TEM analysis of neural terminals on autoradiographically identified regenerated hair cells

Regenerated tall and short hair cells identified by autoradiography ([3H]thymidine) were analyzed for their neural contacts using transmission electron microscopy. Ears from mature Coturnix quail (N = 5) exposed to pure tone overstimulation (1500 Hz, 115 dB, 12 h) and treated with [3H] thymidine for 10 days were fixed, embedded, sectioned serially in 100 mu intervals and prepared for autoradiography. At fifty percent length along the papilla, alternating semi-thick (1 micron) and thin (70 nm) sections were taken at 50 microns intervals. Semi-thick sections were analyzed at the light microscope level for autoradiographic labeling of [3H]thymidine over the hair cell nucleus. When an autoradiographically labelled hair cell was identified the corresponding serial thin sections were analyzed in the transmission electron microscope. Seven autoradiographically labeled hair cells in semi-thin sections were positively identified in immediately adjacent thin serial sections. Labeled hair cells were morphologically similar to adjacent cells with no label and generally appeared to receive similar innervation. Regenerated short hair cells showed large chalice shaped, efferent terminals, intermediate hair cells received both afferent and efferent innervation and tall hair cells were contacted by two to three afferent terminals with synaptic specializations. These results provide conclusive evidence of both efferent and afferent synaptic contacts on newly regenerated hair cells of all types 10 days following acoustic trauma.