Morphogenesis of the peripheral sensory nervous system

描述(申请人提供)：有许多临床情况表现为面部、头部和/或内脏的感觉信号改变，例如糖尿病、艾滋病毒/艾滋病和疱疹引起的周围神经疾病、三叉神经痛、各种遗传性感觉神经疾病、创伤或手术结果引起的神经疾病，以及其他。我们最近设计了一个转基因鱼类品系，它在颅感觉神经节(CSG)的神经元中表达一种EGFP标记蛋白。产生的绿色荧光蛋白是可溶的，并与其外围和中心突起一起填充细胞体。这种表达开始于这些神经元形成神经节并延伸其轴突的时候，并保持到至少三周的年龄。我们建议将这些转基因FISH用于基于乙基亚硝脲的突变筛选，以努力识别参与以下三个过程中的任何一个的基因：单个脑神经节的形成，不同神经节对特定靶点的神经支配，以及同一神经节内单个神经元对不同靶点的神经支配。本申请中提出的工作有两个目的：(1)将ENU突变基因导入TG(3.2：：eGFPGR)系，并使用F3隐性筛选范式来鉴定产生CSG发育和连通性缺陷的胚胎的携带者家族；(2)鉴定已鉴定的突变体。我们已经开始了使用少量航母家族的初步筛选(21架)。到目前为止，我们已经从筛选的11个家系中鉴定出两个突变体，它们的CSG在96HPF中具有发育异常。等位基因SL14表现出三叉神经节的破坏形成，而具有正常的鳃上神经节，而等位基因SL19不正确地靶向颧上神经节的中心和周围投射，而三叉神经节似乎没有改变。这个R21提案旨在将我们的筛查大小增加到150-200个家族(基因组)，以产生覆盖CSG形成的广泛机制的突变体。这一提议的成功结果还将加强Voigt实验室(SLU)和阿佩尔实验室(Vanderbilt)之间的新合作，前者专注于外周神经元和信号问题，并将斑马鱼作为模型，后者是一家老牌斑马鱼实验室，对中枢和外周神经元髓鞘形成的潜在机制感兴趣。公共卫生相关性全世界数百万人由于周围感觉神经病而遭受健康和生活质量下降的问题。目前，此类疾病的治疗因缺乏临床有用的工具而受到限制。该项目的主要目标是通过识别与感觉神经系统正常发育有关的基因以及潜在的治疗靶点，为开发治疗这些疾病的新方法提供跳板。

DESCRIPTION (provided by applicant): There are numerous clinical conditions in which altered sensory signaling from the face, head and/or viscera are manifested, e.g. peripheral neuropathies arising from diabetes, HIV/AIDS and herpes, trigeminal neuralgia, the various hereditary sensory neuropathies, neuropathies arising from trauma or surgical outcomes, and others. We have recently engineered a transgenic fish line that expresses an eGFP marker protein in neurons of the cranial sensory ganglia (CSG). The eGFP produced is soluble and fills the cell body together with its peripheral and central projections. This expression begins at the time these neurons are forming the ganglia and extending their axonal processes, and is maintained through at least three weeks of age. We propose using these transgenic fish in a ethylnitrosourea-based mutagenesis screen in an effort to identify genes involved in any of the following three processes: formation of the individual cranial ganglia, innervation of specific targets by different ganglia, and innervation of different targets by individual neurons within the same ganglia. There are two aims for the work proposed in this application: (1) to introduce ENU mutated genes into the TG(3.2::eGFPGR) line and use an F3 recessive screening paradigm to identify carrier families that produce embryos exhibiting defects in CSG development and connectivities and (2) to characterize the identified mutants. We have initiated a preliminary screen using a small number of carrier families (21). To date, we have already identified two mutants (out of 11 families screened) that have developmental abnormalities in their CSG by 96 hpf. Allele sl14 exhibits disrupted formation of the trigeminal ganglia while having normal epibranchial ganglia, whereas allele sl19 has improper targeting of the central and peripheral projections of the epibranchial ganglia while the trigeminal ganglia appear unaltered. This R21 proposal is designed to increase the size of our screen to 150-200 families (genomes) in order to generate a body of mutants that cover the wide spectrum of mechanisms responsible for CSG formation. A successful outcome of this proposal will also strengthen a nascent collaboration between the Voigt lab (SLU), which is focused on peripheral neurons and signaling issues and is adopting zebrafish as a model, and the Appel lab (Vanderbilt), an established zebrafish lab which is interested in mechanisms underlying the myelination of central and peripheral neurons. PUBLIC HEALTH RELEVANCE Millions of people worldwide suffer degraded health and quality of life issues due to peripheral sensory neuropathies. Currently, treatment of such disorders is limited by the lack of clinically useful tools. The principal goal of this project is to provide a springboard for the development of new approaches to the treatment of these diseases by identifying genes, and thus potential therapeutic targets, involved in the normal development of the sensory nervous system.