基于NRXNs-NLGNs-SHANK3通路活性及相关基因甲基化修饰变化探讨电针调控自闭症模型大鼠海马突触可塑性的机制

Recently, an article reported postsynaptic protein SHANK3 closely related with autism in Nature magazine. The clinical research found that,acupuncture treatment is effective on autism. The experiment showed that the electric acupuncture can promote the change of autism model rat's hippocampus synaptic plasticity. Another related experiments show NRXNs-NLGNs-SHANK3 signal pathway is closely related to synaptic plasticity and autism. This topic selects autism rat models for object, through the NRXNs-NLGNs-SHANK3 signal pathway , observe hippocampus synaptic plasticity changes of the rats by the transmission electron microscopy,observe hippocampus NRXN1 、NLGN3、 NLGN4 、PSD-95、 p-SHANK3 、SHANK3 and Homer2 protein and gene expression by the Western Blot and immunohistochemical RT-PCR technology,observe the pathway related genes NRXN1、NLGN3、NLGN4 methylation modify by MSP and BDS technology, then explain the effect of autism model rat's hippocampus synaptic plasticity by electric acupuncture in molecular biology and epigenetics level,try to clarify cupping treatment mechanism of autism.

近期Nature杂志报道突触后蛋白SHANK3与自闭症密切相关。本课题组临床和实验研究表明：临床针刺治疗自闭症有效，且电针可促进自闭症模型大鼠海马区突触可塑性的变化；另有相关实验表明NRXNs-NLGNs-SHANK3通路与突触可塑性及自闭症密切相关。本课题以自闭症模型大鼠海马区NRXNs-NLGNs-SHANK3信号通路为切入点，采用透射电镜观察大鼠海马区超微结构的变化，采用免疫组织化学、Western Blot和RT-PCR技术检测大鼠海马区NRXN1、NLGN3、NLGN4、PSD-95、p-SHANK3、SHANK3和Homer2蛋白和基因的表达情况，采用MSP和BDS技术检测NRXN1、NLGN3、NLGN4基因甲基化修饰情况，进而从分子生物学和表观遗传学层面阐述电针对自闭症模型大鼠海马区突触可塑性的影响，尝试阐明电针治疗自闭症的机制。

本课题选用自闭症大鼠为观察对象，观察针刺长强穴（动物身上该穴称为“后海”）后其学习记忆的变化及海马超微结构的影响，结合NRXNs-NLGNs-SHANK3通路上NRXN-1、NLGN-3、PSD-95及缝隙连接相关蛋白的变化，探讨针刺长强穴治疗自闭症的机制。得出：⑴针刺后海穴可改善自闭症大鼠的学习和记忆能力；⑵针刺后海穴可改善自闭症大鼠大脑CA1区超微结构的表达；⑶针刺后海穴可提高自闭症模型大鼠大脑皮层M1区和海马CA1区PSD-95蛋白的表达；⑷针刺后海穴可提高自闭症模型大鼠大脑海马NRXN-1和NLGN-3蛋白的表达；⑸针刺后海穴能通过提高额叶皮层缝隙连接相关蛋白CX43、CX32、CX36蛋白的表达来改善自闭症模型大鼠学习和记忆能力。