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NEUROENDOCRINE PACERS, PEPTIDES, RECEPTORS, ANTIBODIES

神经内分泌起搏器、肽、受体、抗体

基本信息

批准号：
3401795
负责人：
FELIX STRUMWASSER
金额：
$ 28.09万
依托单位：
MARINE BIOLOGICAL LABORATORY
依托单位国家：
美国
项目类别：
财政年份：
1984
资助国家：
美国
起止时间：
1984-08-01 至 1992-12-31
项目状态：
已结题

项目摘要

A broad interdisciplinary research program is proposed that would concentrate on understanding one model neuroendocrine system that commands a reproductive act, the laying of eggs with associated behaviors. This research would combine the talents of individuals at six institutions, each specializing in a different branch of biology and chemistry (neurophysiology, immunology, receptor biochemistry, peptide structure and chemical synthesis). Diverse techniques are used including NMR, X-ray diffraction, the generation of monoclonal antibodies (MAbs) as function- interfering (FI) highly specific reagents, microsequencing and electrical techniques. We choose a marine mollusk, Aplysia, for this work because of certain advantages, over mammalian neuroendocrine systems, and the lessons of the past, that generalizable neurobiological principles are often obtained more efficiently from research on "simple" systems. The command of egg-laying in Aplysia is mediated by the neuropeptide hormone, egg-laying hormone (ELH), synthesized by the bag cell (BC) neurons, and released in a 30 minute discharge of synchronous action potentials before spontaneous egg-laying. This "afterdischarge" can be triggered in vitro by a few seconds of electrical stimulation. This long-lasting discharge is mediated by cAMP-dependent protein phosphorylation modulating at least two independent K+-channels. The research described in this application is concerned with: 1) Identifying, purifying and microsequencing those proteins, cytoplasmic and membrane, that mediate the special electrophysiological properties in two neuroendocrine pacemaker systems. We will develop MAbs as FI-probes, for intracellular injection, in order to identify the proteins mediating pacemaker mechanisms. 2) Determining the structure-activity relationships of the neuropeptide egg-laying hormone (ELH), made by the BCs, by using fragments and modifications of synthetic ELH, in conjunction with behavioral egg-laying assays, electrophysiological assays, and assays involving cubes of isolated gonad. In addition NMR spectroscopy of ELH and interesting analogs in solution and X-ray diffraction of crystals will be performed to gain information on the secondary and tertiary structure of these peptides. 3) Identifying, purifying and microsequencing the ELH receptor from non-neural tissue, the gonad, generating antibodies to it and using the antibody to determine binding sites in the nervous system.

提出了一个广泛的跨学科研究计划，专注于了解一个模型神经内分泌系统命令生殖行为，产卵，相关的行为。这项研究将联合收割机的人才，六个机构的个人，每个机构都专门从事不同的工作，生物学和化学的分支（神经生理学，免疫学，受体生物化学、肽结构和化学合成）。使用各种技术，包括NMR，X射线衍射，产生单克隆抗体（MAb）作为功能- 干扰（FI）高度特异性试剂，微测序和电技术。我们选择了一种海洋软体动物，这种工作是因为某些优势，超过哺乳动物神经内分泌系统，以及过去的教训，一般的神经生物学原理往往获得更多从研究“简单”的系统中有效地获得。在Aesthesia中，产卵的命令是由神经肽激素，产卵激素（ELH），由袋细胞（BC）神经元，并在30分钟的放电中释放自发产卵前的同步动作电位这种“后放电”可以在体外由几秒钟的电刺激这种持久的放电是由 cAMP依赖性蛋白磷酸化调节至少两种独立的K+通道本申请中描述的研究涉及：1）鉴定、纯化和微测序这些蛋白质，细胞质和膜，介导特殊的两种神经内分泌起搏器的电生理特性系统. 我们将开发单克隆抗体作为FI探针，用于细胞内注射，以鉴定介导起搏器的蛋白质机制等 2)确定结构-活性关系神经肽产卵激素（ELH），由BC制造，通过使用合成ELH的片段和修饰，结合行为产卵测定，电生理测定和涉及分离的立方体的测定生殖腺此外，ELH的NMR光谱和感兴趣的溶液中的类似物和晶体的X射线衍射将是以获得关于二级和三级的信息这些肽的结构。 3)识别、纯化和对来自非神经组织的ELH受体进行微测序，性腺，产生抗体，并利用抗体，确定神经系统中的结合位点。