Midbrain structures, including the periaqueductal gray (PAG), are essential nodes in vertebrate motor circuits controlling a broad range of behaviors, from locomotion to complex social behaviors such as vocalization. Few single unit recording studies, so far all in mammals, have investigated the PAG's role in the temporal patterning of these behaviors. Midshipman fish use vocalization to signal social intent in territorial and courtship interactions. Evidence has implicated a region of their midbrain, located in a similar position as the mammalian PAG, in call production. Here, extracellular single unit recordings of PAG neuronal activity were made during forebrain-evoked fictive vocalizations that mimic natural call types and reflect the rhythmic output of a known hindbrain-spinal pattern generator. The activity patterns of vocally active PAG neurons were mostly correlated with features related to fictive call initiation. However, spike trains in a subset of neurons predicted the duration of vocal output. Duration is the primary feature distinguishing call types used in different social contexts, and these cells may play a role in directly establishing this temporal dimension of vocalization. Reversible, lidocaine inactivation experiments demonstrated the necessity of the midshipman PAG for fictive vocalization, while tract-tracing studies revealed the PAG's connectivity to vocal motor centers in the fore- and hindbrain comparable to that in mammals. Together, these data support the hypotheses that the midbrain PAG of teleosts plays an essential role in vocalization and is convergent in both its functional and structural organization to the PAG of mammals.