ALPHA-CONOTOXIN MII--SELECTIVE NICOTINIC RECEPTOR PROBE

描述（来自申请人摘要）： 尼古丁通过一个多样化的家庭发挥其许多行为影响 烟碱乙酰胆碱受体尤其是尼古丁引起的 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，分离自捕食性锥 蜗牛Conus magus，阻断α 3-β 2亚型烟碱受体 具有高效力和选择性。α-芋螺毒素MII也选择性地 并有效地阻断尼古丁诱发的多巴胺释放， 体外，因此可能是一个工具，探索一个行为 显著烟碱受体亚型。天然和放射性标记 （[125 I]-alphaCtx MII）α-芋螺毒素MII可用，并将用于 以表征α-芋螺毒素MII结合位点。我们打算测试 抗[3 H]地棘蛙素结合的天然毒素（在α 3 β 2-转染的 细胞和小鼠脑制备物）和烟碱激活的测量 体外[125]alphaCtx MII结合将用于表征 转染的细胞和集中表达的受体，使用生物化学 和放射自显影技术。这些研究旨在定义 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。细胞中表达位点的比较 与大脑中的那些线将确定本地和模型的相似程度 网站是。此外，我们还将尝试开发一套新的 基于α-芋螺毒素MII的探针， 生物素标签插入毒素的一级肽序列这些探针 旨在探索α-芋螺毒素MII结合 网站的组成和结构。拟议的研究将提供 深入了解一种新颖的、天然的尼古丁的性质和重要性， 受体的希望这些知识能让我们更好地理解 尼古丁的中枢介导作用的基础，并可能导致 涉及改进的烟碱药物和/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.