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, whereas 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.

The midbrain periaqueductal gray (PAG) plays a central role in the descending control of vocalization across vertebrates. The PAG has also been implicated in auditory-vocal integration, though its precise role in such integration remains largely unexplored. Courtship and territorial interactions in plainfin midshipman fish depend on vocal communication, and the PAG is a central component of the midshipman vocal-motor system. We made focal neurobiotin injections into the midshipman PAG to both map its auditory-vocal circuitry and enable evolutionary comparisons with tetrapod vertebrates. These injections revealed an extensive bidirectional pattern of connectivity between the PAG and known sites in both the descending vocal-motor and ascending auditory systems, including portions of the telencephalon, dorsal thalamus, hypothalamus, posterior tuberculum, midbrain and hindbrain. Injections in the medial PAG produced dense label within hindbrain auditory nuclei, while those confined to the lateral PAG preferentially labeled hypothalamic and midbrain auditory areas. Thus, the teleost PAG may have functional subdivisions playing different roles in vocal-auditory integration. Together, the results confirm several pathways previously identified by injections into known auditory or vocal areas and provide strong support for the hypothesis that the teleost PAG is centrally involved in auditory-vocal integration.

The avian forebrain nucleus, the lateral magnocellular nucleus of the anterior neostriatum (LMAN), is necessary for normal song development because LMAN lesions made in juvenile birds disrupt song production but do not disrupt song when made in adults. Although these age-limited behavioral effects implicate LMAN in song learning, a potential confound is that LMAN lesions could disrupt normal vocal motor function independent of any learning role by altering LMANЈs premotor target, the song nucleus, the robust nucleus of the archistriatum (RA). To date, however, no studies have examined directly the effects of LMAN lesions on the circuitry of the RA. We report here that juvenile LMAN lesions rapidly and profoundly affect RA, altering synaptic connectivity within this nucleus, including descending inputs from the song nucleus HVc. Specifically, morphological assays of the dendritic spines of RA projection neurons and axon terminal boutons arising from HVc show a numerical decline in the density of connections in RA in LMANlesioned juveniles compared with controls. Concurrently, LMAN lesions alter excitatory transmission within the juvenile RA: after LMAN lesions, the stimulus-response relationship between HVc fibers and RA neurons steepens, and the amplitude of spontaneous monophasic EPSCs increases. Rather than arresting RA in a juvenile state, LMAN lesions transform the structure and function of RA and its connections, such that it is distinct from that of the normal juvenile. In many ways, RA circuitry in LMAN-lesioned juveniles resembles that of normal adults, suggesting that LMAN lesions induce a premature maturation of the vocal motor pathway, which may lead to a loss of behavioral plasticity and abnormal song development.