Axon Guidance in the Olfactory System

描述（由申请人提供）：嗅觉感觉神经元（OSN）的发生是一个持续的过程，在发育早期开始，并在成人中继续。它需要新产生的轴突成功地穿过固有层和筛板进入中枢神经系统。神经发育事件的中断是导致广泛疾病的原因，包括自闭症、脆性X染色体、无脑畸形和卡尔曼综合征。我的最终目标是提高我们对驱动和调节轴突生长的机制的理解。具体来说，我的目标是在这个建议是研究嗅觉通路的发展和测试的假设，Dishevelled（Dvl）蛋白影响轴突的指导和突触的形成。轴突导向信号、细胞粘附分子和气味受体诱导的活性都与OSN轴突最终精确靶向特定肾小球有关。然而，OSN轴突从嗅上皮（OE）向嗅球（OB）延伸轴突的机制仅部分了解，并且经常有争议。类似地，参与在OB中的OSN和投射神经元之间建立突触的分子也没有很好地理解。Wnt分泌的分子通过卷曲（Fz）受体起作用，涉及许多过程，包括轴突引导和突触形成。我们最近在嗅觉系统中发现了几种这样的分子，其模式暗示了OSN轴突生长中的作用。Fz活化后信号通路中的一个关键分子是Dvl。该方案包括对嗅觉系统中所有三种Dvl（Dvl-1、Dvl-2和Dvl-3）分子和一些候选下游信号分子（例如RhoA、Daam-1、ROCK）的表达模式的系统表征。将研究蛋白质（Western印迹和免疫组织化学）和mRNA（RT-PCR和原位杂交）表达。Dvl分子的作用将通过子宫内电穿孔注射表征的沉默RNA（siRNA）来评估，我最近优化了该方法以标记OE中的OSN。该提议的结果将促进我们对Wnt-Fz信号通路（包括Dvls）在OSN轴突中的作用的理解。反过来，这将对理解嗅觉系统中的轴突引导和突触形成产生重大影响，并广泛地在中枢神经系统的其他区域。

DESCRIPTION (provided by applicant): Olfactory sensory neuron (OSN) genesis is an ongoing process that begins early in development and continues in adults. It requires that newly generated axons successfully navigate through the lamina propria and the cribiform plate, into the central nervous system. Disruptions in neurodevelopmental events are responsible for a wide range of diseases, including Autism, Fragile X, Lissencephaly, and Kallman's syndrome. My ultimate goal is to improve our understanding of the mechanisms that drive and regulate axonal outgrowth. Specifically, my goal in this proposal is to study the development of the olfactory pathway and test the hypothesis that Dishevelled (Dvl) proteins influence both axon guidance and synapse formation. Axon guidance cues, cell adhesion molecules, and odor receptor induced activity have all been implicated in the precise final targeting of OSN axons to specific glomeruli. However, the mechanisms used by OSN axons to extend an axon from the olfactory epithelium (OE) toward the olfactory bulb (OB) are only partially understood, and often controversial. Similarly, the molecules involved in establishing the synapses between the OSNs and the projection neurons in the OB are not well understood. Wnt secreted molecules, acting through Frizzled (Fz) receptors are implicated in many processes, including axon guidance and synapse formation. We recently found several of these molecules in the olfactory system, in a pattern suggestive of a role in OSN axon outgrowth. One key molecule in the signaling pathway after Fz activation is Dvl. This proposal includes a systematic characterization of the expression patterns of all three Dvl (Dvl-1, -2 and -3) molecules and some candidate downstream signaling molecules (e.g. RhoA, Daam-1, ROCK) in the olfactory system. Both protein (Western blot and immunohistochemistry), and mRNA (RT-PCR and in situ hybridization) expressions will be studied. The role of Dvl molecules will be assessed by injecting a characterized silencing RNA (siRNA) by in utero electroporation, which I recently optimized to label OSNs in the OE. The results from this proposal will promote our understanding of the role of the Wnt-Fz signaling pathways, including Dvls, in OSN axons. This, in turn, will have a significant impact on understanding axon guidance and synapse formation in the olfactory system, and broadly in other regions of the central nervous system.