尼古丁受体和多巴胺受体的相互调节及其在尼古丁成瘾中的作用

Tobacco abuse is the leading cause of preventable morbidity and mortality in the world. Nicotine is the principal addictive component of tobacco smoke and more than 5 million deaths result from nicotine addiction every year. Nicotine dependence is characterized as a chronic, relapsing disorder in which compulsive drug-seeking and drug-taking behaviours persist despite serious negative consequences. Nicotine exerts its reinforcing effects through its action on nicotinic acetylcholine receptors (nAChRs), a heterogeneous family of pentameric ligand-gated ion channels in the mesocorticolimbic dopaminergic (DAergic) system, which projects from the ventral tegmental area (VTA) to the nucleus accumbens, cortical areas, hippocampus and amygdale. Among different nAChRs expressed in this region, previous experiments established the crucial role of α4β2-containing nAChRs (α4β2* nAChRs) in the positive rewarding properties of nicotine. Furthermore, studies have also suggested dopamine D1 receptor (D1R) plays a critical role in nicotine addiction as D1R antagonist significantly reduces nicotine self-administration and nicotine conditioned single-trial place preference. However, the underlying neurobiological mechanisms of these two receptors in nicotine addiction remain largely unclear and further work is required to identify novel smoking cessation targets. We have previously shown that α7 nAchR couples with NMDA receptor and disrupting this coupling blocks cue-induced nicotine reinstatement in the rat model of nicotine self-administration. Interestingly, we recently obtained preliminary data showing that α4 nAchR forms a protein complex with D1R. Activation of α4β2 nAchR significantly decreases D1R stimulated cAMP increase in both HEK-293 cells co-expressing α4β2 and D1R and cultured striatal neurons. These preliminary studies lead to the hypothesis that α4-D1 coupling may play a role in nicotine addiction. We are planning to elucidate the functional crosstalk between these two receptors both in vitro and in vivo to: characterize the protein-protein interaction between α4 nAchR and D1R; investigate the molecular and functional modulation of the observed α4 nAchR to D1R; study whether α4 nAchR-D1R coupling plays a role in nicotine addiction. Understanding the interplay between α4 nAchR and D1R may not only help to reveal the molecular mechanisms underlying nicotine addiction but also create a unique opportunity to develop novel therapeutic targets for treatment of nicotine addiction.

尼古丁是烟草成瘾主要的强化组分，尼古丁通过作用于中脑边缘多巴胺能系统中的尼古丁受体（nAchR）来完成其对成瘾行为的激活和强化作用。前期的研究确立了包含α4β2亚基的nAchR在尼古丁强化作用中的关键作用。此外，研究还表明多巴胺D1受体（D1R）在尼古丁成瘾中起到重要作用，如D1受体的拮抗剂显著降低尼古丁的自我给药和尼古丁条件性位置偏爱。迄今为止，这两种受体在尼古丁成瘾机制中的作用细节尚不清楚。我们发现α4 nAchR和D1R存在偶联，激活α4β2可以降低共转染细胞和原代纹状体神经元中D1R诱发的cAMP增强。本项目拟研究α4β2和D1R偶联的生化和分子生物学机制，α4β2调节D1R介导的细胞生物学功能，以及二者偶联在尼古丁成瘾中的作用，从而阐明α4β2 nAchR和D1R在尼古丁成瘾中的作用和机制，提供尼古丁成瘾治疗新策略。

多巴胺（DA）是中枢神经系统的重要神经递质，主要参与运动、动机行为，学习和记忆以及情感等功能。多巴胺通过D1 和D2 两种亚型受体发挥对腺苷酸环化酶和环磷酸腺苷（cAMP）水平的激活或者抑制作用。近年来的研究发现，除经典信号途径外，受体与众多胞内蛋白形成结构性、调节性、信号相关性的蛋白复合物（DRIP）。 DRIP不仅调节受体信号传导，还有助于受体运输和稳定性以及多巴胺受体信号传导的形成。迄今为止，已发现与DA相互作用的蛋白包括神经递质受体，转运蛋白，离子通道，细胞内信号蛋白，细胞骨架蛋白，蛋白激酶和适配体/伴侣蛋白。.我们既往的研究表明，多巴胺D1/D2受体复合物在抑郁症，D2/DISC1受体复合物在精神分裂症的发病和治疗中起重要作用。在本研究中，我们发现4 烟碱受体亚型（nAchR）和D1R也存在直接蛋白偶联，二者的相互作用通过D1受体C末端t2的15个氨基酸片段介导 （t2-1）。使用人工合成的干扰性多肽可以抑制尼古丁依赖的位置偏爱行为，后者是尼古丁成瘾的主要动物模型之一，表明二者的相互作用在尼古丁成瘾中起到重要的作用。细胞水平上的研究表明，t2-1片段的多肽在打断4-D1二者复合物的同时，阻断了D1受体介导的cAMP增高，而4 nAchR的激活对D1受体介导的cAMP功能没有影响。.α4受体亚型不仅存在于中脑被盖(VTA)的突触前膜，同时存在于伏隔核(NAc)的突触后GABA能小多棘神经元。α4受体亚型的激活如何影响这一多巴胺能成瘾通路成为一个关键的问题。既往在海马神经元的研究表明，D1受体的激活降低了D1与NMDA受体NR1a亚基通过t2的偶联, 而对t3介导的D1/NR2a没有影响。功能上，t3干扰肽阻断了D1激活所致的NMDA细胞毒作用降低。而t2多肽解聚D1/NR1a, 促进NR1a/CaMKII的偶联和CaMKII激活，以及LTP的形成和工作记忆。我们将进一步在VTA多巴胺和NAc的GABA能神经元中阐述t2-1多肽的作用机制。这一研究结果，不仅阐明了4 nAchR/D1相互作用的机制，在不同类型细胞中的功能作用，同时为t2-1多肽应用于尼古丁成瘾的治疗打下基础。