AMYGDALA AMPA GLUR2 DELETION AND OPIOID DEPENDENCE

描述(由申请人提供)：阿片依赖是慢性阿片类药物使用的主要并发症，导致成瘾综合征。除了避免戒断症状外，戒断相关线索的体验也可能在吸毒行为中发挥重要作用。因此，阐明介导长期使用阿片类药物不良后果的神经生物学过程，特别是涉及厌恶学习的作用，可能为管理成瘾性疾病提供关键信息。有证据表明，杏仁中央核(CEA)中的AMPA型谷氨酸受体可能是阿片依赖的重要分子底物。AMPA-GluR2(GluR2)受体亚基是AMPA受体的主要组成部分，决定着钙的通透性，与药物依赖有关，并在CEA中表达。表达AMPA受体的GluR2的活性受NMDA型谷氨酸受体的调节，NMDA型谷氨酸受体本身就是神经和行为适应性的分子底物。尽管有这些发现，但没有直接证据表明CEA中GluR2基因的表达是阿片依赖表达所必需的。这项应用的主要目的是建立AMPA-GluR2受体亚单位的时空缺失，以检验中央杏仁核神经元突触后GluR2表达是阿片类药物戒断诱导的条件性位置厌恶(CPA)所必需的假说。这一假说将通过局部微量注射表达Cre重组酶的嗜神经性腺相关病毒载体在GluR2小鼠的中央杏仁核神经元中产生突触后GluR2的缺失来进行研究。局部基因缺失的可行性将通过光电子免疫组织化学和原位杂交进行检测。CEA GluR2缺失的表型效应将通过行为测量来确定，包括戒断诱导的条件性位置厌恶。这个项目有望加深我们对神经元谷氨酸受体基因表达在阿片依赖中的作用的理解，特别是关于基于AMPA-GluR2的小分子或基因治疗的可能价值。尽管阿片类药物具有临床疗效，但它们的滥用和成瘾责任是公共卫生问题的重要来源。阿片类药物可能通过激活杏仁核谷氨酸受体产生长期效应，杏仁核谷氨酸受体是在神经可塑性以及学习和记忆中发挥重要作用的分子。利用最先进的分子神经药理学技术，这项提议将确定杏仁核中AMPA型谷氨酸受体GluR2亚单位在阿片类药物依赖方面的作用。通过阐明调节依赖不良后果的神经生物学过程，我们可能会提供必要的关键信息，以开发减少这些有害行为的药物干预措施。

DESCRIPTION (provided by applicant): Opioid dependence is a major complication of chronic opioid use that contributes to the syndrome of addiction. In addition to the avoidance of withdrawal symptoms, experience of withdrawal-associated cues may also play an important role in drug taking behavior. Thus, elucidating the neurobiological processes that mediate the adverse consequences of chronic opioid use, particularly involving the role of aversive learning, may provide critical information for managing addictive disease. There is evidence that ionotropic AMPA-type glutamate receptors in the central nucleus of the amygdala (CeA) may be important molecular substrates of opioid dependence. The AMPA-GluR2 (GluR2) receptor subunit is a major component of AMPA receptors, determines calcium permeability, has been implicated in drug dependence, and is expressed in the CeA. The activity of GluR2 expressing AMPA receptors is modulated by NMDA-type glutamate receptors, which are themselves, established molecular substrates of neural and behavioral adaptability. Despite these findings, there is no direct evidence that GluR2 gene expression in the CeA is necessary for the expression of opioid dependence. The primary goal of this application is to develop a spatial-temporal deletion of the AMPA-GluR2 receptor subunit to test the hypothesis that postsynaptic expression of GluR2 in central amygdala neurons is necessary for opioid withdrawal- induced conditioned place aversion (CPA). This hypothesis will be investigated by producing a postsynaptic deletion of GluR2 in central amygdala neurons of floxed GluR2 mice via local microinjection of a neurotropic adenoassociated viral vector expressing Cre recombinase. The viability of local gene deletion will be tested by light and electron immunohistochemistry and in situ hybridization. The phenotypic effects of CeA GluR2 deletion will be determined by behavioral measurements, including withdrawal-induced conditioned place aversion. This project is expected to enhance our understanding of the role of neuronal glutamate receptor gene expression in opioid dependence, particularly with respect to the possible value of AMPA-GluR2 based small molecule or gene therapeutics. Despite the clinical efficacy of opioids, their abuse and addictive liabilities are significant sources of public health problems. Opioids may produce their long-lasting effects by activating amygdala glutamate receptors, molecules that play important roles in neural plasticity, as well as learning and memory. Using state of the art molecular neuropharmacological techniques, this proposal will identify the role of the AMPA-type glutamate receptor GluR2 subunit in the amygdala with respect to opioid dependence. By elucidating the neurobiological processes that mediate the adverse consequences of dependence, we may provide critical information needed to develop pharmacological interventions for reducing these deleterious actions.