Structural differentiation of mechansosensory hair cells during inner ear development and innervation: Emergence of kinocilia and synapic ribbon

This dissertation examines the emergence of kinocilia and synaptic ribbons during mechanosensory hair cell differentiation in Xenopus laevis inner ear development. As part of this effort, immunohistochemical procedures were optimized that permitted examination of the innervation of larval and post metamorphic Xenopus laevis inner ears by 8th cranial nerve axons through detection of acetylated-alpha-tubulin, a protein that is an established label of axons in neural development.; Data from several microscopy techniques (transmission electron microscopy, scanning electron microcopy, confocal microcopy, and brightfield microscopy) show that the acetylated-alpha-tubulin positively labels and detects target protein in areas where the 8th cranial nerve (auditory-vestibular nerve) and where mechanosensory cell components are present. In the embryonic stage, stage 31 (S31), where the otic primordial epithelium is emerging, acetylated-alpha-tubulin positively labels the initial outgrowth of the kinocilium which seems to be devoid of any innervation contacts to the otic vesicle from the 8th cranial nerve. In the hatchling and early larval stages of Xenopus laevis inner ear development, results suggest that the axonal field is chiefly restricted to regions of the epithelia where hair cells are present. Additionally, hair cells begin to differentiate and organize their accessory structures, such as the kinocilum, in conjunction with developmental extension of 8th nerve axons into the sensory epithelium. Tubulin expression in the 8th nerve, cell body, and kinocilia increases as the otic vesicle enlarges and differentiates in development, suggesting that the level of tubulin expression appears indicative and correlated to the stage of development of Xenopus laevis. Furthermore acetylated-alpha-tubulin positively labels only specifically where the sensory field starts to become organized, and tubulin expression forms visible patterns where auditory and vestibular endorgans are emerging.; A second study was initiated to examine if, and when, ribbon synapses appear during the development of Xenopus laevis inner ear end organs. Ribbon synapses are specialized presynaptic structures in primary sensory cells that are involved in the fast release of neurotransmitters. Transmission Electron Microscopy (TEM) data collected from two Xenopus laevis stages (pre-metamorphic stage 55 and post-metamorphic juvenile) confirms the presence of "ribbon-like" synapses. This result prompts the hypothesis that synaptic ribbons begin to appear early in auditory development after the invagination of the sacculus and when the sensory epithelia is first beginning to form end organs.; Results from this research have established a time-line for key anatomical events during synaptogenesis, morphogenesis, organogenesis, and neurogenesis which presently are not well characterized as part of inner ear developmental studies. Knowledge of the emergence of kinocilia and synaptic ribbons during mechanosensory hair cell differentiation, as well as of the developmental innervation patterns of the 8th nerve, can aid in furthering the identification of physiological mechanisms and factors which may influence normal development of synaptic connections for acoustico-vestibular sensory processing. Thus, results have relevance for ongoing biomedical research that aims to identify disorders of hearing and balance which are based on mechanosensory hair cell dysfunction.