Wang Shu-Qiu, Li Xiao-Jie, Qiu Hong-Bin, Jiang Zhi-Mei, Simon Maria, Ma Xiao-Ru, Liu Lei, Liu Jun-Xing, Wang Fang-Fang, Liang Yan-Feng, Wu Jia-Mei, Di Wei-Hua, Zhou Shaobo
Department of Pathophysiology, School of Basic Medical Sciences, Jiamusi University, Jiamusi, P. R. China.
Children Neural Rehabilitation Laboratory of Jiamusi University, School of Rehabilitation Medical Sciences, Jiamusi University, Jiamusi, Heilongjiang Province, P. R. China.
PLoS One. 2014 Jul 10;9(7):e102161. doi: 10.1371/journal.pone.0102161. eCollection 2014.
To investigate the mechanism of the anti-epileptic effect of Ganoderma lucidum polysaccharides (GLP), the changes of intracellular calcium and CaMK II α expression in a model of epileptic neurons were investigated.
Primary hippocampal neurons were divided into: 1) Control group, neurons were cultured with Neurobasal medium, for 3 hours; 2) Model group I: neurons were incubated with Mg(2+) free medium for 3 hours; 3) Model group II: neurons were incubated with Mg(2+) free medium for 3 hours then cultured with the normal medium for a further 3 hours; 4) GLP group I: neurons were incubated with Mg(2+) free medium containing GLP (0.375 mg/ml) for 3 hours; 5) GLP group II: neurons were incubated with Mg(2+) free medium for 3 hours then cultured with a normal culture medium containing GLP for a further 3 hours. The CaMK II α protein expression was assessed by Western-blot. Ca(2+) turnover in neurons was assessed using Fluo-3/AM which was added into the replacement medium and Ca(2+) turnover was observed under a laser scanning confocal microscope.
The CaMK II α expression in the model groups was less than in the control groups, however, in the GLP groups, it was higher than that observed in the model group. Ca(2+) fluorescence intensity in GLP group I was significantly lower than that in model group I after 30 seconds, while in GLP group II, it was reduced significantly compared to model group II after 5 minutes.
GLP may inhibit calcium overload and promote CaMK II α expression to protect epileptic neurons.
研究灵芝多糖（GLP）抗癫痫作用的机制，探讨癫痫神经元模型中细胞内钙及CaMK II α表达的变化。
原代海马神经元分为：1）对照组，神经元用Neurobasal培养基培养3小时；2）模型组I：神经元用无镁培养基孵育3小时；3）模型组II：神经元用无镁培养基孵育3小时，然后再用正常培养基培养3小时；4）GLP组I：神经元用含GLP（0.375mg/ml）的无镁培养基孵育3小时；5）GLP组II：神经元用无镁培养基孵育3小时，然后再用含GLP的正常培养基培养3小时。通过蛋白质免疫印迹法评估CaMK II α蛋白表达。使用添加到更换培养基中的Fluo-3/AM评估神经元中的Ca(2+)周转，并在激光扫描共聚焦显微镜下观察Ca(2+)周转。
模型组中CaMK II α表达低于对照组，然而，在GLP组中，其高于模型组。30秒后，GLP组I中的Ca(2+)荧光强度明显低于模型组I，而在GLP组II中，5分钟后与模型组II相比明显降低。
GLP可能抑制钙超载并促进CaMK II α表达以保护癫痫神经元。
