ALPHA-CONOTOXIN MII--SELECTIVE NICOTINIC RECEPTOR PROBE

描述(摘自申请人的摘要)： 尼古丁通过一个不同的家庭发挥其许多行为影响 烟碱型乙酰胆碱受体。特别是，尼古丁能唤起 多巴胺的释放被认为在 尼古丁依赖的建立和维持。α-芋螺毒素 MII是最近发现的一种毒素，从捕食性锥体中分离出来 蜗牛圆锥，阻断α3-β2亚型烟碱受体 具有高效力和高选择性。甲芋毒素MII还选择性地 并有效地阻断尼古丁引起的多巴胺释放的措施 在体外，因此可能是一种探索行为的工具 重要的烟碱受体亚型。包括原生的和有无线电标记的 ([125I]-alphaCtx Mii)α-芋螺毒素Mii可用并将被使用 鉴定a-芋螺毒素MII结合位点。我们打算测试一下 抗[~3H]表巴替丁结合的天然毒素(转Alpha3beta2) 细胞和小鼠脑制剂)和烟碱激活的测量 在试管中。[125]将使用AlphaCtx MII绑定来表征两者 转基因细胞和集中表达的受体，使用生化 和放射自显影技术。这些研究旨在确定 中华按蚊α-芋螺毒素MII位点的数量、分布和功能 哺乳动物(小鼠)中枢神经系统。细胞中表达的位点的比较 与大脑中的那些一致将决定本机和模型的相似程度 网站是。此外，我们将尝试开发一套进一步的 以α-芋螺毒素MII为基础的可光激活和 生物素标记到毒素的主肽序列中。这些探头 被设计成允许探索α-芋螺毒素MII结合 网站的组成和结构。拟议的研究将提供 洞察一种新颖的本土尼古丁的性质和重要性 受体。希望这些知识能让我们更好地理解 尼古丁的中枢调节作用的基础，并可能导致 用于开发改进的尼古丁药物和/或治疗 尼古丁依赖。

DESCRIPTION (from applicant's abstract): Nicotine exerts many of its behavioral effects through a diverse family of nicotinic acetylcholine receptors. In particular, nicotine evoked dopamine release is thought to play an important role in the establishment and maintenance of nicotine dependence. alpha-Conotoxin MII is a recently-discovered toxin, isolated from the predatory cone snail Conus magus, which blocks alpha3-beta2subtype nicotinic receptors with high potency and selectivity. Alpha-Conotoxin MII also selectively and potently blocks measures of nicotine evoked dopamine release in vitro, and may thus be a tool with which to explore a behaviorally significant nicotinic receptor subtype. Both native and radiolabeled ([125I]-alphaCtx MII) alpha-conotoxin MII are available and will be used to characterize a-conotoxin MII bindings sites. We intend to test the native toxin against [3H]epibatidine binding (at alpha3beta2-transfected cell and mouse brain preparations) and measures of nicotinic activation in vitro. [125]alphaCtx MII binding will be used to characterize both transfected cell and centrally expressed receptors, using biochemical and autoradiographic techniques. These studies are intended to define the numbers, distribution and function of alpha-conotoxin MII sites in the mammalian (mouse) CNS. Comparison of the sites expressed in the cell line with those in brain will determine how similar the native and model sites are. In addition, we will attempt to develop a further set of probes based on alpha-conotoxin MII incorporating photoactivatable and biotinyl tags into the toxin's primary peptide sequence. These probes are designed to allow the exploration of alpha-conotoxin MII bindings sites' composition and structure. The proposed studies will provide insights into the nature and importance of a novel, native nicotinic receptor. Hopefully, this knowledge will lead to a better understanding of the basis (of) nicotine's centrally mediated effects, and may lead to the development of improved nicotinic drugs and/or treatments for nicotine dependence.