Molecular and anatomical correlates of spiral ganglion neuron heterogeneity

 描述（由申请人提供）：我们的听力是我们与周围世界沟通的关键。与大多数哺乳动物一样，人类的听力损失随着年龄的增长而逐渐减弱。然而，由于自愿或非自愿地暴露在嘈杂的环境中，听力障碍在年轻人中变得越来越普遍。内毛细胞（IHC）和螺旋神经节神经元（SGN）的损失是年龄相关的和噪声诱导的听力障碍的标志。适当地，正在努力通过使用基于干细胞的方法取代这两种细胞类型来恢复听力。几十年的电生理学研究表明，SGN在功能上是异质的，因此通过替换丢失或受损的神经元来恢复正常听力需要仔细考虑不同的SGN亚型。这一点尤其重要，因为已经发现具有低自发放电率的SGN子集特别容易受到噪声引起的损害。不幸的是，除了基于自发放电率的分类之外，关于I型SGN的异质性知之甚少，I型SGN是唯一支配IHC的SGN。值得注意的是，对于I型SGN分类的任何生物学相关基础，没有已知的分子标志物。这一直是一个主要的障碍，不仅在调查的意义SGN异质性的听觉编码，而且在产生精确的亚群后，噪音创伤。在这里，我建议定义SGN异质性的分子和解剖学相关性，并检查不同SGN亚型的功能意义。具体目标1涉及使用下一代测序和单细胞转录谱分析工具， 生成出生后第20天小鼠的个体SGN的转录组，其将用于鉴定对应于生物学相关SGN亚型的转录簇。该目的还试图通过急性和选择性沉默来了解所鉴定的SGN亚型对小鼠听力的重要性。具体目标2侧重于确定SGN异质性的解剖学相关性，并寻求回答该领域长期存在的问题-SGN队列是否支配单个IHC，其在神经节和分子谱中的定位不同？为了实现这一点，将使用跨突触病毒追踪方法在小鼠中用GFP稀疏地标记个体IHC及其神经支配SGN。这些实验的结果应该大大提高我们的理解SGN异质性独立于他们的apico-basal位置，并承诺打开新的大门，调查听觉编码的SGNs的水平。此外，SGN异质性的分子指纹的知识应有助于进一步研究不同SGN亚型的发展，这将有利于该领域正在进行的努力，以通过替代丢失的SGN来恢复听力。

 DESCRIPTION (provided by applicant): Our ability to hear well is key to our communication with the world around us. As in most mammals, loss of hearing occurs with age in humans with debilitating results. However, hearing deficits are becoming increasingly more common in younger people due to voluntary or involuntary exposure to noisy environments. Loss of inner hair cells (IHCs) and spiral ganglion neurons (SGNs) are the hallmarks of age-related and noise- induced hearing disorders. Appropriately, efforts are underway to restore hearing by replacing both of these cell types using stem-cell based approaches. Several decades of electrophysiological studies have shown that SGNs are functionally heterogeneous, so restoring the full range of normal hearing by replacing lost or damaged neurons will require careful consideration of different SGN subtypes. This is especially important given that a subset of SGNs with low spontaneous firing rates has been found to be particularly vulnerable to noise-induced damage. Unfortunately, little is known beyond the spontaneous firing rate-based classification regarding heterogeneity of Type I SGNs, which are the only ones that innervate IHCs. Notably, no molecular markers are known for any biologically relevant bases of Type I SGN classification. This has been a major roadblock, not only in investigating the significance of SGN heterogeneity for auditory coding, but also in generating the precise subpopulations lost after noise trauma. Here I propose to define the molecular and anatomical correlates of SGN heterogeneity and to examine the functional significance of different SGN subtypes. Specific aim 1 involves use of next generation sequencing and single-cell transcriptional profiling tools to generate transcriptomes of individual SGNs of postnatal day 20 mice, which will be used to identify transcriptional clusters that correspond to biologically relevant SGN subtypes. This aim also seeks to understand the significance of the identified SGN subtypes for hearing in mice by acutely and selectively silencing them. Specific aim 2 focuses on determining the anatomical correlates of SGN heterogeneity and seeks to answer a longstanding question in the field- are the cohort of SGNs innervating a single IHC diverse in their positioning within the ganglion and molecular profiles? To accomplish this, a transsynaptic viral tracing approach will be used to sparsely label individual IHCs and their innervating SGNs with GFP in mice. Results of these experiments should vastly improve our understanding of SGN heterogeneity independent of their apico-basal position, and promise to open new doors to investigation of auditory coding at the level of SGNs. Furthermore, knowledge of molecular fingerprints of SGN heterogeneity should facilitate further study of development of different SGN subtypes, which will be of benefit to ongoing efforts in the field to restore hearing by replacing lost SGNs.