DEVELOPMENT AND FUNCTION OF SYNAPTIC CO-TRANSMISSION

这项申请的目的是加强正在进行的研究， 交感神经节突触共传递生理学， 学习将应用于新研究的分子和生物学方法 神经元的分化。 私家侦探是一名助理教授， 生理学，是在05年的独立研究与RO 1的支持。 良好的大学环境和充足的支持服务 在当地大学社区。 拟议的计划将确保 继续发展P.I.的研究生涯。它将限制增长 非研究相关的活动，并提供资源，以追求新的 方法、实验和合作。 实验将 利用电生理学、解剖学和生物化学的组合， 分离制备物和原代细胞培养物中的方法。 新 需要学习的方法包括核酸分离、北方分析和 原位杂交 该提案的实验目标是确定综合 功能和发育起源的突触，利用缓慢 除了经典的肽能共递质外， 递质乙酰胆碱（ACH）和肾上腺素（EPI）。 促黄体生成素释放激素、 （LHRH）、神经肽Y（NPY）、降钙素基因相关肽（CGRP）和 P物质（SP），将在牛蛙的交感神经系统进行分析。 尽管这些肽在许多细胞中与其他递质共表达， 周围和中枢神经系统的区域，功能性 突触共传递的动力学和个体发生在很大程度上是未知的。 牛蛙交感神经节表达多种突触机制 但它们可以在细胞和分子水平上进行分析， 程度. 在腰神经节中，存在3个交感神经亚类， 神经元可以通过其功能、电生理学来区分 特性，与脊髓和外周靶点的连接， 以及通过它们的毒蕈碱胆碱能受体的表达， 神经肽 在血管交感神经C系统中，LHRH和ACH是 由节前神经元共同释放并引发3种突触后电位 在共表达EPI和NPY样的神经节细胞亚群中， 免疫反应性。 本提案的第一个具体目标是测试 神经节和终末器官突触共传递假说 该回路增强了血管收缩的动态范围， 是由不同的节前刺激引起的。 的 第二个具体目的是检验CGRP是共同释放的假设 与ACH通过节前B神经元，并产生营养作用， B神经元兴奋性突触后电位。 第三个目标是 在蝌蚪发育过程中建立时间表， 神经肽基因及其产物的表达。 使用组织 文化，该项目的最终目标将是测试假设， 神经肽Y的表达受细胞外的分子控制， 环境和/或细胞相互作用，并且这些因素还 控制血管神经元的电生理分化。

The objectives of this application are to intensify ongoing research on the physiology of synaptic cotransmission in sympathetic ganglia and to learn molecular and biological methods that will be applied in new studies of neuronal differentiation. The P.I. is an assistant professor of Physiology and is in the 05 year of independent research with RO1 support. An excellent collegial environment and adequate support services exist within the local University community. The propose plan will insure continued development of the P.I.'s research career. It will limit growth of non-research related activities and provide resources to pursue new methodologies, experiments and collaborations. The experiments will utilize a combination of electrophysiological, anatomical and biochemical methods in isolated preparations and in primary cell cultures. The new methods to be learned include nucleic acid isolation,Northern analysis and in situ hybridization. The experimental goals of the proposal are to determine the integrative function and the developmental origins of synapses that utilize slowly acting peptidergic co-transmitters in addition to the classical transmitters acetylcholine (ACH) and epinephrine (EPI). The roles of 4 neuropeptides, luteinizing hormone releasing hormone (LHRH), neuropeptide Y (NPY), calcitonin gene-related peptide (CGRP) and substance P (SP), will be analyzed in the sympathetic system of bullfrog. Although these peptides are co-expressed with other transmitters in many regions of the peripheral and central nervous system, the functional dynamics and ontogeny of synaptic co-transmission is largely unknown. Bullfrog sympathetic ganglia express a diversity of synaptic mechanisms and yet they are accessible to analysis at the cellular and molecular levels. In the lumbar ganglia, there exist 3 subclasses of sympathetic neurons that can be distinguished by their functions, electrophysiological properties, connectivity with the spinal cord and with peripheral targets, and by their expression of muscarinic cholinergic receptors and neuropeptides. In the vasomotor sympathetic C system, LHRH and ACH are co-released by preganglionic neurons and elicit 3 postsynaptic potentials in a subset of ganglion cells that co-express EPI and NPY-like immunoreactivity. The first specific aim of this proposal is to test the hypothesis that co-transmission at ganglionic and end-organ synapses in this circuit enhances the dynamic range of vascular contractions that can be elicited by different patterns of preganglionic stimulation. The second specific aim is to test the hypothesis that CGRP is co-released with ACH by preganglionic B neurons and produces trophic effects and an excitatory postsynaptic potential in B neurons. The third goal is to establish the timetable during development in tadpoles for cell-specific expression of neuropeptide genes and their products. Using tissue culture, the final goal of the project will be to test the hypothesis that expression of NPY is controlled by molecules in the extracellular environment and/or by cellular interactions, and that such factors also control the electrophysiological differentiation of vasomotor neurons.