THE ROLE OF AGRIN IN THE DEVELOPMENT OF CENTRAL NEURONS

描述（由申请人提供）：在过去的二十年中，大量的注意力集中在哺乳动物中枢神经系统突触发生的研究上。许多这样的研究都是由这样的信念驱动的，即通过更好地了解中枢神经元突触形成的基本机制，可以实现与几种神经退行性疾病相关的突触损失的诊断和预防。尽管这些机制还没有被完全理解，但人们很容易推测，在神经肌肉连接形成中起关键作用的蛋白质，如agrin，也可能在神经元之间突触接触的形成中起重要作用。最近，我们已经表明，在海马神经元轴突和树突的延伸率以及突触的形成，agrin调节差异。然而，这些神经元中的agrin受体尚未被确定。本文提出的实验旨在验证以下假设：ror蛋白，两种被认为是“孤儿受体”的酪氨酸激酶受体，可以介导agrin对中枢神经元早期发育的影响。结合Western blot分析、免疫细胞化学、RT-PCR、零突变生成、结合试验和免疫沉淀等技术分析：1)中枢神经元中ror 1和ror 2的表达模式和定位；2)这些酪氨酸激酶受体在神经突伸长和突触形成中的作用；3) ror 1和ror 2在agrin信号通路中的参与。

DESCRIPTION (provided by applicant): In the past two decades, a great deal of attention has been directed to the study of synaptogenesis in the mammalian central nervous system. Many of these studies are driven by the belief that the diagnosis and the prevention of synapse loss associated with several neurodegenerative diseases can be achieved through a better understanding of the basic mechanisms of synapse formation in central neurons. Although these mechanisms are not completely understood, it is tempting to speculate that proteins, such as agrin, that play a key role in the formation of the neuromuscular junction might also play an important role in the formation of synaptic contacts between neurons. Recently, we have shown that agrin differentially regulates the rates of axonal and dendritic elongation as well as synapse formation in hippocampal neurons. However, the agrin receptor(s) has yet to be identified in these neurons. The experiments proposed here are intended to test the following hypothesis: ror proteins, two tyrosine kinase receptors considered "orphan receptors", could mediate the effects of agrin on early stages of development in central neurons. A combination of techniques including: Western blot analysis, immunocytochemistry, RT-PCR, the generation of null mutations, binding assays, and immunoprecipitation will be used to analyze: 1) the pattern of expression and localization of ror 1 and ror 2 in central neurons; 2) the role of these tyrosine kinase receptors in neurite elongation and synapse formation; and 3) the participation of ror 1 and ror 2 in the agrin-signaling pathway.