内源性大麻素对海马神经元AMPA受体GluR2的作用

Effect of endocannabinoids pretreatment on the expression of AMPA receptor GluR2 subunit in the hippocampal pyramidal neurons of mouse

目的探索内源性大麻素预处理对小鼠全脑缺血再灌注损伤后海马神经元α-氨基-3羟基-5-甲基-4-异恶唑丙酸(AMPA)受体2亚基(GluR2)表达的影响及机制。方法雄性C57BL/6小鼠随机分为假手术组、模型组、2-花生酰基甘油(2-AG)处理组、大麻素1型受体(CB1R)拮抗剂(AM251)+2-AG组和溶剂组。2-AG处理组腹腔注射2-AG 5 mg/kg;AM251+2-AG组腹腔注射AM251 1 mg/kg,30 min后腹腔给予2-AG 5 mg/kg;溶剂组腹腔给予0.1 ml二甲基亚砜(DMSO)。各组小鼠在预处理后30 min采用夹闭双侧颈总动脉20 min行再灌注的方法制备全脑缺血再灌注损伤模型。再灌注2 h取材行Western blot及免疫荧光检测。结果 GluR2高表达于正常小鼠海马CA1区锥体神经元;C57小鼠全脑缺血20 min,再灌后2 h海马组织GluR2表达明显下调(P<0.05);与模型组比较,2-AG处理组海马组织GluR2明显增高(P<0.05);与2-AG处理组比较,AM251+2-AG组海马组织GluR2明显下降(P<0.05)。结论内源性大麻素2-AG作用于神经元CB1R,逆转全脑缺血损伤导致的海马神经元GluR2表达下降。

Objective Whether endocannabinoids pretreatment affects the expression of the α-amino-3-hydroxy-5-methylisoxazole-4-propionate （AMPA） receptor 2 subunit （GluR2） in mice after global cerebral ischemia-reperfusion （I/R） injury and its mechanism are investigated.Methods Adult male C57BL/6 mice were randomly divided into five groups：sham group, I/R group, 2-arachidonoyl glycerol （2-AG） group, cannabinoid 1 receptor （CB1R） antagonist（AM251）＋2-AG group and vehicle group.The mice in 2-AG group were injected 2-AG with a dose of 5 mg/kg intraperitoneally.AM251 was injected intraperitoneally with a dose of 1 mg/kg 30 min before pretreatment.2-AG was administered in AM251＋2-AG group mice with a dose of 5 mg/kg.A total of 0.1 ml Dimethylsulfoxide （DMSO） （the solvent of 2-AG or AM251） was injected intraperitoneally in vehicle group mice. Global cerebral I/R was induced at 30 min after preconditioning by occlusion of bilateral common carotid artery for 20 min and then followed by reperfusion.Immunofluorescence and Western blot were employed to detect the expression of GluR2 in the hippocampus.Results The AMPA receptor GluR2 subunit was highly expressed in the pyramidal cell layers of hippocampal CA1.Two hours after reperfusion, global cerebral ischemia induced a marked reduction in GluR2 expression in the pyramidal neurons （P〈0.05）.In 2-AG group, hippocampal neuronal GluR2 expression was significantly up-regulated compared with that of I/R group （P〈0.05）.The GluR2 expression decreased in the AM251 ＋2-AG group compared with that in 2-AG group （ P 〈0.05 ）.Conclusion Endocannabinoids preconditioning increases the expression of hippocampal neuronal AMPA receptor GluR2 via CB1R after global cerebral ischemia.