自噬对孤独症大鼠海马组织兴奋性、抑制性突触相关蛋白表达的影响及机制  

Effects of autophagy on expression of excitatory and inhibitory synaptic related proteins in the hippocampus of autistic rats

作　　者：罗瑜平 文敏[1] 熊江福 周波[1] 艾戎 童雪涛[1] LUO Yuping;WEN Min;XIONG Jiangfu;ZHOU Bo;AI Rong;TONG Xuetao(Guizhou Medical University, Guiyang 550004, China)

机构地区：[1]贵州医科大学,贵阳550004 [2]贵州医科大学附属医院

出　　处：《山东医药》2019年第11期34-39,共6页Shandong Medical Journal

基　　金：贵州省科技合作计划项目[黔科合LH字(2015)7351]

摘　　要：目的探讨自噬对孤独症大鼠海马组织兴奋性、抑制性突触相关蛋白表达的影响及机制。方法取繁殖期Wistar雌鼠20只、雄鼠10只合笼过夜,于雌鼠受精后第12. 5天,将10只孕鼠腹腔注射丙戊酸(VPA),其后代鼠标记为VPA干预组;另取5只孕鼠腹腔注射同等剂量生理盐水,其后代鼠标记为对照组。在VPA干预组后代鼠出生后第35天,随机取30只分为模型组、自噬抑制组[3-甲基腺嘌呤(3-MA)组]和自噬增强组(Rap组),每组各10只,分别腹腔注射生理盐水、3-MA 5 mg/kg和雷帕霉素5 mg/kg;对照组10只腹腔注射同等剂量的生理盐水。后代鼠出生后第42天时,采用自梳理实验评价刻板重复行为,采用三箱实验评价各组社交能力;然后处死各组大鼠,取海马组织,采用Western blotting法及免疫组化法检测海马组织突触素(Syn)、兴奋性突触[突触后致密蛋白95(PSD-95)]和抑制性突触[桥尾蛋白(Gephyrin)]的表达。比较各组PSD-95/Gephyrin比值变化。结果 (1)自梳理实验及三箱实验示,与对照组相比,模型组出现刻板重复行为和社交能力障碍,Rap组上述行为改善,3-MA组上述行为加重(P均<0. 05)。(2)Western blotting及免疫组化示,与对照组相比,模型组海马Syn、PSD-95蛋白表达均上调,Gephyrin蛋白表达下调(P均<0. 05);与模型组相比,3-MA组Syn、PSD-95蛋白表达均上调,Gephyrin蛋白表达下调(P均<0. 05);与模型组相比,Rap组Syn、PSD-95蛋白表达均下调,Gephyrin蛋白表达水平上调(P均<0. 05)。(3)与对照组相比,模型组PSD-95/Gephyrin比值上调;与模型组相比,3-MA组PSD-95/Gephyrin比值上调,Rap组PSD-95/Gephyrin比值下调(P均<0. 05)。结论孤独症大鼠存在海马组织兴奋性/抑制性突触失衡,增强自噬可恢复兴奋性/抑制性突触失衡,从而改善孤独症症状。Objective To investigate the effects and mechanism of autophagy on the expression of excitatory and inhibitory synaptic related proteins in the hippocampus of autistic rats. Methods Twenty female Wistar rats and 10 male Wistar rats were bred in cages overnight. On the 12.5 day after fertilization, 10 pregnant mice were intraperitoneally injected with valproic acid (VPA). Their offspring were labeled as the VPA intervention group. Another 5 pregnant mice were intraperitoneally injected with the same dose of normal saline, and their offspring were labeled as the control group. On the 35th day after birth, 30 mice in the VPA intervention group were randomly divided into the model group, autophagy inhibition group (3 methyladenine, 3-MA group) and autophagy enhancement group (rapamycin, Rap group). Ten mice in each group were intraperitoneally injected with normal saline, 5 mg/kg 3-MA and 5 mg/kg rapamycin. In the control group, 10 rats were intraperitoneally injected with the same dose of normal saline. On the 42nd day after birth, the self-grooming experiment was used to evaluate the stereotyped and repetitive behaviors of each group, and the three-box experiment was used to evaluate the social skills of each group. Then, rats in each group were sacrificed and hippocampal tissues were collected. The expression of synaptophysin(Syn), postsynaptic density 95 (PSD-95) and Gephyrin in the hippocampal tissues were detected by Western blotting and immunohistochemistry. The PSD-95/Gephyrin ratio of each group was compared. Results ① Self-combing experiment and three-box experiment: compared with the control group, the stereotyped repetitive behaviors and social dysfunction occurred in the model group, and the above behaviors improved in the Rap group, and the above behaviors aggravated in the 3-MA group (all P <0.05).② in Western blotting and immunohistochemistry: compared with the control group, the expression of hippocampal Syn and PSD-95 was up-regulated and the expression of Gephyrin was down-regulated in the mode

关 键 词：孤独症 自噬 突触 兴奋性 抑制性 海马 突触素 突触后致密蛋白95 桥尾蛋白 大鼠 

分 类 号：R729[医药卫生—儿科]