基于脑fMRI及DNA甲基化修饰途径探讨电针治疗恒河猴氯胺酮成瘾效应的机制研究

Ketamine is widely abused as a new drug, which is especially popular with young people. Since Ketamine addiction poses health hazard, it is very important to find the effective method of detoxification.Acupuncture has been widely used in the clinical treatment of drug addiction and achieved a certain effect, but its mechanism is still unclear.Most addictive drugs mainly acted upon the mesolimbic system mediated drug reward -- the core parts of the ventral tegmental area and the nucleus accumbens, which results in adaptive changes in the dopamine pathway and thereby leads to addiction.On the basis of our previous studies on drug addiction, this study was designed to establish rhesus monkey models of ketamine addiction by conditioning stimulation of rings and then giving them Electroacupuncture (EA) treatment. The changes of the structure and function of the critical brain regions of ketamine addiction in monkeys were observed and analyzed by brain fMRI scanning, and the effects of EA treatment on these brain regions were observed as well.Moreover, the DNA methylation mediated regulation of gene expression was taken as the starting point,and with the help of the methods and techniques of immunohistochemistry, Western blot, RT-PCR, pyrosequencing and other molecular biology research, this study explored the mechanism of EA treatment of ketamine addiction from the overall organization of cellular and molecular level,which will provide scientific basis for the clinical application of EA in the treatment of drug addiction.

氯胺酮作为新型毒品而被广泛滥用，特别受到青年人的青睐,氯胺酮成瘾危害严重，寻找有效的戒毒方法具有重要意义。针灸已被广泛用于临床治疗药物成瘾且取得一定的疗效，但其作用机制尚不明确。大多数成瘾性药物主要是通过作用于中脑边缘系统介导成瘾药物奖赏效应的核心部位--中脑腹侧被盖区和伏隔核，引起多巴胺通路发生适应性改变，从而导致成瘾。本课题将在前期相关药物成瘾研究的基础上，建立恒河猴氯胺酮成瘾模型，并给予电针治疗；通过脑fMRI扫描观察和分析恒河猴氯胺酮成瘾关键性奖赏脑区的结构及功能活动变化规律，以及电针治疗对这些脑区的影响；并以DNA甲基化修饰介导调节基因表达作为研究切入点，运用免疫组化、Western blot、RT-PCR、焦磷酸测序等分子生物学的方法和技术，从整体-组织-细胞分子水平进行研究，探讨电针治疗氯胺酮成瘾疗效的作用机制，将为临床推广应用电针治疗药物成瘾提供科学依据。

针灸已被广泛用于临床治疗药物成瘾并取得很好的疗效，其能够有效改善戒断症状和防治复吸，但相关作用机制尚未明确。中脑腹侧被盖区(Ventral tegmental area, VTA)-伏隔核(Nucleus Accumbens, NAc)-前额叶皮质(Prefrontal cortex, PFC)构成的神经环路是药物成瘾最重要的神经解剖学基础，本项目正是针对这一神经通路进行深入的研究探索。本研究成功建立了非人灵长类动物恒河猴氯胺酮慢性成瘾模型，并给予电针治疗，运用行为学、BOLD-fMRI技术、代谢组学及免疫组化、Western blot、RT-PCR等分子生物学方法和技术相结合的方法，从整体-组织-细胞分子水平进行研究，观察和分析恒河猴氯胺酮慢性成瘾关键性神经通路相关VTA-NAc-PFC脑区的功能活动变化规律，同时检测上述脑区代谢物变化情况以及多巴胺受体D2(Dopamine Receptor D2, DRD2)和多巴胺转运体( Dopamine Transporter, DAT)、脑源性神经营养因子(Brain Derived Neurotrophic Factor, BDNF)的表达情况，进而探讨氯胺酮慢性成瘾可能的功能连接途径及相关神经分子机制，以及电针刺激对氯胺酮成瘾戒断症状的治疗效果及其相关中枢神经机制。研究结果表明电针治疗能够有效改善氯胺酮成瘾戒断症状；电针治疗可能通过调节VTA-NAc-PFC脑区与相关脑区的脑功能连接，从而改善氯胺酮成瘾导致的相关功能连接损伤及对应的行为学改变；另一方面电针治疗可以通过调节VTA-NAc-PFC脑区相关蛋白和基因的表达，从而改善相关脑区的病理形态及脑功能连接损伤改变，进一步改善氯胺酮成瘾相关戒断症状，这可能是电针治疗氯胺酮成瘾起效的重要作用机制；此外本研究还发现基于氢质子代谢指纹谱结合模式识别方法能有效区分氯胺酮药物组和对照组恒河猴两组间的差异代谢物，从代谢水平上探寻了氯胺酮慢性成瘾的分子基础和机制，将为氯胺酮慢性成瘾的临床防治和治疗提供客观的理论依据和指导。