NMDA受体和IP3受体介导NMDA诱导的小脑颗粒神经元内Ca2+超载
NMDA-induced calcium overload via NMDA and IP3 receptors in cultured cerebellar granular neurons
方小霞 1韩新华 2马颂华 1邱一华 1彭聿平1
作者信息
- 1. 南通大学医学院机能学实验室,江苏南通226001;南通大学医学院生理学系,江苏南通226001
- 2. 南通大学医学院法医学系,江苏南通226001
- 折叠
摘要
目的 探讨N-甲基-D-天门冬氨酸(NMDA)诱导的小脑颗粒神经元(CGNs)钙超载与NMDA受体(NMDAR)及细胞内钙库受体三磷酸肌醇受体(IP3R)和兰尼定受体( RyR)之间的关系.方法 取出生后8d SD大鼠小脑进行颗粒神经元体外培养.用NMDA( 100 μmol/L)急性损伤神经元,在激光扫描共聚焦显微镜(LSCM)扫描开始前30 min,或扫描开始后4,9,14 min分别加入NMDAR、IP3R、RyR拮抗剂MK-80I、2-APB和DAN,检测神经元内Ca2+浓度的动态变化.结果 MK-80I预孵育神经元经NMDA急性刺激后,神经元内Ca2+的荧光强度不再升高,NMDA刺激后加入MK-801,上升的ca2+水平立即下降,最终下降至基线水平；NMDA急性刺激2-APB预孵育神经元,神经元内Gf+的荧光强度升高,但升高幅度明显低于未经NMDA刺激组,NMDA刺激后加入2-APB,细胞内升高的Ca2+水平急剧下降,最终下降至接近基线水平；DAN预孵育的神经元经NMDA急性刺激后,胞内Ca2+的荧光强度急剧升高,达到NMDA刺激组水平,NMDA刺激后加入DAN,神经元内Ca2+水平无明显下降.结论 NMDA诱导的神经元ca2+超载,主要由细胞膜钙通道NMDAR和细胞内钙释放通道IP3R介导,而细胞内钙释放通道RyR不起主导作用.
Abstract
Objective To explore the relationship between NMDA-induced calcium overload and calcium receptors in the cerebellar granular neurons (CGNs). Methods CGNs from postnatal 8-day rats were cultured for 8 days. On the 8th day, NMDA(100 μmol/L) was added to the onedinm to stimulato the neurons. MK-801(NMDAR antagonist) , 2-APB(IP3R antagonist) , and DAN(RyR antagonist) were added before and after Laser scanning confocal microscope (LSCM) scanning. Dynamic changes of intra-cellular Ca2+ fluorescence intensity in the neurons were measured during 15 min of the NMDA stimulation. Results The NMDA stimulation of the CGNs without any pretreatment caused a notable and sustaining enhancement of Ca2+ fluorescence intensity in the cultured neurons; MK-801 (NMDAR antagonist) , 2-APB (IP3R antagonist) , and DAN (RyR antagonist) were added into the culture before NMDA for 30 min or after the beginning of LSCM scanning at the 4th, 9lh and 14th min respectively. MK-801 or 2-APB, but not DAN reduced Ca2+concentrations sharply induced by NMDA. When pretreatment with MK-801 , 2-APB, and DAN for 30 min before LSCM scanning, fluorescence intensity of Ca2+ in neurons no longer increased in MK-801 group after the NMDA acute stimulation, while enhanced in 2-APB but still lower than NMDA group, and increased rapidly in DAN group. Conclusion NMDA-induced calcium o-verload via NMDA and IP3 receptors in cultured cerebellar granular neurons, and Ryanodine receptor is not playing a leading role.
关键词
小脑颗粒神经元/N-甲基-D-天门冬氨酸/三磷酸肌醇受体/兰尼定受体/钙超载Key words
cerebellar granular neuron/N-methyl-D-aspartate/inositol 1,4,5-trisphosphate receptor/Ryanodine receptor/calcium overload引用本文复制引用
基金项目
国家自然科学基金(30870819)
南通市应用研究计划项目(K2010047)
出版年
2012
NETL
NSTL
维普
万方数据