人参皂苷Rb1经鼻给药对戊四唑慢性点燃小鼠的抗癫痫作用

Effect of Intranasal Ginsenoside Rb1 Against Epilepsy in Mice Induced by Chronic Ignition of Pentylenetetrazole

目的:观察人参皂苷Rb_(1)经鼻给药的抗癫痫作用并对作用机制进行初探研究。方法:采用小鼠戊四唑(PTZ)腹腔注射构建慢性癫痫模型,造模成功后(21 d)将癫痫小鼠随机分为PTZ组、丙戊酸钠(VPA)组、人参皂苷Rb_(1)低、高剂量组(20、40 mg·kg^(-1))。各组小鼠分别经鼻给予相应药物30 d,每日2次,空白组经鼻给予等体积生理盐水。给药期间记录小鼠体质量变化、癫痫发作潜伏期、癫痫发作级别,远程无线记录动物脑电图(EEG)变化。采用神经元特异核蛋白(NeuN)观察大脑皮层及海马神经元损伤,采用离子钙结合衔接分子-1(IBA-1)、胶质纤维酸性蛋白(GFAP)分别观察小胶质细胞激活、星形胶质细胞活化现象。用谷氨酸(Glu)转运体-1(GLT-1)、谷氨酰胺合成酶(GS)观察Glu调节关键分子变化。结果:与空白组比较,PTZ明显抑制小鼠体质量增加(P<0.05,P<0.01),显著缩短癫痫发作潜伏期(P<0.01),显著升高癫痫发作最高级别(P<0.01),癫痫样脑电波增加。与PTZ组比较,人参皂苷Rb_(1)低、高剂量组小鼠体质量增加(P<0.05,P<0.01),癫痫发作潜伏期延长(P<0.05,P<0.01),癫痫发作级别降低(P<0.05,P<0.01),癫痫样脑电波减少。免疫荧光(IF)结果显示,人参皂苷Rb_(1)治疗显著改善PTZ引起的小鼠大脑皮层运动感觉区及海马CA1区神经元损伤(P<0.05,P<0.01),并明显抑制小胶质细胞激活(P<0.05,P<0.01)、星形胶质细胞活化。进一步研究发现,人参皂苷Rb_(1)治疗显著改善癫痫小鼠大脑星形胶质细胞GLT-1(P<0.01)和GS表达(P<0.01)。结论:人参皂苷Rb_(1)经鼻给药能显著改善PTZ引起的小鼠癫痫症状,对癫痫小鼠大脑神经元损伤具有明确保护作用,并能显著抑制癫痫小鼠大脑小胶质细胞激活和星形胶质细胞活化,其抗癫痫作用可能与调节星形胶质细胞Glu代谢关键分子GLT-1和GS有关。

Objective:To observe the effect of intranasal ginsenoside Rb_(1) against epilepsy and preliminarily explore the mechanism.Method:The mouse model of chronic epilepsy was established by intraperitoneal injection of pentylenetetrazole(PTZ).After successful modeling(21 d),the epileptic mice were randomly divided into PTZ group,sodium valproate(VPA)group,and low-dose(20 mg·kg^(-1))and high-dose(40 mg·kg^(-1))ginsenoside Rb_(1) groups.Mice in each group were given corresponding drugs intranasally for30 days,twice a day,and the control group was given the equal volume of normal saline.During the intranasa administration,the weight change,epilepsy latency,and epilepsy stage of the mice were recorded,and the changes in the electroencephalography(EEG)were recorded wirelessly.Neuronal Nuclei(NeuN)was used to observe the damage of neurons in cerebral cortex and hippocampus.The activation of microglia and astrocytes was observed with ionized calcium binding adapter molecule-1(IBA-1) and glial fibrillary acidic protein(GFAP),respectively.Glutamate(Glu)transporter-1(GLT-1)and glutamine synthetase(GS)were used to observe the key molecular changes in Glu regulation.Result:Compared with the control group,the PTZ group decreased body weight(P<0.05,P<0.01),shortened the epilepsy latency(P<0.01),and increased the epilepsy stage(P<0.01).The epileptic EEG waves were increased in the PTZ group.Compared with the PTZ group,the low and high-dose ginsenoside Rb_(1) groups increased body weight(P<0.05,P<0.01),prolonged the epilepsy latency(P<0.05,P<0.01),decreased the epilepsy stage(P<0.05,P<0.01),and decreased epileptiform EEG waves.Immunofluorescence staining(IF)showed that ginsenoside Rb_(1) significantly ameliorated PTZ-induced neuronal damage(P<0.05,P<0.01)in the motor sensory area of the cerebral cortex and hippocampal CA1 area,and significantly inhibited PTZ-induced activation of microglia(P<0.05,P<0.01)and astrocytes.Further research found that ginsenoside Rb_(1) significantly improved the expressions of astrocytic GLT-1(P<0.01)and GS(P<0.01)in the brains of epileptic mice.Conclusion:Intranasal ginsenoside Rb_(1) can significantly improve the symptoms of epilepsy caused by PTZ in mice,which has a clear protective effect on neuronal damage in the brains of epileptic mice and significantly inhibits the activation of brain microglia and astrocyte activation.Its anti-epileptic mechanism may be related to the regulation of GLT-1 and GS of the key molecules of astrocyte Glu metabolism.