical Response of Nerve nerve chamber containing nerve with electrical connections, E f/. Thread tied to lower end of nerve, and attached to short arm of optic lever, O. Beam of light from L reflected from mirror of optical lever, O, falls on recording-drum, d. Adjustment of reflected spot of light made by micrometer screw, s. Periodic electric stimulation at intervals of one minute is automatically made by means of key regulated by clock-work. Air bubbles through water at vv, and is led on by india-rubber tubing, T, to nerve-chamber, thus kept humid. By proper manipulation of stop-cock apy vapour— as chl oroform— con- tained in vessel v, may be passed through nerve-chamb er, subsequent responses showing eff"ect. 512 COMPARATIVE ELECTRO-PHYSIOLOGY glass cover. The nerve is placed vertically within this, and held, at its upper end, by a clamp. The lower end of the nerve is connected with the short arm of the Optical Lever by means of a thread, which passes through a hole in the floor of the chamber. A second thread of cotton moistened with saline solution hangs loosely from the end of the nerve, and is connected with the electrode e'. When the electrodes E and e' are put in connection with the secondary of an induction coil R, the entire length of the nerve is subjected to direct excitation. When, on the other hand, we wish to study the effect of transmitted excitation, the nerve is lightly clamped at B (fig. 315). Excitation is then induced in the portion of the nerve A a, and after transmission through the inter- vening tract, causes the motile effect in the responding portion of the nerve B c. One precaution which I find to be very necessary is the maintenance of the properly humid condition in the nerve-chamber. This is specially important in the warm weather which characterises the greater part of the year in India. The usual means of keeping the chamber moist, by a large quantity of blotting-paper soaked in water, is not sufficient to bring about the maintenance of the normal excitability of the nerve for any length of time. This need was met by keeping moist vapour in uniform circulation through the nerve-chamber. An air-bag is kept under suitable pressure, and the air, bubbling through water in the vessel w, is made to enter the nerve-chamber through an entrance-pipe, and to escape by an exit-pipe. In warm weather it is well to keep fragments of ice in the water-vessel. By proper mani- pulation of the stop-cock of the air-bag, a gentle stream of cooled and humid air is kept in constant circulation through Fig. 315. Diagrammatic Representation of Ar- rangement for Obtaining Transmitted EflFect of Stimulus. L, indicating lever MECHANICAL RESPONSE OF NERVE 513 the chamber. Observing these precautions, I have been able to obtain responses from a given nerve for as much as three hours continuously, whereas, without this care, they would have come to a stop in a very short time. By a modification of this arrangement, we are also enabled to study the effect on the excitability of the nerve of various gases and vapours contained in a second vessel, v. A series of responses is first taken, under normal conditions—that is to say, when the nerve is surrounded simply by a moist atmosphere. On now turning a three-way tap in a given direction, the water-vapour can be made to pass through the vessel V, filled with the given gas or vapour, before reaching the nerve-chamber. The series of responses then obtained will show either the immediate or the after-effect of the reagent at will. For it is easy, by means of the three-way cock, to shut off the gas and re-establish the first or normal condition, after which the responses will afford an indication of the nature of the after-effect. The lower end of the nerve, as has been said, is attached to the arm of the lever which passes through the fulcrum-rod. A light mirror is fixed on the fulcrum-rod, its face being downwards. The pull caused by the excitatory contraction of the nerve causes rotation of the fulcrum-rod, and this in turn gives rise to a deflection of the spot of light reflected from the mirror. A responsive relaxation of the nerve would give rise, on the other hand, to a deflection of the spot of light in the opposite direction. The long arm of the lever, it will be noticed, is here the ray of light. The responsive movement of the spot of light is recorded on a moving photographic plate vertically below the mirror, and whose [■Movement, regulated by clockwork, is in a direction at right angles to that of the spot of light. The photographic plate, I or the film wrapped round the drum, moves under a fixed ■R^ooden cover, not shown in the figure, which is provided with a narrow incised slit. The length of this is parallel to the (direction of the movement of light, and at right angles to that ©f the plate or film. The advantage of having the plate 514 COMPARATIVE ELECTRO-PHYSIOLOGY vertically below the mirror lies in the fact that a lighted candle may be placed in the dark room without spoiling the record by diffuse illumination. The only way in which such diffuse light could now find access to the plate would be by reflection from the ceiling. But if the ceiling of the experimental room is blackened, or a black cover placed over the nerve-chamber at a