Opioids and Conditional Amygdala NMDA Receptor Knockout

阿片类药物和条件性杏仁核 NMDA 受体敲除

• 批准号: 7033079
• 负责人: MICHAEL J GLASS
• 金额: $ 14.27万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-15 至 2008-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7033079
• 关键词: NMDA receptors; amygdala; behavior test; dendrites; drug addiction; drug tolerance; drug withdrawal; electron microscopy; gene expression; genetically modified animals; immunocytochemistry; in situ hybridization; integrase; intracellular transport; laboratory mouse; morphine; neurophysiology; nitric oxide synthase; opiate alkaloid; protein localization; protein transport; transfection /expression vector

DESCRIPTION (provided by applicant): This revised application is being submitted to provide Dr. Michael Glass, with four years of mentored training in molecular neuropharmacological and stereological methods to study drug abuse under the primary sponsorship of Dr Charles Inturissi and co-sponsorship of Dr. Virginia Pickel. This project will allow the candidate to engage in full-time research which will be facilitated by his appointment to tenure-track assistant professor at the Weill Medical College of Cornell University. The projects in this proposal will address the role of amygdala gene expression of the essential NMDA-NR1 (NR1) receptor subunit in the development and expression of opiate dependence by creating spatial-temporal knockouts using a specific and versatile form of Cre-loxP technology. This Cre-loxP system has been successfully used to achieve conditional NR1 gene knockout in spinal cord (i.e. spatial knockout) in adult animals (i.e. temporal knockout) with changes in injury-induced pain. The viability of this conditional gene knockout approach to achieve significant reductions of NR1 gene expression in a specific brain region of adult animals, and its consequences for behavior, neural activation, and protein targeting in dendritic spines has never been determined. This proposal will integrate innovative methods for conditional gene knockout and high resolution quantitative electron microscopic methods along with established behavioral paradigms to achieve the following aims: 1). produce NR1 knockout limited to the amygdala of adult mice, 2). examine the effects of conditional NR1 knockout in the amygdala on the development and expression of morphine dependence and tolerance, 3). determine the effects of conditional NR1 knockout in the amygdala on neural activation induced by opiate withdrawal, 4). examine the effects of amygdala NR1 knockout on the cellular targeting of proteins that are structurally and functionally linked to NR1, including the NMDA-R2 (NR2) subunit and neuronal nitric oxide synthase (nNOS) in dendritic spines. These studies will provide important new information on the role of amygdala NR1 subunit gene expression in the development of opiate dependence, and provide novel insights into the mechanisms mediating neural plasticity induced by chronic opiate use with implications for maintenance of drug use and relapse.

描述（由申请人提供）： 提交修订后的申请是为了向 Michael Glass 博士提供四年的分子神经药理学和体视学方法的指导培训，以研究药物滥用，该培训由 Charles Inturissi 博士的主要赞助和 Virginia Pickel 博士的共同赞助。该项目将允许候选人从事全职研究，这将通过他被任命为康奈尔大学威尔医学院的终身教授助理教授来促进。该提案中的项目将通过使用特定且多功能形式的 Cre-loxP 技术创建时空敲除，解决重要 NMDA-NR1 (NR1) 受体亚基的杏仁核基因表达在阿片依赖的发展和表达中的作用。该Cre-loxP系统已成功用于在成年动物脊髓中实现条件性NR1基因敲除（即空间敲除）（即时间敲除），从而改变损伤引起的疼痛。这种条件性基因敲除方法能否显着降低成年动物特定大脑区域的 NR1 基因表达，及其对行为、神经激活和树突棘蛋白靶向的影响尚未确定。该提案将整合条件基因敲除的创新方法和高分辨率定量电子显微镜方法以及已建立的行为范式，以实现以下目标：1）。产生仅限于成年小鼠杏仁核的 NR1 敲除，2)。检查杏仁核中条件性 NR1 敲除对吗啡依赖性和耐受性的发展和表达的影响，3)。确定杏仁核中条件性 NR1 敲除对阿片戒断引起的神经激活的影响，4)。研究杏仁核 NR1 敲除对结构和功能上与 NR1 相关的蛋白质的细胞靶向的影响，包括树突棘中的 NMDA-R2 (NR2) 亚基和神经元一氧化氮合酶 (nNOS)。这些研究将为杏仁核NR1亚基基因表达在阿片依赖发展中的作用提供重要的新信息，并为介导长期阿片使用引起的神经可塑性的机制提供新的见解，这对维持药物使用和复发具有影响。