Role of Lipid Rafts in Adenylyl Cyclase Sensitization

描述（由申请人提供）：阿片类药物依赖在这些药物的滥用中起着重要作用，是临床关注的问题。如果我们要改善疼痛的治疗，治疗和/或预防阿片类药物滥用，了解依赖的机制是很重要的。急性时，mu-阿片类药物激活Gai/o蛋白来抑制腺苷酸环化酶，但慢性治疗后，腺苷酸环化酶激活的代偿性增加可以在细胞水平上证明其依赖性，称为致敏。该项目的目的是更好地了解细胞水平上依赖性发展的机制，特别是腺苷酸环化酶的代偿性变化是如何通过与质膜微域（称为脂筏）中的其他信号蛋白共定位来调节的。类阿片受体存在于脂筏中，被致敏的腺苷酸环化酶亚型倾向于在脂筏中定位，而一些在致敏中起作用的信号蛋白能够重新定位到脂筏内或脂筏外。为了确定脂筏在mu-阿片介导的腺苷酸环化酶的发育和表达中的作用，我们将：1)检查mu-阿片信号成分在脂筏和非脂筏中的分布，并开发检测其功能的方法。2)从经过μ -阿片激动剂处理的细胞中制备富含脂筏和非脂筏的质膜组分，比较腺苷酸环化酶的致敏性，并确定致敏性的变化是否与信号蛋白进入或离开脂筏的再定位有关。3)确定脂筏是否在不同受体和不同细胞类型的腺苷酸环化酶敏化中起必要作用。在长期暴露于阿片类药物（如吗啡或海洛因）后，神经细胞膜中的信号蛋白（如腺苷酸环化酶）发生变化。这些代偿性变化导致阿片类药物去除后的戒断。为了更好地治疗和预防阿片类药物依赖，我们的目标是了解引起这些变化的机制，特别是与细胞膜微域（脂筏）内外调节信号蛋白的位置有关。

DESCRIPTION (provided by applicant): Opioid dependence plays a major role in abuse of these drugs and is a concern in the clinic. It is important to understand mechanisms underlying dependence if we are to improve the treatment of pain and to treat and/or prevent opioid abuse. Acutely, mu- opioids activate Gai/o proteins to inhibit adenylyl cyclase, but following chronic treatment dependence can be demonstrated at the cellular level by the compensatory increase in adenylyl cyclase activation, referred to as sensitization. The objective of this project is to better understand the mechanisms of the development of dependence at the cellular level, particularly how the compensatory changes in adenylyl cyclase are modulated by co-localization with other signaling proteins in plasma membrane microdomains referred to as lipid rafts. Mu-opioid receptors are found in lipid rafts and the adenylyl cyclase isoforms which are sensitized tend to be localized in lipid rafts, while some signaling proteins which have a role in sensitization are able to relocalize into or out of lipid rafts. To determine the role of lipid rafts in the development and expression of mu-opioid mediated adenylyl cyclase sensitization, we will: 1) Examine the distribution of mu-opioid signaling components in lipid rafts and non-raft fractions and develop assays to measure their function. 2) Prepare lipid-raft enriched and non-raft plasma membrane fractions from cells treated with mu-opioid agonist to compare sensitization of adenylyl cyclase and determine if changes in sensitization are related to relocalization of signaling proteins into or out of lipid rafts. 3) Determine if lipid rafts play a necessary role in adenylyl cyclase sensitization across different receptors and in different cell types. Changes occur in signaling proteins, such as adenylyl cyclase, in membranes of neuronal cells following long term exposure to opioid drugs, such as morphine or heroin. These compensatory changes result in withdrawal on removal of the opioid. To better treat and prevent opioid dependence, the goal is to understand the mechanisms causing these changes, particularly in relation to the location of modulatory signaling proteins in or out of cell membrane microdomains referred to as lipid rafts.