Component for Institution # 264313

对药物滥用和成瘾的研究需要多学科的方法。生物化学 药理核心为不进行生化药理研究的研究人员提供服务 化验。该核心提供的技术允许询问与药物相关的受体 虐待和他们发出的信号。大多数使用核心的研究人员要么是行为科学家，要么是 化学家。核心提供的服务对这些调查人员来说很有价值，扩大了他们的 进行了大量研究，并出版了许多出版物。核心还提供了赠款的初步数据 申请和几笔美国国立卫生研究院的补助金被授予。核心所拥有的滥用药物受体 迄今调查的包括阿片受体(MOPR、DOPR、KOPR、NOPR)、大麻素(CB1和 受体、烟碱胆碱能受体(NAChRs)以及多巴胺和西格玛受体。在 在请求的资金期限内，核心将执行以下服务：(1)与放射性配体结合 上述受体与药物滥用有关，必要时与其他相关；(2)G 蛋白偶联受体激活，包括cAMP水平，[35S]GTPγS结合，p44/42 MAP 激酶磷酸化，β-arrestin募集；(3)确定GPCRs的激动剂偏倚。 [35S]GTPγS结合或cAMP和β-arrestin募集与CRISPR细胞株；(4)放射自显影 与阿片和多巴胺受体结合的放射性配基和(5)阿片激动剂的放射自显影 促进脑切片中[35S]GTPγ与S的结合；(6)免疫印迹 在培养细胞和啮齿类动物脑中的磷酸化KOPR； 培养细胞中的KOPR和CB1以及小鼠脑中的KOPR；(8)提供纯化的抗体 向研究人员提供MOPR、KOPR和磷酸KOPR；(9)对其他实验室的人员进行培训。为 在要求的赠款期限内，核心将与美国国立卫生研究院资助的至少15个合作项目 全国各地的调查人员。创新之处在于拟议的研究将涉及广泛的 与药物滥用有关的各种问题，包括说明对青少年和成年人的影响 长期接触吗啡、可卡因或尼古丁对MOPR、多巴胺和/或nAChR表达的影响 和/或分别在成虫和后代中发出信号；寻找不会导致 KOPR磷酸化(可能偏向于G蛋白)；AMP激活的蛋白激酶(AMPK)的作用 MOPR和nAChR表达缺失；筛选CB1受体变构调节剂和 它们如何改变配基结合的测定；CB1和CB2激动剂偏向的研究 通过使用删除了单个G蛋白或阻滞素的CRISPR细胞系来识别受体。这些研究 都是新颖的，核心将扩大这些研究项目超出原来的范围。因此，核心 将对药物滥用研究领域做出重大贡献。

Research on drug abuse and addiction requires multi-disciplinary approaches. The Biochemical Pharmacology Core provides services for researchers who do not perform biochemical pharmacology assays. The techniques provided in this Core allow interrogation of receptors associated with drugs of abuse and their signaling. Most researchers who used the Core are either behavioral scientists or chemists. The services provided by the Core are valuable to these investigators, expanded their research and have resulted in many publications. The Core also has provided preliminary data for grant applications and several NIH grants were awarded. The receptors of drugs of abuse that the Core has investigated to date include opioid receptors (MOPR, DOPR, KOPR, NOPR), cannabinoid (CB1 and CB2) receptors, nicotinic cholinergic receptors (nAChRs) and dopamine and sigma receptors. In the requested funding period, the Core will perform the following services: (1) Radioligand binding to the aforementioned receptors related to drug abuse and others when necessary; (2) Assessment of G protein-coupled receptor (GPCR) activation, including cAMP level, [35S]GTPγS binding, p44/42 MAP kinases phosphorylation, β-arrestin recruitment; (3) Determination of agonist bias for GPCRs using [35S]GTPγS binding or cAMP and β-arrestin recruitment and CRISPR cell lines; (4) Autoradiography of radioligand binding to opioid and dopamine receptors and (5) Autoradiography of opioid agonist- promoted [35S]GTPγS binding in brain sections; (6) Immunoblotting of MOPR, KOPR and phosphorylated KOPR in cultured cells and in rodent brains; (7) Internalization of MOPR, DOPR and KOPR and CB1 in cultured cells and KOPR in mouse brains; (8) Providing purified antibodies against MOPR, KOPR and phospho-KOPR to researchers; (9) Training of personnel of other laboratories. For the requested grant period, the Core will perform at least 15 collaborative projects with NIH-funded investigators across the country. The innovation lies in the proposed studies, which will address a wide variety of questions related to drug abuse, including elucidation of impact of adolescent and adult chronic exposure to morphine, cocaine or nicotine on MOPR, dopamine and/or nAChR expression and/or signaling in adults and offspring, respectively; search for KOPR agonists that do not cause KOPR phosphorylation (likely G protein-biased); effects of AMP-activated protein kinase (AMPK) deletion on expression of MOPR and nAChR; screening for allosteric modulators of CB1 receptors and determination of how they alter ligand binding; and investigation of agonist biases for CB1 and CB2 receptors by using CRISPR cell lines with either individual G proteins or arrestins deleted. The studies are novel and the Core will augment these research projects beyond the original scope. Thus, the Core will make significant contribution to the area of substance abuse research.