The overall goals of the Program Project are to study the neural and cellular bases of labyrinthine function, relevant to balance and equilibrium. The investigators will pursue a multifaceted, interdisciplinary research approach to test the central hypothesis of this Program, that the origins of diverse primary vestibular afferent response dynamics can be found distributed among the steps of the transduction cascade from head acceleration to afferent discharge modulation. Those steps are (1) biomechanics, (2) mechano-transduction, (3) basolateral currents, (4) hair cell synapses, and (5) postsynaptic factors. The proposed studies will evaluate the contributions of each of these categories to the differential response dynamics of individual canal nerve afferents, utilizing the vertebrate fish Opsanus tau (toadfish) as the model experimental system for all proposed experiments. Specifically, cupular structure, and the relative motion of the cupula and single hair cell stereocilia in response to mechanical stimuli will be evaluated. Transduction will be investigated through voltage clamp studies of semicircular canal hair cells in-situ. These experiments will examine the regional differences in membrane currents generated in response to hair bundle motion, and will test the hypothesis that such differences influence the construction of individual nerve response dynamics. The relationships of transduction voltage and/or current to membrane and endolymphatic potentials will be measured, to establish a working model for the generation of transduction in-vivo. Results from these studies will establish the electro-chemical characteristics of hair cell apical transduction. Other experiments will document the transcupular pressure differentials that drive cupular motion. Cupular deformation, and stereociliary coupling to the cupula will be observed with video microscopy and quantitated by image analysis. Afferent nerve responses will be further evaluated in plugged semicircular canals. The ultrastructural organization and synaptology of functionally-characterized, biocytin-injected primary afferents will be characterized and quantified using light and electron microscopy. The effects of efferent vestibular nuclei stimulation on hair cell receptor and membrane potentials, and upon transmitter release as measured by mEPSPs will be determined. These experiments will provide a systematic study of the origins of primary vestibular afferent response dynamics in a single vertebrate species. As such, the work will offer new insights into labyrinthine function, and is thereby critical for the development of effective therapies for motion sickness and peripheral vestibular disorders
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