Mechanisms of sensory hair cell reinnervation following lateral line cranial nerve damage in Danio rerio

项目总结 听力丧失在美国是一种流行的感官病理，影响着3000多万人。一个 很大比例的耳聋归因于感觉神经性听力损失，这通常涉及到 传入神经纤维将听觉信息从内耳的机械敏感毛细胞传递到 大脑。生理听力的恢复需要感觉神经传入纤维的再生。 耳蜗管上皮，然后适当的毛细胞靶点的神经再支配。神经再生 由于内耳的可及性有限，目前还缺乏对人类和其他哺乳动物模型的研究。这个 斑马鱼侧线系统由浅表液流检测毛细胞和传入神经纤维组成， 提供了一种简单易用的神经再生模型。在这个模型中，可能有各种旁分泌， 旁分泌和神经元自主信号机制协同工作，指导轴突寻径和 目标选择。该方案的目标1将确定目标感官所表达的分子线索 毛细胞通过再生侧线的传入轴突来引导神经再支配。在横切之后 斑马鱼的侧线神经、毛细胞将在多个时间点被分离。在这些毛细胞中， 可用于吸引轴突生长的典型和非典型分子线索的表达变化 锥体将通过转录组测序进行量化。目标2将调查神经元的偏向 发育相关毛细胞群体的神经再生。尽管研究表明神经元保留了 对于它们最初的毛细胞目标的记忆，这种记忆是如何建立或维持的是未知的。一个 转基因成像技术将用于标记和追踪下列再生轴突的克隆群体 横断侧线神经。假设神经元更喜欢重新支配毛细胞，这些毛细胞起源于 发育过程中共有的感官代用品。目标3将揭示当地物理环境的变化 以允许单个传入纤维进入它们的靶器官。通过成像 用荧光标记的雪旺细胞和胶原蛋白转基因鱼，雪旺细胞将发生变化 神经束和上皮性基底膜，以允许单个轴突进入斑马鱼神经肥大， 其中含有目标毛细胞。推测雪旺细胞和基底中存在物理缝隙。 靠近失神经毛细胞的膜层，允许再生轴突分支通过 从主神经束上剥离。总之，这些研究将阐明支配传入神经的机制。 毛细胞的再生和再神经支配，这可能为恢复听力提供洞察力 耳聋的人类耳蜗虫。这些研究将在A.J.哈德斯佩斯博士的直接指导下进行 洛克菲勒大学和#年三所院校医学博士项目的支持环境 纽约市。

PROJECT SUMMARY Loss of hearing is a prevalent sensory pathology in the United States that affects over 30 million people. A significant proportion of deafness is attributed to sensorineural hearing loss, which often involves the damage of afferent nerve fibers which relay auditory information from the mechanosensitive hair cells of the inner ear to the brain. The restoration of physiologic hearing would require the regeneration of afferent fibers into the sensory epithelium of the cochlea, followed by the reinnervation of appropriate hair cell targets. Nerve regeneration studies in humans and other mammalian models are lacking due to the limited accessibility of the inner ear. The zebrafish lateral line system, composed of superficial fluid-flow detecting hair cells and afferent nerve fibers, offers a simple and accessible model of nerve regeneration. In this model, there are likely various paracrine, juxtacrine, and neuron-autonomous signaling mechanisms working in coordination to guide axon pathfinding and target selection. Aim 1 of this proposal will determine the molecular cues expressed by target sensory hair cells to guide reinnervation by regenerating afferent axons of the lateral line. Following transection of the lateral line nerve, hair cells from the zebrafish will be isolated at multiple timepoints. In these hair cells, the expression changes of canonical and non-canonical molecular cues that may be used to attract axonal growth cones will be quantified through transcriptome sequencing. Aim 2 will investigate the neuronal bias for reinnervation of developmentally related hair cell populations. Although studies suggest that neurons retain a memory for their original hair cell targets, how this memory is established or maintained is unknown. A transgenic imaging technique will be used to label and trace clonal populations of regenerating axons following transection of the lateral line nerve. It is hypothesized that neurons prefer reinnervating hair cells that arose from a shared sensory placode during development. Aim 3 will reveal changes in the local physical environment of the regenerating nerve to allow entry of individual afferent fibers into their target organ. By imaging transgenic fish with fluorescently labeled Schwann cells and collagen, changes will be shown in Schwann cell tracts and the epithelial basement membrane to permit entry of individual axons into the zebrafish neuromast, which contains target hair cells. It is hypothesized that physical gaps form in Schwann cell and basement membrane layers in close proximity to denervated hair cells to allow passage of regenerating axons branching off the main nerve bundle. Together, these studies will elucidate the mechanisms governing afferent nerve regeneration and the reinnervation of hair cells, which may provide insight towards restoring hearing in the deafened human cochlea. These studies will be carried out under the direct mentorship of Dr. A. J. Hudspeth at The Rockefeller University and within the supportive environment of the Tri-Institutional MD-PhD Program in New York City.