摘要
目的:研究多巴胺(dopamine,DA)对星形胶质细胞谷氨酸(glutamate,Glu)摄取能力的影响,以及DA通过哺乳动物雷帕霉素靶蛋白(m TOR)-兴奋性氨基酸转运体2(EAAT2)信号通路对星形胶质细胞Glu摄取能力的影响。方法:采用Amplex Red谷氨酸测定试剂盒检测经过干预的原代皮层星形胶质细胞对Glu摄取含量的变化,RT-q PCR、Western blot和免疫荧光染色等检测EAAT2和m TOR mRNA和蛋白质相对表达量,m TOR拮抗剂雷帕霉素或m TOR兴奋剂MHY1485干预在DA中共培养的星形胶质细胞,检测m TOR和EAAT2的表达情况,以及培养上清液Glu的含量。结果:DA干预的原代星形胶质细胞中m TOR表达下调,EAAT2表达下调,培养上清液Glu水平上升;雷帕霉素干预后,EAAT2表达下调,培养上清液中Glu的含量增加;MHY1485干预后,EAAT2表达上调,培养上清液中Glu的含量下降。结论:DA通过与星形胶质细胞m TOR-EAAT2通路相互作用,减弱星形胶质细胞摄取Glu的能力,引起细胞外Glu蓄积,最终损伤星形胶质细胞的功能。
AIM：To investigate the effect of dopamine （ DA） on the glutamate （ Glu）-uptake ability of astro-cytes, and the role of mammalian target of rapamycin （mTOR）-excitatory amino acid transporter 2（EAAT2） pathway in this process .METHODS：Extracellular Glu levels in DA-treated primary cortical astrocytes （ PCAs） were measured by a fluorimetric method .The relative expression of EAAT 2 and mTOR at mRNA and protein levels was measured by RT-qPCR and Western blot .PCAs stimulated with or without DA in the presence or absence of mTOR antagonist rapamycin or mTOR agonist MHY1485 were used to determine the expression of mTOR and EAAT 2, and Glu content in the culture supernatant was also measured.RESULTS： The expression of mTOR in DA-treated PCAs was decreased, the expression of EAAT2 was also decreased .Extracellular Glu levels of DA-treated PCAs were elevated significantly .When the PCAs were stimula-ted with DA in the presence of rapamycin , the expression of EAAT2 was decreased , and the levels of extracellular Glu was significantly increased.In the presence of MHY1485, the expression of EAAT2 was elevated, and significant decrease in the levels of extracellular Glu was also observed .CONCLUSION：DA interacts with mTOR-EAAT2 pathway to reduce the Glu-uptake ability of the astrocytes , and causes extracellular Glu accumulation , ultimately destroys the function of astro-cytes.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2017年第10期1746-1750,共5页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.81671042
No.81300308)
