Molecular mechanisms of vestibular hair cell exocytosis

描述(申请人提供)：前庭系统中的机械感觉毛细胞将头部位置和运动的信息转化为神经元信号。本研究的目的是阐明前庭毛细胞向传入纤维突触传递的分子机制。毛细胞传入突触表现出解剖学上的(突触小体或突触带)和功能上的(对分级去极化的反应)专门化，区别于神经系统中的其他突触。在这里，我们建议研究在常规突触中重要的蛋白质及其相互作用对毛细胞突触传递的重要性。此外，前庭生理学对I型和II型毛细胞之间的功能差异缺乏了解。只在羊膜中发现的I型毛细胞被传入神经吞噬，这表明突触生理上的差异。因此，拟议的实验旨在进一步了解前庭I型和II型毛细胞在神经递质释放方面的分子和功能差异。这些实验将进一步加深我们对前庭和听觉系统中第一个突触的分子和生物物理理解。 毛细胞从细胞体的基底外侧端释放神经递质，提供了相对容易进入突触区域的途径，从而引入毒素和多肽，从而急剧扰乱蛋白质功能。胞吐作用将通过测量囊泡与质膜融合时细胞膜面积的增加来直接监测。特定目的1验证了胞吐作用受SNARE蛋白特定子集调控的假说。具体目标2解决囊泡回收是否限制胞吐作用。在具体目标3中，验证了谷氨酸在正反馈环路中的作用，以促进释放，特别是在I型毛细胞中。对最初感觉突触释放的蛋白质和机制的详细了解有助于确定针对前庭功能障碍的新治疗方法。

DESCRIPTION (provided by applicant): Mechanosensory hair cells in the vestibular system transduce information about head position and movement into neuronal signals. The aim of this application is to elucidate the molecular mechanisms underlying synaptic transmission from vestibular hair cells to afferent fibers. The hair cells afferent synapse exhibits anatomical (synaptic bodies or ribbons) and functional (responding to graded depolarizations) specializations that distinguish them from other synapses in the nervous system. We here propose to investigate which proteins and protein-interactions important at conventional synapses are important for hair cell synaptic transmission. Furthermore, vestibular physiology lacks an understanding of the functional difference between type I and type II hair cells. Type I hair cells, found only in amniotes, are engulfed by the afferent nerve, suggesting differences in synaptic physiology. The proposed experiments are therefore further aimed at understanding the molecular and functional difference in neurotransmitter release between vestibular type I and type II hair cells. These experiments will further our molecular and biophysical understanding of the first synapse in the vestibular and auditory system. Hair cells release neurotransmitter from the basolateral end of the cell body, offering relatively easy access to synaptic areas for the introduction of toxins and peptides to acutely perturb protein function. Exocytosis will be monitored directly by measuring the increase in cell membrane area that occurs when vesicles fuse with the plasma membrane. Specific Aim 1 tests the hypothesis that exocytosis is regulated by a specific subset of SNARE proteins. Specific Aim 2 addresses whether vesicle recycling is limiting for exocytosis. In Specific Aim 3 the hypothesis is tested that glutamate acts in a positive feedback loop to enhance release especially in type I hair cells. A detailed understanding of the proteins and mechanisms of release at this initial sensory synapse aid in identifying novel therapeutic approaches targeting vestibular dysfunction.