帕金森病运动并发症发生机制中G蛋白偶联受体激酶6与N-甲基-D-天冬氨酸受体关系的研究

摘要

目的 研究G蛋白偶联受体激酶6(GRK6)在帕金森病(PD)运动并发症发生机制中与N-甲基-D-天冬氨酸(NMDA)受体的关系.方法 建立PD运动并发症大鼠模型,25只大鼠分为3组.异动症(LID)组10只,腹腔注射左旋多巴甲酯23 d;MK-801处理组10只,第23天左旋多巴甲酯注射前腹腔注射MK-801;PD组5只,腹腔注射0.2%维生素C液.另设假手术组5只为对照组.观察MK-801处理左旋多巴诱导的运动并发症模型大鼠的行为变化,并用免疫组织化学方法和Western印迹方法检测大鼠纹状体区GRK6蛋白的表达情况.结果 PD组大鼠长期使用左旋多巴后出现明显的异常不自主运动,与人类LID具有相似特征.免疫组化结果显示.PD组损伤侧纹状体区GRK6阳性细胞指数减少至(4.81±1.31)×103(P＜0.05),LID组损伤侧GRK6阳性细胞指数进一步减少至(3.23±0.41)×103(P＜0.01).MK-801组LID大鼠异常不自主运动评分减少,药效期延长,同时损伤侧纹状体区GRK6阳性细胞指数增多至(4.64±1.39)×103(P＜0.05).Western印迹法检测结果同免疫组化基本相符,PD组损毁侧/未损毁侧纹状体区GRK6含量比值为(83.7±2.1)%,LID组GRK6值降低为(76.8±2.2)%,而MK-801组GRK6值升高至(91.1±2.7)%(P＜0.01).结论 NMDA受体拮抗剂能逆转大鼠运动并发症的发生,其机制可能与GRK6增多,抑制了谷氨酸受体的过度活化有关.%Objective To investigate the relationship between G protein-coupled receptor kinase 6 (GRK6) and N-methyl-D-aspartate (NMDA) receptor in the mechanism study underlying motor complications in Parkinson's disease (PD).Methods The rat models (n= 25) of Parkinsonism related motor complications were established and were randomly divided into levodopa-induced dyskinesia (LID) group (n= 10,intraperitoneal injection of levodopa for 23 d),MK-801 treatment group (n= 10,intraperitoneal injection of MK-801 at day23 after intraperitoneal injection with levodopa for 22 days) and PD group (n= 5,intraperitoneal injection of vitamin C).Another 5 rats were served as controls (sham-operation group).The behavior changes of rats in MK-801 treatment group were observed,and the expression of GRK6 in the striate of rats was detected by immunohistochemistry and Western blot.Results After the chronic treatment with levodopa methyl ester,PD rats displayed abnormal involuntary movements,which was similar to levodopainduced dyskinesia in PD patients.Immunohistochemistry showed that GRK6-positive cells of lesion side were decreased in LID rats as compared with PD rats [(3.23±0.41 ) × 103 vs.(4.81 ± 1.31 ) ×103,P＜0.01].Rats in MK-801 treatment group displayed the decreased AIM scores and increased peak rotation,and the increased GRK6-positive cells of lesion side as compared with LID rats (P＜0.05).Western blot showed that the levels of GRK6 was 83.7% ±2.1% in PD group (presented as lesion side/unlesion side),76.8% ± 2.2% in LID group and 91.1% ± 2.7% in MK-801 treatment group (intergroups comparison:all P＜0.05).These results were in accordance with the results of immunohistochemistry.Conclusions Antagonist of NMDA receptor can be used to reduce the motor complications in rats.It may be due to increased GRK6 which inhibits the overactivation of glutamic acid receptors.