伏隔核D1/D2共表达中等多棘神经元在孤独症小鼠社交奖赏障碍中的作用及机制研究

Impaired social interaction is one of the core symptoms to define autism spectrum disorder (ASD). It has become increasingly serious social issues. Impaired social rewards are likely to be important factors in social deficits of ASD patients. The nucleus accumbens (NAc) is a crucial brain region for regulating social rewards, which may result in lacking of social motivation for ASD patients. Previously, a large proportion of studied focused on the rule of medium-sized spiny neurons expressing D1 receptors (D1 MSNs) and D2 MSNs expressing D2 receptors in NAc related function. However, several groups and our results found that about one-third of MSNs in the NAc expressed D1-D2 heteromers formed by D1 and D2 receptors (D1/D2 MSNs). The function of D1/D2 MSNs in the NAc remains largely elusive, especially in social reward process. We speculate that D1/D2 MSNs may participate in NAc's regulation of social rewards deficits in ASD mouse model. Shank3 knockout (KO) mice exhibit typical social behavioral deficits. This proposal is planning to characterize the changes in neuronal activities (including intrinsic properties and glutamatergic synaptic function) and morphology of D1/D2 MSNs and explore the pharmacological effects of D1-D2 heteromers on D1/D2 MSNs in the NAc of Shank3 KO ASD mouse model. Based upon the above in vitro results, our proposal then focuses on the functional role of D1/D2 MSN in the NAc underlying social reward behavior in Shank3 KO mice by specifically activating or inhibiting with pharmacological and optogenetic methods. Combined with these results, our work will help to provide a scientific pathological mechanisms of ASD social reward disorders for finding potential drug intervention targets.

社交障碍是孤独谱系障碍（autism spectrum disorders，ASD）患者的核心症状之一，带来严重社会问题。伏隔核调控社交奖赏，致ASD患者社交动机不足。以往伏隔核多关注表达多巴胺D1受体的中等多棘神经元（D1 Medium-sized spiny neurons，D1 MSNs）和表达D2受体的D2 MSNs在ASD的作用。他人和我们均发现伏隔核存在近1/3 MSNs共表达D1和D2受体（D1/D2 MSNs），在伏隔核中作用尚不清楚。我们推测伏隔核的D1/D2 MSNs参与ASD社交奖赏障碍。Shank3敲除小鼠具有典型ASD社交障碍。我们拟研究该小鼠伏隔核D1/D2 MSNs电活动（内在特性和兴奋性突触功能）及形态学改变，受体机制，药理学和光遗传学特异性调控该群细胞对社交奖赏行为的影响，为揭示ASD社交障碍发生的神经环路机制及可能的干预靶点提供科学依据。

社会交往活动对群居生活的种群至关重要，社交活动涉及了社交识别，社交记忆，社交奖赏，社交认知等多个复杂环节，关联着众多脑区。社交奖赏的功能受损很可能是孤独症患者社交行为障碍的重要因素，伏隔核是社交奖赏信息和其他社交信息的重要汇聚点，临床研究显示伏隔核异常可导致社交奖赏异常。多巴胺D1受体和D2受体分属Gs和Gi蛋白偶联受体，引发不同的信号传导途径并作用于不同的靶点产生相互拮抗的效应，调控伏隔核的最终输出。本项目聚焦于探究D1-D2受体共表达神经元在孤独症小鼠社交奖赏障碍中的作用及机制研究。在体功能研究方面，首先通过三箱测试的方法来检测Shan3 WT和KO小鼠的社交行为，结果表明相较于WT小鼠，Shank3 KO小鼠社交行为异常，主要体现在KO小鼠对于三箱两侧的社交对象和非社交对象没有明显偏好。接下来使用sCPP（social conditioned place preference）实验检测小鼠的社交奖赏行为。sCPP实验结果表明wt小鼠社交偏好评分均值在1.2，而ko小鼠的社交偏好评分均值在1左右，说明Shank3 ko小鼠社交奖赏行为可能受损。为了特异性标记并调控D1-D2 MSNs来研究它在社交奖赏行为中的作用，我们进行了高通量单细胞测序筛选D1-D2 MSNs的marker基因，同步使用病毒标记的方法来实现这一目标。离体功能研究方面，首先我们采用了免疫组织荧光染色的方法对伏隔核脑区共染了DARPP-32，结果表明，伏隔核D1-D2 MSNs是投射神经元。全细胞膜片钳记录结果表明，相比较2种单阳性神经元，D1-D2 MSNs放电数更多，也就是兴奋性更高，并且在静息状态下更去极化，它的内向整流钾Kir电流明显更小。此外我们还进行了躁狂症和睡眠障碍模型的神经机制和干预靶点研究，结果表明槲皮素可通过激活小鼠的漏钾通道TREK-1来缓解由安非他命诱导的躁狂样行为。此外，TNF信号通路介导的小胶质细胞活化是小鼠急性睡眠剥夺诱导的焦虑样行为的基础。本项目的支持帮助我们了解神经障碍性疾病有着至关重要的作用，对相关神经机制和干预靶点将继续深入探究。

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