Chronic Ethanol Effects on CNS Opiate Receptors

慢性乙醇对中枢神经系统阿片受体的影响

基本信息

批准号：
6506027
负责人：
LINDA C SALAND
金额：
$ 7.5万
依托单位：
UNIVERSITY OF NEW MEXICO
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-08-01 至 2004-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Chronic ethanol consumption and its continued reinforcement is an ongoing major health and societal problem. In the Central nervous system, the reinforcement of ethanol intake has been linked to enhanced release of endogenous opiates which act at opiate receptors. A non-selective opiate antagonist drug, naltrexone, is approved for humans to reduce ethanol consumption, craving and relapse. Use of the more selective delta receptor antagonist, naltriben, in animals and human trials, suggests that delta receptors are important in use and abuse of alcohol. However, mechanisms by which endogenous opiates and opiate receptors lead to continued ethanol consumption remain unclear. This proposal outlines the use of both histochemical and pharmacologic techniques to study mechanisms which may link chronic ethanol consumption to modulation of the delta opiate receptor, in a rat animal model. The major hypothesis to be tested is that immunoreactive delta opiate receptor expression in the forebrain and midbrain regions is increased during chronic ethanol intake, and the change in expression is accompanied by a reduction in functional coupling of the receptor to G proteins. Changes in delta receptor expression may affect neuronal intracellular signaling pathways in those brain areas to maintain ethanol consumption. Aim #1: To localize and quantify levels of delta opiate receptor subtype in the nucleus accumbens (NA) of the forebrain, and the midbrain ventral tegmental area (VTA), as well as other brain regions, in rats chronically exposed to ethanol. Confocal microscopy will be used, together with computer-assisted quantification, for immunofluorescent-labeled delta opiate receptors, with comparisons to mu receptor expression. We will compare neurons in brain areas of control and ethanol-consuming animals, and in animals which have been withdrawn from ethanol. Aim #2: To determine if chronic ethanol consumption affects functional coupling of delta receptors to second messenger systems in neurons of the NA and VTA, as well as other brain areas, with comparisons to mu receptor coupling. We will use a recently developed method with [35S]-GTPgammaS, whereby receptor-linked G-protein activation can be measured directly with autoradiographic techniques on sections of the brain areas. The direct effects of delta or mu opiate ligands can be examined to determine if they are functionally linked to G-proteins in the neurons of the selected brain areas. By using both quantitative immunohistochemical methods, and functional coupling studies, to examine the interactions of delta receptor ligands in animals after chronic ethanol consumption, it will be possible to determine receptor subtype-specific effects of chronic ethanol. It will also be possible to study the functional changes that may occur in the receptors after withdrawal from consumption. A future potential may be then to target delta receptors for treating chronic alcoholism.

描述（由申请人提供）：长期乙醇消耗及其持续强化是一个持续存在的重大健康和社会问题。在中枢神经系统中，乙醇摄入的加强与作用于阿片受体的内源性阿片的释放增加有关。纳曲酮是一种非选择性阿片拮抗剂药物，被批准用于人类，以减少乙醇消耗、渴望和复发。在动物和人体试验中使用更具选择性的 δ 受体拮抗剂 naltriben，表明 δ 受体在酒精的使用和滥用中很重要。然而，内源性阿片剂和阿片剂受体导致持续乙醇消耗的机制仍不清楚。该提案概述了在大鼠动物模型中使用组织化学和药理学技术来研究可能将长期乙醇消耗与 δ 阿片受体调节联系起来的机制。待测试的主要假设是，在长期摄入乙醇期间，前脑和中脑区域的免疫反应性 δ 阿片受体表达增加，并且表达的变化伴随着受体与 G 蛋白的功能耦合的减少。 δ受体表达的变化可能会影响这些大脑区域的神经元细胞内信号传导途径，以维持乙醇消耗。目标#1：定位和量化长期暴露于乙醇的大鼠前脑伏隔核 (NA) 和中脑腹侧被盖区 (VTA) 以及其他大脑区域中 δ 阿片受体亚型的水平。共聚焦显微镜将与计算机辅助定量一起用于免疫荧光标记的 δ 阿片受体，并与 mu 受体表达进行比较。我们将比较对照动物、消耗乙醇的动物以及戒断乙醇的动物大脑区域的神经元。目标#2：通过与 mu 受体耦合进行比较，确定长期乙醇消耗是否影响 NA 和 VTA 以及其他大脑区域神经元中 δ 受体与第二信使系统的功能耦合。我们将使用最近开发的 [35S]-GTPgammaS 方法，通过放射自显影技术可以直接测量大脑区域切片上受体相关的 G 蛋白激活。可以检查 delta 或 mu 阿片配体的直接作用，以确定它们是否与所选脑区神经元中的 G 蛋白功能相关。通过使用定量免疫组织化学方法和功能耦合研究，检查长期乙醇消耗后动物体内δ受体配体的相互作用，将有可能确定长期乙醇对受体亚型的特异性影响。还可以研究停止食用后受体中可能发生的功能变化。未来的潜力可能是针对δ受体来治疗慢性酒精中毒。