姜黄素对AMPA／KA受体介导大鼠海马神经元钙内流的影响

Effect of curcumin on AMPA and kainate receptor-mediated calcium influx in cultured rat hippocampal neurons

目的探讨姜黄素对α-氨基-3-羧基-5-甲基异恶唑-4-丙酸（AMPA）／海人酸（KA）受体介导大鼠海马神经元钙内流的影响。方法选用胚胎17d SD鼠分离海马，离体培养海马神经元，借助活体钙荧光染色和激光共聚焦钙成像技术观察100μmol／L KA刺激海马神经元内钙的变化，不同浓度（5、10、15、30、50μmol／L）姜黄素预孵育海马神经元30min对100μmol／L KA刺激下细胞内钙变化的影响，15μmol／L姜黄素对不同浓度（10、30、50、100、200、300μmol／L）KA刺激海马神经元内钙变化的影响。应用钴染色技术观察（30、100μmol／L KA）刺激后海马神经元钴阳性染色细胞变化，姜黄素预孵育30min对KA刺激导致钴阳性染色细胞变化的影响。结果不同浓度姜黄素预孵育30min均可以明显缓解100μmol／L或300μmol／L KA导致的细胞内钙升高程度，差异均有统计学意义（P〈0．05），其中15μmol／L姜黄素作用最为明显。30μmol／L或100μmol／L KA刺激均可以引起海马神经元钻染色阳性细胞增加，15μmol／L姜黄素预处理30min后明显减少钴染色阳性细胞，差异有统计学意义（P〈0．05），而其他浓度（5μmol／L或30μmol／L）姜黄素未见明显影响。结论一定浓度的姜黄素可以影响AMPA／KA受体介导大鼠海马神经元钙内流，这可能是姜黄素抗癫痫作用的一个机制。

Objective To investigate the effect of curcumin on alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid （AMPA） and kainate （KA） receptor-mediated calcium influx in cultured rat hippocampal neurons. Methods The hippocampal neurons from SD rat embryos （17 days old） were cultured for 9 days, and fluorescent calcium chelator and confocal microscopy calcium imaging were used to observe the changes in intracellular flee calcium in the neurons following stimulation with 100 μmol/L KA. The effect of curcumin pretreatment at different concentrations （10, 30, 50, 100, 200 and 300 μmol/L） for 30 min on 100 μmol/L KA-induced intracellular calcium changes in the neurons were evaluated, and the effect of 15 μmol/L curcumin on intracellular calcium in the neurons stimulated with 10, 30, 50, 100, 200 and 300 μmol/L KA were also assessed. Cobalt staining was used to examine the expression of calcium permeable AMPA/KA channel in the hippocampal neurons treated with 30 and 100 μmol/L KA. Results Curcumin pretreatment at different concentrations for 30 min significantly relieved the elevation of free calcium concentration induced by 100 or 30 μmol/L KA （P〈0.05）, and the effect was especially obvious with 15 μmol/L KA curcumin. Both 30 and 100 μmol/L KA increased the ratio of cobalt staining-positive hippocampal neurons, and curcumin pretreatment at 15 μmol/L, but not at 5 or 30 μmol/L, significantly reduced the ratio of the positive neurons （P〈0.05）. Conclusion Curcumin at appropriate concentrations can modulate calcium influx mediated by AMPA and KA in cultured rat hippocampal neurons, which might be one of the anti-epileptic mechanisms of curcumin