GSK-3β在铝致原代海马神经元网络电活动损伤的机制初探  

Preliminary study on the mechanism of GSK-3βin electrical activity damage of primary hippocampal neuronal network induced by aluminum

作　　者：卢丽媛 李萌 贺小玉 路小婷[1] 秦小江[1] 张慧芳[1] LU Li-yuan;LI Meng;HE Xiao-yu;LU Xiao-ting;QIN Xiao-jiang;ZHANG Hui-fang(School of Public Health Shanxi Medical University,Taiyuan Shanxi 030001,China)

机构地区：[1]山西医科大学公共卫生学院,山西太原030001

出　　处：《毒理学杂志》2023年第2期158-165,共8页Journal of Toxicology

基　　金：山西省基础研究计划“自然科学研究面上项目”(202103021224226)。

摘　　要：目的初步探讨GSK-3β在铝致原代海马神经元网络电活动损伤中的作用及机制。方法24 h内新生的ICR小鼠海马原代神经元进行体外培养至第10天,进行麦芽酚铝[Al(mal)3)]染毒及干预,实验分组为空白对照组、二甲基亚砜(DMSO)组、SB(GSK-3β抑制剂SB216763,1μmol/L)组、Al(20μmol/L)组、SB(1μmol/L)+Al(20μmol/L)组,于干预0和48 h点使用微电极阵列分析仪(Micro-Electrode Array,MEA)检测海马神经元网络电活动;染毒48 h后,免疫荧光观察神经元树突棘密度;Western blot法检测GSK-3β、p-GSK-3β、PSD95和Arc蛋白的表达水平。结果Al(mal)3作用0 h时间点,各组神经元未见明显差异,在48 h时间点,各组神经元均呈节律性簇发放电。空白对照组神经元自发放电频率、簇发放电频率、网络簇发放电频率、同步指数分别为172.82%±44.56%、134.65%±25.60%、209.48%±70.09%、120.10%±15.32%;与空白对照组相比,DMSO组及SB组未见明显差异(F=6.93,F=4.35,F=1.81,F=13.43,P>0.05),而Al组神经元网络电活动明显下降(P<0.05);与Al组相比,SB+Al组神经元网络电活动明显升高。与空白对照组相比,Al组的树突棘密度明显降低(F=16.82,P<0.05);与Al组相比,SB+Al组的树突棘密度明显增加(P<0.05)。与空白对照组相比,各干预组的GSK-3β蛋白表达水平均无明显变化(F=2.27,P>0.05);与空白对照组相比,SB组的p-GSK-3β蛋白表达水平明显升高(F=45.84,P<0.05),Al组和SB+Al组明显降低(P<0.05),但与Al组相比,SB+Al组的p-GSK-3β蛋白表达水平明显升高(P<0.05);Al组和SB+Al组的PSD-95、Arc蛋白表达水平与空白对照组相比均明显降低(F=37.61,F=37.13,P<0.05),但与Al组相比,SB+Al组的PSD-95、Arc蛋白表达水平明显升高(P<0.05)。结论铝可明显抑制大鼠原代海马神经元的网络电活动,GSK-3β可能通过影响神经元突触结构进而参与铝致神经元的网络电活动损伤。Objective To preliminary explores the mechanism of GSK-3βin the electrical activity damage of primary hippocampal neuron network induced by aluminum.Methods The hippocampus of the newborn ICR mice within 24 h was used for in vitro culture of primary neurons until the 1oth day,and aluminum intervention was performed.The experimental groups were:blank control group,dimethyl sulfoxide(DMSO)group,SB(GSK-3βinhibitor,1μmol/L)group,Al(20μmol/L)group,SB(1μmol/L)+Al(20μmol/L)group,microelectrodes were used at O h and 48 h of the intervention Array analyzer(Micro-Electrode Array,MEA)was used to detect the electrical activity of hippocampal neuron network;48 h after exposure,the density of neuronal dendritic spines was observed by immunofluorescence;Western blot method was used to detect the expression levels of CSK-3β,p-GSK-3β,PSD95 and Arc proteins.Results At the time point of O h of Al(mal)exposure,there was no statistical difference among the neurons in each group.At the time point of 48 h,the neurons in each group showed rhythmic cluster discharge.The spontaneous discharge frequency,cluster discharge frequency,network cluster discharge frequency and synchronization index of neurons in the blank control group were 172.82%±44.56%,134.65%±25.60%,209.48%±70.09%and 120.10%±15.32%respectively.Compared with the blank control group,there was no significant difference between DMS0 group and SB group(F=6.93,F=4.35,F=1.81,F=13.43,P>0.05),the network electrical activity of neurons in Al group decreased significantly(P<0.05);Compared with Al group,the parameters of SB+Al group were significally higher.Compared with the blank control group,the density of dendritic spines in the Al group was significally decreased(F=16.82,P<0.05),and compared with the Al group,the density of dendritic spines in the SB+Al group was significantly increased(P<0.05).Compared with the blank control group,the expression level of GSK-3βprotein in each intervention group had no significal difference(F=2.27,P>0.05);Compared with the blank control

关 键 词：铝 糖原合成酶激酶-3Β 突触后致密蛋白95 活性调节细胞骨架蛋白 神经元网络电活动 

分 类 号：R135[医药卫生—劳动卫生] R99[医药卫生—公共卫生与预防医学]