Anxiety, a debilitating sense of fear, involves an interconnected brain network that includes the medial prefrontal cortex (mPFC), amygdala, and periaqueductal grey (PAG). However, the specific organization and cellular physiology of distinct pathways within this network are undefined but critical for understanding anxiety disorders. This thesis elucidated the circuitry, pharmacology, and intrinsic physiology of defined pathways within the mPFC-amygdala-PAG circuit. Comparison of mPFC neurons projecting to the PAG versus amygdala revealed significant differences in intrinsic properties, local circuitry, and neuromodulation by stress peptides. In the central amygdala (CeA), neurons projecting to the PAG (CeA-PAG neurons) expressed different levels of hyperpolarization-activated current. CeA-PAG neurons were altered by anxiogenic peptide corticotropin-releasing factor (CRF) but not anxiolytic neuropeptide-Y (NPY). Optogenetic activation of axons originating in the PAG or IL drove both excitation and inhibition in CeA-PAG neurons. Together, this thesis has uncovered critical characteristics of a neural network implicated in anxiety.