certain height, even this possibility is eliminated. The advantage which the observer enjoys, when, instead of groping in semi-darkness, he can work in a fairly well-lighted room is obvious. By making the arm of the lever to which the nerve is attached sufficiently short, and by placing the recording plate sufficiently far away, a wide range of magnification, from several hundreds to several thousands, may be obtained. It may sometimes be desirable to subject the nerve to a certain amount of tension, and this is secured by placing a small weight on the arm of the lever. With high magnification, due adjustment, which is very troublesome, lies in bringing the spot of light con- veniently over the recording plate. This difficulty is obviated, however, by means of a fine micrometer screw S which moves the whole nerve-chamber up or down, in relation to the Optical Lever. The adjustment of this screw in a right- handed manner then moves the spot of light in one direction, say to the left, while its left-handed rotation moves it to the right. This movement can be made very fine, and the spot adjusted to any part of the photographic field. It remains to deal with the possible disturbances inci- dental to the high magnification employed. Apprehension, in this matter, is often more fanciful than real. Disturbances might no doubt occur, however, when proper conditions are not secured for the experiment. If the nerve-chamber, for example, be supported on a different stand from that of the Optical Lever, then the slightest tremor of the common pedestal would result in relative movements of the two supports, causing constant disturbance of the spot of light. Under these conditions, heavy stone pedestals, erected on steady foundations, afford no security against the ground- MECHANICAL RESPONSE OF NERVE 515 vibrations of a busy city. But when both the nerve- chamber and the Optical Lever are fixed to the same supporting-rod, relative movements, due to external disturb- ance, are practically eliminated. This common supporting- rod may be screwed securely to a wall. With these precautions, I have been able to take records, without the least dis- turbance from the adjacent electric tram line. As a matter of fact, when the magnification required is only of a few hundred times, nothing but gross carelessness could allow any source of disturbance to remain. It is only when the magnification has to be pushed to the order of a hundred thousand that unusual care is necessary to avoid errors of disturbance. One precaution which should, however, be taken, is that arising from disturbance of the mirror by convection currents of air. The remedy for this is obvious, namely* a suitable glass cover. This is the order of magnification which is necessary for the recording of response under a degree of stimulation usual in making observations of excitatory electrical variation with a very sensitive galvanometer. But while the sensitive- ness of the galvanometric method of detecting respons^ is here nearing its limit, that of the mechanical method is in* its first stage only, and how greatly the sensitiveness of the latter may be exalted when required will be shown in the next chapter. I shall now explain how easy it is to study the physio- logical variations induced in the animal nerve under various agencies by means of the mechanical response. The following experiments were performed on specimens of the sciatic nerve of frog. A well-known reagent for abolition of ex- citability of the nerve is ammonia. Its effect on mechanical response is seen in fig. 316. In all the following experiments, the stimulus applied was by fairly strong tetanising electrical shocks, which were usually of two seconds' duration. Two series of records were taken, successive responses being recorded at intervals of one minute, before and after the application of thq chemical reagent. In fig, 316, the normal L L 3 I 5i6 COMPARATIVE ELECTRO-PHYSIOLOGY responses seen in the first series are found to be abolished when the nerve has been subjected to strong vapour of ammonia for some time. It should be mentioned here that this abolition takes place under the action of a strong dose. When highly diluted with air, the vapour may cause a temporary exaltation. In the next figure (fig. 317) is shown the effect of morphia. After the application of this solution for a certain length of time, the response is seen to be abolished. The strength of application which brings about this abolition I find to vary according to the condi- FlG. 316. Photographic Record of Effect of Ammonia on Mechanical Response of Frog's Nerve First series of responses are nor- mal. Second series show effect of ammonia in practical aboli- tion of response. Fig. 317. Photographic Record showing Abolition of Mechanical Response of Frog's Nerve by Action of Solution of Morphia tion of the nerve. Another agent by which the mechanical response of the nerve is found to be abolished is aconite. And it is of special interest to note that I have often found this to act as an antidote, for the revival of response previously almo.st completely abolished by morphia. The condition of the nerve here also appears to be a determining factor in the mutually antidotal action of these two poisons. A strong application of