安神定志灵调控ADHD多巴胺D1/D2受体信号通路及脱敏/复敏的机制研究

Attention-deficit/hyperactivity disorder (ADHD) is a common behavioral disorder in childhood and adolescent, and the etiology of ADHD is still unknown. The dopamine deficit theory is potential to be further studied. Our researches revealed that Anshen Dingzhi Ling (ADL) which has valid clinical effects could up-regulate the expression of D1R, D2R mRNA and protein in the prefrontal cortex and striatum, and also decrease the expression of DAT mRNA as well as protein in striatum of spontaneously hypertensive rat (SHR) model of ADHD. However, the complete mechanism of how D1R/D2R and DAT alleviate ADHD symptoms is not yet clear. Researches demonstrated that D1R/D2R signaling pathway regulate synthesis, release and clearance of dopamine, and also participate in receptor desensitization/resensi-.tization. The interaction of D1R/D2R signaling pathway and receptor desensiti- zation/resensitization induce the occurrence of core symptoms of ADHD which prove to be a new research target. Hence, we infer that ADL may exert its effect through regulating some target spots and links of D1R/D2R signaling pathway and receptor desensitization/resensitization. In order to confirm this hypothesis, we intend to apply various technologies such as cellular and molecular biology techniques to observe the expression of protein involved in D1R/D2R signaling pathway and factors regulating receptor desensitization/resensitization in prefrontal cortex and striatum of SHR rats. It has significant academic and clinical values to go further study of multiple targets of ADL.

注意力缺陷多动障碍（ADHD）是儿童期常见的行为障碍疾病，该病的发病机制至今不明，多巴胺缺陷理论是一具有潜力的研究热点。我们研究显示，临床疗效确切的安神定志灵可上调ADHD模型SHR大鼠D1R/D2R表达和下调DAT表达，但如何通过D1R/D2R及DAT而改善ADHD症状的机制尚未明确。研究表明，D1R/D2R信号通路参与了多巴胺的合成、释放及清除，同时参与了多巴胺受体的脱敏/复敏，两者相互影响共同介导了ADHD核心症状的发生，成为疾病研究的新靶点。因此，我们推测安神定志灵可能通过调控D1R/D2R信号通路、受体脱敏/复敏的某些靶点或环节而发挥效应。为了证实这一假说，我们运用多种细胞分子生物学技术，观察SHR大鼠前额叶及纹状体中，D1R/D2R信号通路蛋白及受体脱敏/复敏调控因子的表达变化，进一步探讨安神定志灵的作用靶点，具有重要的学术和临床应用价值。

注意力缺陷多动障碍（ADHD）是儿童期常见的行为障碍性疾病，该病的发病机制至今不明，多巴胺缺陷理论是当前国际上的研究重点和热点。前期研究表明，临床疗效确切的安神定志灵可上调ADHD模型SHR大鼠D1R/D2R表达和下调DAT表达，但如何通过D1R/D2R及DAT而改善ADHD症状的机制尚未明确。为明确安神定志灵调控D1/D2受体表达及效应的作用机制，并分析可能参与的信号分子之间的相互作用，我们运用动物行为学实验方法及Western Blot/Elisa/Real-time PCR/IHC/IF等现代实验技术进行了深入研究。通过研究我们发现，安神定志灵能够减少SHR大鼠的运动量、降低平均运动速度，并能够提高SHR大鼠水迷宫实验中的学习记忆能力，通过减少SHR大鼠多动、冲动及注意力不集中的行为表现，控制ADHD的核心症状。在作用机制研究中，安神定志灵能够下调突触前膜促进DA清除的ERK1/2信号通路的表达并上调促进DA合成的AC/cAMP/PKA信号通路及TH、DDC等相关因子的表达，同时调控介导多巴胺释放的CaMKII信号通路及VMAT2、Syntaxin-1A、SNAP-25等因子，共同影响突触前膜多巴胺的合成、释放及清除。在突触后膜中，安神定志灵能够调控多巴胺D1/D2受体共同介导的AC/cAMP/PKA/DARPP-32经典通路及多巴胺D2受体优势通路AKT/GSK-3的表达，同时提高多巴胺受体复敏相关的β-arrestin2、NCS-1、PSD-95、PP2A、PKA表达并降低脱敏相关的GRK-6的表达，发挥增强受体复敏、减少脱敏的干预作用。这些研究结果共同表明多巴胺D1/D2受体可能是安神定志灵的药效作用靶点，安神定志灵通过调控突触前膜多巴胺的合成、释放及清除及突触后膜多巴胺受体信号通路及受体的脱敏/复敏相关因子的表达，调控脑内多巴胺系统，影响脑内多巴胺的水平，改善SHR大鼠前额叶-纹状体回路的调控异常而控制ADHD的核心症状。这些研究结果进一步揭示了安神定志灵的作用靶点，对认识ADHD的发病机制及中医药治疗ADHD的潜在药效机制具有指导意义，有利于发挥中医药在ADHD疾病研究中的特色和优势，并为安神定志灵治疗ADHD中药新药的开发奠定一定的实验基础，为后续的研究开辟了道路，具有重要的学术和临床应用价值。