IN VIVO AND IN VITRO HAIR BUNDLE REGENERATION AFTER GENT

Project: Research project

Project Details

Description

Mature amphibian otolith hair cells display a variety of adaptation kinetics which confer stimulus encoding and frequency selective properties. Adapting hair cells do not retain information about sustained hair bundle displacement, and thus are most sensitive to dynamic stimuli. Adaptation is mediated by calcium and a cytoplasmic motor which releases tension on the stereociliary tip-links, mechanically closing the transduction channels. The most likely candidate for this cytoplasmic motor is a myosin isoform. Thus, antisera against stereociliary myosin and transduction channels can be used as markers of hair cell functional maturity. Previous studies have also shown that hair cell regeneration in amphibian otolith organs can occur in mitotically-blocked organotypic cultures, and suggested that non-mitotic hair cell regeneration might be accomplished by the phenotypic conversion of supporting cells into hair cells. Alternatively, immature hair bundles might develop in damaged hair cells undergoing repair, or post-mitotic progenitor cells undergoing terminal differentiation. We will use gentamicin sulfate to induce hair cell degeneration in bullfrogs in vivo, and in organotypic cultures of bullfrog otolithic organs. We will then use immunocytochemical, histochemical, and ultrastructural techniques to study the subsequent labeling patterns in regenerating hair bundles. The proposed experiments have been designed to determine if in vitro labeling patterns for hair bundle regeneration accurately reflect those in vivo. We will test the hypotheses that immature hair bundles develop on either surviving. but damaged, hair cells, and/or immature hair cells derived from either post-mitotic progenitors or converting supporting cells in vivo and in mitotic and mitotically-blocked organotypic cultures. We will also investigate the temporal organization of soluble and filamentous actin in the early stages of hair bundle repair in damaged hair cells and/or cuticular plate and hair bundle development in post-mitotic progenitors and converting supporting cells in mitotically-blocked cultures. This will reveal labeling pafterns in damaged hair cells-undergoing repair, immature hair cells derived from post-mitotic progenitors, and/or converting supporting cells. We will also test the hypothesis that immature hair bundles possess myosin and transduction channels, and if so, at what size of hair bundle development they are expressed. The expression of myosin I- and transduction channel proteins in a specific bundle size or larger will allow these bundles to be selected and tested for hair cell function in future electrophysiological studies. This work might suggest new directions for rehabilitation of hair cell loss in hearing and vestibular disorders.
StatusFinished
Effective start/end date1/1/9712/31/98

Funding

  • National Institute on Deafness and Other Communication Disorders

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