伏隔核多巴胺D3受体与NMDA受体相互作用参与吗啡行为敏化的分子机制

Opioids addiction has very highly relapse and strong drug craving, which is shown as behavioral sensitization in rodents. The molecular mechanisms underlying the behavioral sensitization are one of the hot topics in addiction field. Both dopamine and glutamate receptors systems are the important pathway and transmitters in the synapse plasticity after drug addiction. However, how dopamine-glutamate receptors interaction modulates behavioral sensitization is poorly understood. Our previously study indicated that the D3Rs are involved in morphine induced mice behavioral sensitization. The D3R knock out (D3R KO) mice or mice administered with dopamine D3R antagonist nafadotride could not develop behavioral sensitization to intermittent morphine administration. The D3 KO mice also can reverse acute morphine or NMDAR (N-methyl-D-aspartic acid receptor) antagonist MK-801 induced mice overexercise compared with WT mice. The D3R antagonist nafadotride can reverse the higher expressed p-NR2B in the NAc after the acute morphine administration. Our study and other published literatures indicated that D3Rs and NMDARs may interact to modulate mice behavioral, but the molecular mechanisms in brain remain elusive. Based on our preliminary studies, we intends to use behavioral, molecular biology, pharmacology, and electrophysiology methods to explore the protein-protein interaction between D3Rs and NMDARs and its downstream molecular pathway of behavioral sensitization induced by morphine in mice accumbens nucleus (NAc, core or shell). Our aim is to reveal the molecular mechanisms of morphine sensitization, and to provide new ideas and theoretical basis to explore opioid detoxification drugs.

阿片类成瘾者有很高的复吸率及对毒品的渴求感，在啮齿类动物表现为行为敏化。脑内多巴胺和谷氨酸系统是毒品依赖后引起突触可塑性改变的重要通路和递质。两类递质受体亚型在脑内如何相互调节从而导致行为敏化的分子机制还不清楚。我们前期工作显示D3R参与吗啡行为敏化并调节NMDAR功能，提示D3R与NMDAR相互作用影响动物行为，但细胞分子机制不明。本研究拟在前期研究基础之上，利用行为学、分子生物学、药理学及电生理学等方法，从整体行为水平到亚细胞水平、蛋白质分子水平研究吗啡诱导小鼠行为敏化前后，伏隔核（NAc，core or shell）D3R与 NMDAs之间的相互作用，阐明受体-受体复合物对受体功能的影响以及对其下游Ca2+信号通路中CaMKII及CREB等关键蛋白调控机制，通过药理学手段调节该受体-受体复合物，观察其对动物行为敏化的影响。以期揭示吗啡敏化的分子机制，为发掘阿片类戒毒药物提供新的思路。

毒品成瘾是一种慢性复发性脑疾病，其病因复杂，机理不清，严重危害公共健康和社会安全。本项目主要研究吗啡成瘾中，小鼠大脑内涉及成瘾的中脑边缘多巴胺环路系统中关键核团内多巴胺与谷氨酸受体间相互关系及下游通路的改变。.主要发现：.1.揭示了吗啡成瘾中关键脑区内NMDA受体表达改变。急性吗啡腹腔注射后，伏隔核（Nac）和前额叶皮质（PFc）内pNR2B/NR2B均呈剂量依赖性增高。吗啡诱导位置条件偏爱（CPP）模型中，NR2B参与小鼠CPP的形成，消退及复吸过程，是小鼠吗啡成瘾的关键蛋白。.2.阐明小鼠敏化过程中多巴胺与谷氨酸受体相互作用的方式及位点。药理学研究发现小鼠伏隔核（Nac）区多巴胺D3R通过调节NMDA-NR2BR的表达参与吗啡敏化的形成过程。电生理学研究发现D3R通过调节由NMDA受体介导的兴奋性突触后电流从而参与吗啡敏化行为的行程。免疫共沉淀（co-IP）研究发现吗啡敏化模型中，小鼠纹状体内NR2B与D3R相互作用显著升高，应用D3R拮抗剂，我们发现，小鼠纹状体内NR2B与D3R相互作用出现显著下降，并进一步影响小鼠行为敏化的形成。pull down实验证明NR2B-CT端是与D3R相互作用的作用的位点。.3.研究了吗啡不同成瘾状态下，关键脑区表观遗传学改变。采用吗啡急性给药及吗啡慢性给药诱导的小鼠行为敏化(behavioral sensitization)模型，观察小鼠大脑PFC、NAc和Hip中MeCP2、CREB/p-CREB蛋白及mRNA的表达变化。揭示了吗啡不同成瘾状态下关键核团表观遗传学改变的差异性，和成瘾形成的表观遗传学机制。.4. 提出了毒品成瘾的干预及治疗策略，探讨了毒品成瘾及精神分裂症易感基因。特异性多巴胺D3R拮抗剂；天然黄酮类药物木犀草素Luteolin均可抑制毒品成瘾行为。我们研究了精神分裂症的相关遗传易感基因，发现在CACNG4和 CACNG5基因之间的SNP rs17645023与中国汉族人群精神分裂症密切相关，另外发现CACNG8基因两个SNP单倍型SNPrs10420331, SNPrs11084307与精神分裂症相关。.本项目系统研究吗啡成瘾的分子机制，为发掘阿片类戒毒药物提供新的思路。