嘌呤受体P2X7激活谷氨酸受体NMDA引起视网膜神经节细胞凋亡.DOCVIP

临床医学论文-嘌呤受体P2X7激活谷氨酸受体NMDA引起视网膜神经节细胞凋亡 ????????????????? 作者：张秀兰 张梅 胡慧玲 ?Claire H. Mitchell? ?葛坚 【摘要】? 【目的】 研究嘌呤受体P2X7和谷氨酸受体NMDA激活诱导大鼠视网膜神经节细胞（RGC）凋亡的相互作用机制。 【方法】 ①对新生Long-Evan大鼠进行上丘注射荧光标记物Aminostilbamidine标记RGC，NMDA受体通道拮抗剂MK-801、APV和Memantine分别与P2X7受体激动剂BzATP共培养，检测它们对体外培养RGC存活率的影响；②未经Aminostilbamidine标记的新生大鼠RGC，以10 μmol/L钙离子（Ca2+）荧光染料Fura-2标记后，利用Ca2+影像测定仪分别测定BzATP及三种NMDA拮抗剂对RGC胞内Ca2+浓度的影响。 【结果】 （1）三种NMDA拮抗剂均分别不同程度阻断BzATP引起的RGC凋亡。BzATP在50 μmol/L 浓度下，约杀死（36%±2%）的RGC，而MK-801（10 μmol/L）、APV（100 μmol/L）和Memantine（100 μmol/L）则均可明显减轻BzATP对RGC的毒性作用，使RGC 存活率分别提高至（96% ± 4%，P 0.001）、（80% ± 5%，P= 0.010）和（76%±9%，P =0.144）。（2）BzATP可引起RGC 胞内Ca2+持续升高，在50 μmol/L浓度下可使胞内Ca2+升高至（1183±109）nmol/L。而MK-801（10 μmol/L）、APV（300 μmol/L）和Memantine（30 μmol/L）则均可显著降低BzATP介导的Ca2+升高幅度，分别为（76% ± 7%，P =0.003）、（51%± 17%，P =0.033）和（55% ± 16%，P =0.025）。 【结论】 NMDA受体拮抗剂可阻断嘌呤受体P2X7激活诱导的RGC凋亡。P2X7受体和NMDA受体通道激活可能共同介导着兴奋性神经毒作用且P2X7在NMDA受体的上一环节先起作用。 【关键词】? 视网膜神经节细胞； P2X7受体； NMDA ?????? Abstract: 【Objective】 To demonstrate if P2X7 receptor and N-methyl-D-aspartate (NMDA) receptor link to initiate retinal ganglion cells (RGC) death. 【Methods】 (1) Long-Evan neonatal rats were back labeled with Aminostilbamidine to identify RGC. RGC cell viability was then examined using P2X7 receptor agonist BzATP and NMDA receptor antagonists MK-801, APV and Memantine. (2) RGC were dissociated from the retinas of unlabeled neonatal rats and were loaded with Fura-2, an intracellular calcium indicator. BzATP and three NMDA receptor antagonists were applied to RGC to examine their effects on intracellular Ca2+ levels using Ca2+ imaging system. 【Results】 (1) BzATP (50 μmol/L) could kill about (36% ± 2%) of the RGC. Cell death was prevented by MK-801 (10 μmol/L), APV (100 μmol/L) and Memantine (100 μmol/L) with a increasing the cell viability of (96% ± 4%) (P 0.001), (80% ± 5%, P= 0.010) and (76% ± 9%,P =0.144), respectively. (2) BzATP (50 μmol/L) led to a large, sustained increases of intracellular Ca2+ (1183 ± 109) nmol/L. Calci