Regeneration and reinnervation of mechanosensory hair cells

描述(由申请人提供)： 项目摘要/摘要内耳是所有脊椎动物的主要感觉器官，调节听觉和平衡(前庭)功能。听觉和前庭感觉上皮的共同之处是感觉毛细胞，它们对声音创伤非常敏感。哺乳动物内耳毛细胞的丧失是不可逆转的，会导致听力障碍、耳聋和平衡障碍。与哺乳动物相比，鱼不仅在一生中产生新的毛细胞，而且具有再生功能毛细胞的先天能力。本研究以斑马鱼作为再生动物模型，重点研究了长寿毛细胞前体的形成和调控，以及新生毛细胞再生的遗传学。这项建议的长期目标是提供对斑马鱼毛细胞更新的分子和遗传方面的了解，并利用这些信息开发强制哺乳动物毛细胞再生和再生的新策略。具体目标1集中在斑马鱼中介导非有丝分裂毛细胞再生的有丝分裂后毛细胞前体的形成和调控。利用一个新鉴定的斑马鱼品系，我们发现Notch mRNA在胚胎后感觉上皮细胞中表达，并且3-分泌酶活性的抑制剂在体内诱导性早熟毛细胞的形成。其目的是(A)确定Notch信号在调节前体分化中的作用，(B)确定有丝分裂后前体在感觉上皮生长和再生过程中的功能，(C)确定长寿毛细胞前体的有丝分裂来源，以及(D)确定斑马鱼非有丝分裂毛细胞再生的遗传基础。特定目标2将确定新生感觉毛细胞神经再支配所需的基因。听觉功能的恢复需要细胞更新和与中枢神经系统的重新连接这一密不可分的过程。然而，利用分子遗传学和斑马鱼的突变鉴定，毛细胞的再生和再神经支配在体内是可以分离的。通过利用简单的视觉和行为分析，我们将重点研究(I)不能再生毛细胞的斑马鱼突变体和(Ii)可再生但不能再生新生毛细胞的突变体。克隆这些突变体中受影响的基因将识别新的基因，或具有新功能的已知基因，这些基因是形成新的毛细胞和重建突触连接所必需的。

DESCRIPTION (provided by applicant): Project Summary/Abstract The inner ear is the primary sensory organ mediating hearing (auditory) and balance (vestibular) function in all vertebrates. Common to both the auditory and vestibular sensory epithelia are sensory hair cells, which are acutely sensitive to acoustic trauma. Loss of hair cells in the mammalian inner ear is irreversible and causes hearing impairment, deafness and balance dysfunction. In contrast to mammals, fish not only produce new hair cells throughout life but also have the innate capacity to regenerate functional hair cells. Using zebrafish as a regenerative animal model, this proposal is focused on the formation and regulation of long-lived hair cell precursors and the genetics of nascent hair cell reinnervation. The long-term goal of this proposal is to provide a molecular and genetic understanding of hair cell renewal in zebrafish and use this information to develop novel strategies that force hair cell regeneration and reinnervation in mammals. Specific Aim 1 is focused on the formation and regulation of postmitotic hair cell precursors that mediate non-mitotic hair cell regeneration in zebrafish. Using a newly identified strain of zebrafish, we show that Notch mRNA is expressed in the postembryonic sensory epithelium and that inhibitors of 3-secretase activity induce precocious hair cell formation in vivo. The goals of this Aim are to (a) determine the role of Notch signaling in regulating precursor differentiation, (b) determine the function of postmitotic precursors during growth and regeneration in the sensory epithelium, (c) identify the mitotic source of long-lived hair cell precursors, and (d) determine the genetic basis of non-mitotic hair cell regeneration in zebrafish. Specific Aim 2 will identify genes required for neuronal reinnervation of nascent sensory hair cells. Restoration of auditory function requires the inextricably linked processes of cell renewal and reconnection to the central nervous system. However, hair cell regeneration and reinnervation can be dissociated in vivo using molecular genetics and mutant identification in zebrafish. By exploiting simple visual and behavioral assays we will focus on (i) zebrafish mutants that do not regenerate hair cells and (ii) mutants that regenerate, but do not reinnervate nascent hair cells. Cloning of the affected loci in these mutants will identify novel genes, or known genes with novel functions, required for new hair cell formation and reestablishment of synaptic connectivity.