MOLECULAR MECHANISMS: COCAINE & OTHER UPTAKE INHIBITORS

分子机制：可卡因

• 批准号: 3461306
• 负责人: S Paul BERGER
• 金额: $ 9.61万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-30 至 1996-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3461306
• 关键词: affinity labeling; antiadrenergic agents; binding proteins; brain mapping; catecholamine inhibitor; chemical group; chemical structure function; cocaine; corpus striatum; dopamine; drug abuse; gel electrophoresis; gel filtration chromatography; laboratory rat; molecular weight; neurotransmitter metabolism; nucleus accumbens; photolysis; piperazines; protein denaturation; protein structure; radiotracer; sodium; transport proteins; ultraviolet radiation

The ability of cocaine to inhibit dopamine reuptake in areas of the brain such as the nucleus accumbens where dopamine is reinforcing, is believed to be a major mechanism contributing to the abuse potential of cocaine. Although we and others have characterized the binding of cocaine and other tritiated dopamine reuptake inhibitors to the dopamine reuptake transporter protein less is known about the molecular mechanism by which the binding of a ligand such as cocaine to a sodium dependent transporter results in inhibition of reuptake. Recently, we synthesized a tritiated azido diphenylpiperazine derivative 1-[2bis(phenyl)methoxy]ethyl- 4-[3-(3'-azidophenyl)-{2,3-3H]propyl}piperazine (3azido[3H]GBR-12935.), which can be covalently incorporated (in a sodium dependent manner and with appropriate pharmacological specificity) into a single striatal 80 kDa polypeptide of the dopamine reuptake transporter protein (as detected by sodium dodecyl sulfate-polyacrylamine gel electrophoresis and fluorography). This grant proposes a series of studies that will attempt to determine the molecular composition of the dopamine transporter as well as identify regions of the transporter protein to which uptake inhibitors bind. Specific questions to be addressed are whether there are different regional isoforms of the dopamine transporter, whether several dopamine reuptake inhibitor share, a common molecular mechanism and whether dopamine reuptake inhibitors interact with the substrate (dopamine) binding site on the transporter. Methodologies proposed include further covalent incorporation studies with 3-azido [3H]GBR-12935 and comparison to covalent incorporation studies of other pharmacological probes (BTCP, metaphit, cocaine isothiocyanate, xylamine). Radiation inactivation studies are proposed to determine whether cocaine and other dopamine reuptake inhibitors interact with a protein of similar molecular weight (preliminary data suggests this is not the case). Finally chemical modification / inactivation studies are proposed to identify, compare and contrast important functional groups for the binding of cocaine and other dopamine reuptake inhibitors.

可卡因抑制大脑区域多巴胺再摄取的能力 例如，多巴胺正在加固的伏隔核，相信 成为有助于可卡因滥用潜力的主要机制。 尽管我们和其他人的特征是可卡因的结合 对多巴胺再摄取的其他多巴胺再摄取抑制剂 转运蛋白较少有关分子机制已知 配体（例如可卡因）与钠依赖转运蛋白的结合 导致抑制再摄取。 最近，我们合成了一个曲折的偶氮二苯基哌嗪衍生物 1- [2Bis（苯基）甲氧基]乙基 - 4- [3-（3'-二苯基） - {2,3-3H]丙基}哌嗪（3azido [3H] GBR-12935。）， 可以共价掺入（以钠依赖的方式和 具有适当的药理特异性）成单个纹状体80 多巴胺再摄取转运蛋白的KDA多肽（如检测到的 通过十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳和 荧光图）。 该赠款提出了一系列研究，将尝试 确定多巴胺转运蛋白的分子组成为 以及识别摄取的转运蛋白的区域 抑制剂结合。 要解决的具体问题是是否存在 是多巴胺转运蛋白的不同区域同工型，是否 几种多巴胺再摄取抑制剂共享，这是一种常见的分子机制 以及多巴胺再摄取抑制剂是否与底物相互作用 （多巴胺）在转运蛋白上的结合位点。 提出的方法 包括3-齐达多[3H] GBR-12935的进一步共价结合研究 并与其他药理的共价结合研究进行比较 探针（BTCP，隐喻，可卡因异硫氰酸盐，木胺）。 辐射 提出了灭活研究，以确定可卡因和其他 多巴胺再摄取抑制剂与类似分子的蛋白质相互作用 重量（初步数据表明事实并非如此）。 最后 提出了化学修饰 /灭活研究，以识别， 比较和对比重要的官能团的结合 可卡因和其他多巴胺再摄取抑制剂。