Cell proliferation follows acoustically-induced hair cell bundle loss in the zebrafish saccule

Fishes are capable of regenerating sensory hair cells in the inner ear after acoustic trauma. However, a time course of auditory hair cell regeneration has not been established for zebrafish. Adult zebrafish (Danio rerio) were exposed to a 100 Hz pure tone at 179 dB re 1 microPa RMS for 36 h and then allowed to recover for 0-14 days before morphological analysis. Hair cell bundle loss and recovery were determined using phalloidin to visualize hair bundles. Cell proliferation was quantified through bromodeoxyuridine (BrdU) labeling. Immediately following sound exposure, zebrafish saccules exhibited significant hair bundle damage (e.g., splayed, broken, and missing stereocilia) and loss (i.e., missing bundles and lesions in the epithelia) in the caudal region. Hair bundle counts increased over the course of the experiment, reaching pre-treatment levels at 14 days post-sound exposure (dpse). Low levels of proliferation were observed in untreated controls, indicating that some cells of the zebrafish saccule are mitotically active in the absence of a damaging event. In sound-exposed fish, cell proliferation peaked two dpse in the caudal region, and to a lesser extent in the rostral region. This proliferation was followed by an increase in numbers of cuticular plates with rudimentary stereocilia and immature-like hair bundles at 7 and 14 dpse, suggesting that at least some of the saccular cell proliferation resulted in newly formed hair cells. This study establishes a time course of hair cell bundle regeneration in the zebrafish inner ear and demonstrates that cell proliferation is associated with the regenerative process.