Glutamate-like Immunoreactivity During Hair Cell Recovery After Gentamicin Exposure in the Chinchilla Vestibular Sensory Periphery

Objective: Determine the expression of glutamate by immunohistochemistry in normal and recovering vestibular hair cells in the chinchilla crista ampullaris after gentamicin ototoxicity. Study Design: In five groups of three animals each, ototoxicity was produced by placing gentamicin (50 μg)-impregnated Gelfoam pellets within the perilymphatic space of the superior semicircular canal. Animals were sacrificed at 1, 2, 4, 8, and 16 weeks after treatment. A group of normal (n=3) animals was also processed. Methods: For the detection of glutamate the inner ears of these animals were dissected, and the horizontal cristae ampullaris embedded in plastic. Two-micron-thick tissue sections were obtained and incubated with monoclonal antibodies against glutamate. The immunoreaction was detected using the avidinbiotiny-lated-complex technique and diaminobenzidine was the chromogen. Results: Normal sensory epithelia demonstrated type I and type II hair cells with moderate glutamate-like immunoreactivity. Supporting cells demonstrated no glutamate-like immunoreactivity. Afferent nerve fibers and calyxes surrounding type I hair cells demonstrated strong glutamate-like immunoreactivity. At 1 and 2 weeks after treatment the few type II hair cells surviving ototoxic treatment (15%–18%) contained moderate glutamate-like immunoreactivity, supporting cells showed no immunoreactivity, and nerve terminals and fibers displayed strong immunoreactivity. At 4 and 8 weeks after treatment, recovered hair cells (80%) had greater glutamate-like immunoreactivity when compared with normal hair cells, supporting cells displayed no glutamate-like immunoreactivity, and afferent fibers contained strong glutamate-like immunoreactivity. At 16 weeks, glutamate-like immunoreactivity in hair cells returned to normal level. Conclusion: Glutamate may be used as an indicator of hair cell differentiation and as an index of the molecular recovery of hair cells after ototoxicity.