CLONING AND CHARACTERIZATION OF CANNABINOID RECEPTORS

大麻是目前滥用最广泛的街头毒品。 但 大麻素受体系统在健康和 疾病包括使用大麻素作为止痛剂，止吐剂， 癌症患者，癫痫的抗惊厥药和抗青光眼药， 以及免疫调节剂。 这些不同的药理作用 大麻素暗示了受体亚型的可能性。 两 大麻素受体已被确定的日期;一个位于 主要在中枢神经系统（CB 1），而另一个是 在外周组织中表达（CB 2）。 该项目的目标是 描述大麻素受体亚型的药理学特征， 以阐明受体在体内的作用。 我们将稳定地使用 表达个体cDNA克隆的转染细胞系 受体亚型以提供纯的、同质的受体群体 用于筛选候选药物 在具体的目的1中，结合和信号传导 表达CB 1或CB 2的转染细胞的转导特性 将表征大麻素受体亚型。 这些研究将 识别受体亚型选择性配体，其区分 受体亚型的结合和功能测定。 分子 激动剂与大麻素受体结合的机制不是 知道的 具体目标2将检查CB 1或CB 2的哪些结构域 受体识别配体并激活第二信使 系统. 这些研究将通过定点诱变进行， CB 1/CB 2嵌合受体的构建，以定位区域 对受体识别至关重要的受体cDNA， 信使功能 在具体目标3中，额外的推定大麻素 受体将通过聚合酶链反应和文库分离 筛选技术 这些研究将使用累积的信息 从目前分离的受体克隆中获得，以分离其他 这些亚型可以为分离另外的 大麻素受体亚型 总之，这些实验描述了 研究大麻素受体功能的分子生物学方法。 这些研究将有助于设计新的治疗策略 涉及大麻素系统。 此外，深入了解分子 大麻素受体的激活机制可能会导致更好的 对人类滥用大麻素的了解。

Marijuana is currently the most widely abused street drug. However, the functional significance of the cannabinoid receptor system in health and disease includes the use of cannabinoids as analgesics, antiemetics in cancer patients, anticonvulsant for epilepsy and as antiglaucoma agents as well as immunomodulatory agents. These diverse pharmacological effects of cannabinoids suggest the possibility of receptor subtypes. Two cannabinoid receptors have been identified to date; one is located predominantly in the central nervous system (CB1), whereas the other is expressed in peripheral tissues (CB2). The goal of this project is to characterize the pharmacology of subtypes of the cannabinoid receptor in order to elucidate the role of the receptors in vivo. We will use stably transfected cell lines which express cDNA clones of the individual receptor subtypes to provide a pure, homogeneous population of receptors for screening drug candidates. In specific aim 1, the binding and signal transduction properties of transfected cells expressing CB1 or CB2 cannabinoid receptor subtypes will be characterized. These studies will identify receptor subtype-selective ligands which discriminate between the receptor subtypes in binding and functional assays. The molecular mechanisms by which agonists bind to the cannabinoid receptors are not known. Specific aim 2 will examine which domains of the CB1 or CB2 receptors recognize the ligands and which activate second messenger systems. These studies will be conducted by site-directed mutagenesis and the construction of CB1/CB2 chimeric receptors in order to map the regions of the receptor cDNAs critical to receptor recognition and second messenger function. In specific aim 3, additional putative cannabinoid receptors will be isolated by polymerase chain reaction and library screening techniques. These studies will use the cumulative information obtained from the currently isolated receptor clones to isolate other subtypes which may provide data for the isolation of yet additional cannabinoid receptor subtypes. In summary, these experiments describe a molecular biological approach to studying cannabinoid receptor function. These studies will facilitate the design of new therapeutic strategies involving the cannabinoid system. Furthermore, insight into the molecular mechanisms of activation of cannabinoid receptors may lead to a better understanding of cannabinoid abuse in humans.