Evaluation of Cellular and Molecular Pathology in the Rodent Vestibular Sensory Epithelia by Immunofluorescent Staining and Confocal Microscopy

Hair cells in the vestibular and cochlear sensory epithelia are the main target of ototoxic drugs. Nevertheless, the synapses between the hair cells and the afferent terminals of the post-synaptic ganglion neurons have also been shown to be a target of ototoxic damage. In this chapter, we describe immunohistochemistry protocols adapted to the quantification of hair cells and synapses in the vestibular epithelia to assess ototoxic damage in rodents. Epithelia are immunolabeled intact and are used in whole-mount preparations for the quantification of hair cell numbers by confocal microscopy imaging. For synaptic assessment, the epithelia are first immunolabeled, embedded in a gelatin/albumin block, and then sectioned in a vibrating microtome before confocal microscopy imaging. The data thus obtained offer a thorough evaluation of the damage suffered by the vestibular sensory epithelia, including overt hair cell loss and subtle synaptic loss. Together, these pathological outcomes determine the loss vestibular input and the resulting behavioral alterations.