Characterization of the Role of mTOR Signaling in Opiate Action

描述（由申请人提供）：近年来，阿片类药物的医疗和非医疗用途在美国大大增加。这一增长尤其值得注意的是，滥用处方阿片类药物的青少年人数有所增加。尽管这种增加，很少有人知道与慢性阿片类药物使用发生的神经适应。该项目的目标是确定有助于神经适应慢性阿片类药物使用的潜在分子信号，特别是导致多巴胺能腹侧被盖（VTA）神经元大小减少和相应的奖励耐受行为适应的机制。Nestler实验室先前的工作已经建立了一种慢性吗啡范例，该范例以BDNF和PI 3 K/Akt依赖的方式减少VTA中多巴胺能细胞的大小。此外，VTA中PI 3 K/Akt信号传导的变化被发现是产生奖赏耐受性所必需和充分的。该项目旨在确定Akt下游的信号通路调节细胞大小和行为的这种变化。已知影响CNS以及外周中细胞大小的一个突出的信号传导途径是雷帕霉素（mTOR）途径的哺乳动物靶标。我们假设mTOR信号的改变有助于慢性吗啡产生的形态和行为变化。为了确定mTOR信号传导是否响应于慢性吗啡而改变，将用吗啡处理大鼠，并解剖VTA以通过蛋白质印迹分析生化变化。此外，mTOR抑制剂雷帕霉素将用于抑制mTOR信号传导，以确定降低的mTOR信号传导是否可以阻断吗啡诱导的奖励耐受。最后，将使用mTOR信号传导的关键调节剂、脑中富集的ras同系物（Rheb）的野生型和活化和显性负性形式的局部病毒介导的过表达来模拟由慢性阿片剂诱导的变化并阻断慢性阿片剂作用。 鉴于2005年美国最常用的处方药物是阿片类药物，氢可酮与对乙酰氨基酚，并且非处方使用也有所增加，特别是在青少年中，阿片类药物奥施康定和维柯丁的使用大于更多研究的滥用药物，如可卡因，研究阿片类药物对神经生物学和行为的作用是一个重要的局部药物滥用项目。我们工作的目标是了解慢性阿片类药物使用中发生的神经适应，这可能有助于我们更好地理解成瘾倾向以及奖励耐受现象，即药物奖励减少导致剂量增加。

DESCRIPTION (provided by applicant): Both the medical and nonmedical use of opiate drugs have greatly increased in the U.S. in recent years. This increase is especially noteworthy in the increased number of teenagers abusing prescription opiate drugs. Despite this increase, very little is known about the neuroadaptations that occur with chronic opiate use. The goal of this project is to identify the underlying molecular signals that contribute to neuroadaptation to chronic opiate use, specifically the mechanisms that cause a decrease in the size of dopaminergic ventral tegmental (VTA) neurons and the corresponding behavioral adaptation of reward tolerance. Previous work in the Nestler laboratory has established a chronic morphine paradigm that decreases the size of dopaminergic cells in the VTA in a BDNF- and PI3K/Akt-dependent manner. Further, changes in PI3K/Akt signaling in the VTA were found to be necessary and sufficient to produce reward tolerance. This project aims to determine what signaling pathways downstream of Akt modulate this change in cell size and behavior. One prominent signaling pathway that is known to affect cell size in the CNS as well as in the periphery is the mammalian target of rapamycin (mTOR) pathway. We hypothesize that alterations in mTOR signaling contribute to morphological and behavioral changes produced by chronic morphine. To determine whether mTOR signaling is altered in response to chronic morphine, rats will be treated with morphine and the VTA will be dissected for analysis of biochemical changes by western blot. Additionally, the mTOR inhibitor, rapamycin, will be used to inhibit mTOR signaling in order to determine whether decreased mTOR signaling can block morphine-induced reward tolerance. Finally, local viral-mediated overexpression of wildtype and activating and dominant-negative forms of a key modulator of mTOR signaling, ras homolog enriched in brain (Rheb) will be used to mimic changes induced by chronic opiates and to block chronic opiate effects. Given that the most highly prescribed drug in the U.S. in 2005 was an opiate, hydrocodone with acetaminophen, and that the non-prescription use has also increased, especially in adolescents where use of the opiates Oxycontin and Vicodin is greater than more studied drugs of abuse such as cocaine, investigating the action of opiate drugs on neurobiology and behavior is an important and topical drug abuse project. The goal of our work is to understand the neuroadaptations that occur with chronic opiate use, which may help us to better understand both the addiction liability as well as the phenomenon of reward tolerance, decreased reward for drug leading to escalation of dose.