新型硫化氢供体调控环磷酸鸟苷/蛋白激酶G信号通路对癫痫大鼠海马Kir6.2表达的影响  

A novel hydrogen sulfide donor modulates Kir6.2 via the cyclic guanosine monophosphate-dependent protein kinase signaling pathway in one side hippocampus of epileptic rats

作　　者：刘丹琼 朱紫婷 都昇 陈佩叶 郭曼茹 张艺泷 苏燕妹 王冠淇 徐评议[2] 朱晓琴 Liu Danqiong;Zhu Ziting;Du Sheng;Chen Peiye;Guo Manru;Zhang Yilong;Su Yanmei;Wang Guanqi;Xu Pingyi;Zhu Xiaoqin(Department of Physiology,Guangzhou Medical University,Guangzhou 511436,China;Department of Neurology,the First Affiliated Hospital,Guangzhou Medical University,Guangzhou 510120,China)

机构地区：[1]广州医科大学基础医学院生理教研室,511436 [2]广州医科大学附属第一医院神经内科,510120

出　　处：《中华神经科杂志》2021年第3期242-250,共9页Chinese Journal of Neurology

基　　金：广东省自然科学基金(2020A1515010985);国家级大学生创新创业训练计划项目(20191050003)。

摘　　要：目的探究硫化氢的抗癫痫机制即调控环磷酸鸟苷/蛋白激酶G(cGMP/PKG)信号通路对三磷酸腺苷敏感性钾通道(KATP)亚基Kir6.2表达的影响。方法将60只SD大鼠按随机数字表法分为6个组(每组10只):(1)正常对照组;(2)癫痫组;(3)硫化氢供体干预组;(4)硫化氢供体加PKG抑制剂(KT5823)组;(5)硫化氢供体加KATP抑制剂(格列本脲)组;(6)硫化氢供体正常对照组。采用腹腔注射戊四氮致大鼠癫痫发作,记录潜伏期、首次发作级别、持续时间;记录癫痫发作时海马脑电图的变化;采用蛋白质印迹法和实时荧光定量聚合酶链反应法检测海马组织中PKG、Kir6.2 mRNA和蛋白的表达水平,酶联免疫吸附测定法检测海马组织中cGMP、磷酸化PKG的浓度。结果癫痫组出现Ⅳ~Ⅴ级发作,级别为4.500(4.000,4.875)级,潜伏期短[(10.37±8.21)min],持续时间长[(69.50±24.37)s]。与癫痫组相比,硫化氢供体干预组的发作级别降低(P=0.004),潜伏期延长(P<0.001)和持续时间缩短(P<0.001)。硫化氢供体加PKG抑制剂组与硫化氢供体干预组相比,潜伏期缩短(P<0.001),持续时间延长(P<0.001)。硫化氢供体干预组癫痫波明显减少,波幅与癫痫组相比差异有统计学意义(P<0.001);硫化氢供体加PKG抑制剂组与硫化氢供体干预组相比癫痫波增加,波幅增大(P<0.001)。组间比较结果提示,各组PKG mRNA和蛋白的表达水平存在差异(分别为n=5,H=26.714,P<0.001和n=5,F=30.597,P<0.001);其中与正常对照组相比,癫痫组PKG mRNA(分别为1.000±0.001和0.782±0.064,P=0.023)和蛋白(分别为0.550±0.037和0.145±0.020,P=0.042)的表达水平显著下降。组间比较结果提示,各组Kir6.2 mRNA和蛋白的表达水平存在差异(分别为n=5,H=27.761,P<0.001和n=5,F=60.659,P<0.001);其中与正常对照组相比,癫痫组Kir6.2 mRNA(分别为1.000±0.001和0.897±0.033,P=0.004)和蛋白(分别为0.384±0.035和0.215±0.016,P=0.024)的表达水平显著下降,cGMP(P<0.001)、磷酸化PKG(P<0.001)�Objective To explore the effect of hydrogen sulfide(H2S)on modulating the subunit Kir6.2 of adenosine triphosphate sensitive potassium channels via the cyclic guanosine monophosphate-dependent protein kinase(cGMP/PKG)signaling pathway in epileptic rat models.Methods Sixty adult male SD rats were randomly divided into the following six groups(10 rats in each group)by random number table method:control,epileptic,H2S donor,H2S donor+epileptic,KT5823(one inhibitor of the cyclic guanosine monophosphate-dependent protein kinase)+H2S donor+epileptic,and glibenclamide(one inhibitor of the adenosine triphosphate sensitive potassium channels)+H2S donor+epileptic groups.Except the control group,SD rats were intraperitoneally injected with plentylenetetrazole to make the kindling models and their behaviours were recorded including the latency period,the grade,and the duration of the first epileptic seizure according to the Racine′s standard.The waveforms of electroencephalogram(EEG)in hippocampus were also recorded during the seizure.The mRNA and protein levels of PKG and Kir6.2 in hippocampus were evaluated by Western blotting and quantitative real-time polymerase chain reaction,and the hippocampal concentrations of cGMP and phosphorylation of cyclic guanosine monophosphate-dependent protein kinase(p-PKG)were detected by enzyme linked immunosorbent assay.Results Rats in the epileptic group showedⅣ-Ⅴgrade of epileptic seizure[4.500(4.000,4.875)],short latency period[(10.37±8.21)min]but long duration[(69.50±24.37)s]of seizure.Compared to the epileptic group,rats in the H2S donor group showedⅡ-Ⅲgrade of epileptic seizure(P=0.004),significantly longer latency period(P<0.001),and shorter duration of seizure(P<0.001).Compared to the H2S donor+epileptic group,rats in the KT5823+H2S donor+epileptic group showedⅢ-Ⅳgrade of epileptic seizures,significantly shorter latency period(P<0.001),and longer duration of seizure(P<0.001).The results of EEG showed that the wave patterns in the epileptic group were spike or sharp

关 键 词：癫痫 硫化氢 环磷酸鸟苷 蛋白激酶G 三磷酸腺苷敏感性钾通道 

分 类 号：R742.1[医药卫生—神经病学与精神病学]