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目的:探讨人参皂甙Rd(Ginsenoside Rd,GSRd)对皮层神经元兴奋性毒性损伤后细胞内游离钙离子浓度变化的影响。方法:采用原代方法培养大鼠皮层神经元,免疫荧光染色鉴定神经元纯度。应用激光共聚焦显微镜,观察GSRd对谷氨酸(Glutamate,Glu)和N-甲基-D-天门冬氨酸(NMDA)刺激后神经元胞内游离钙离子浓度变化的影响。使用钙离子荧光探针Fluo-4,AM标记细胞内游离钙,以Fluo-4的荧光强度反映细胞内游离钙浓度变化。结果:空白对照组荧光强度没有明显变化,而高浓度Glu刺激可迅速升高神经元胞内的荧光强度;在给予GSRd干预时,荧光强度升高的幅度明显降低,与MK-801的作用相似;NMDA刺激亦可使神经元胞内荧光强度明显升高,而加入GSRd干预时,荧光强度升高的幅度较NMDA损伤组有明显减小。结论:GSRd能够抑制高浓度Glu和NMDA引起的大量钙内流,提示减轻兴奋性毒性损伤过程中的钙超载可能是GSRd神经保护作用的机制之一。
Objective: To investigate the effect of ginsenoside Rd (GSRd) on the changes of intracellular free calcium concentration in cortical neurons after excitotoxic injury. METHODS: Rat cortical neurons were cultured using primary methods, and neuronal purity was determined by immunofluorescence staining. The confocal laser scanning microscopy was used to observe the effect of GSRd on the intracellular free calcium concentration of neurons stimulated by Glutamate (Glu) and N-methyl-D-aspartate (NMDA). Calcium-ion fluorescence probe Fluo-4 was used to label intracellular free calcium, and the fluorescence intensity of Fluo-4 was used to reflect the change of intracellular free calcium concentration. RESULTS: There was no significant change in the fluorescence intensity of the blank control group, but the high concentration of Glu stimulation could rapidly increase the intracellular fluorescence intensity of the neuron; when the GSRd was administered, the increase of the fluorescence intensity was significantly reduced, similar to that of MK-801. NMDA stimulation also increased the intracellular fluorescence intensity of neurons. When GSRd was added, the increase of fluorescence intensity was significantly decreased compared with NMDA-injured group. Conclusion: GSRd can inhibit a large amount of calcium influx induced by high concentrations of Glu and NMDA, suggesting that mitigating calcium overload during excitotoxic injury may be one of the mechanisms of GSRd neuroprotective effect.