There is evidence from several studies showing that sensory cells which are destroyed by trauma in the chick auditory epithelium are replaced by new cells. The fate of neurons that innervate the injured and degenerating sensory cells in the lesion, and the temporal sequence of re-innervation of regenerated hair cells are not well understood. This study examined efferent terminals in the chick auditory sensory epithelium following acoustic overstimulation using synapsin-specific immunocytochemistry. Chicks were exposed to an octave band noise (1.5 kHz center frequency, 116 dB SPL, 16 h) and killed on each day from 0 to 9 days postexposure. In the proximal half of control whole mounts of the basilar papillae, synapsin-specific immunoreactivity stained efferent terminals throughout the abneural portion of the sensory epithelium (the short hair cell region). In this area, the labeling appeared as 2-3 bouton-shaped clusters along the abneural edge of each hair cell. After acoustic overstimulation, a lesion was observed at the abneural edge of the papilla where many short hair cells were lost. The center of the lesion was located at 40% distance from the proximal end of each traumatized papilla. Synapsin-specific labeling was not found in sites where expanded supporting cells had replaced missing hair cells. Hair cells which survived the trauma exhibited a shrunken apical area, and synapsin-labeled boutons were observed near their basal domains. New hair cells, which first appeared in the papilla 4 days after trauma, were not initially associated with synapsin-labeled boutons. Regenerated hair cells first displayed contacts with synapsin-labeled boutons 7 days after trauma. Nine days after acoustic overstimulation, most new hair cells appeared to be associated with synapsin-labeled boutons which resembled the normal horseshoe configuration of efferent terminals. The data suggest that direct contact with functional efferent synapses is not necessary for the generation and differentiation of new hair cells.