| Objective:Epilepsy is the one of the most common neurological disorders caused by suddenly recurrent abnormal discharge of neurons, leading to temporary dysfunction in the central nervous system. Drug treatment is currently still dominated but fails to effectively control all seizures despite of some progress in regular and new antiepileptic drugs. After regular drug therapy, About one-third of individuals with epilepsy still fail to achieve good conctol effect and develop into refractory epilepsy. Many studies have reported that frequent and continuous seizures can result in neurons apoptosis in the brainwhich play an important role in the formation of chronic epilepsy and epilepctic brain injury. In recent years, more and more reseachers have focused on medicinal plants. Ginsenoside Rgl (G-Rgl), one of the major bioactive ingredients of Panax plant such as ginseng and sanqi, has been shown to possess anti-apoptosis, anti-inflammatory and anti-oxidant effects. Based on above, it is reasonable to hypothesize that G-Rg1 may have the anti-epileptogensis role. Based on the supposition above, rats model of epilepsy were induced by intraperitoneal injection of pilocarpine combine with lithium chloride. To observe nerons apoptosis in hippocampus of epilepsy model after intrapentioneal injection with pilocarpine. To analysis G-Rgl effects on nerons apoptosis in hippocampus. To explore the optimal concentration of G-Rgl and the molecular mechanism of possible neural protection on epilepsy.Methods:One hundred adult male Sprague-Dawley (SD) rats (weight 285±15 g) were randomly divided into five groups:blank control group, pilocarpine-induced epileptic group and different does of G-Rg1 treatment group (50mg/kg,100 mg/kg and 200 mg/kg).20 rats in each group. The treatment group received different dose of G-Rgl 6 hours by the way of intraperitoneal injection before the epileptic model preparation and others were injected intraperitoneal with isometric normal saline (0.9% NaCl solution) at the same time. After 6 hours, in pilocarpine-induced epileptic group and G-Rgl treatment group, lithium chloride (127 mg/kg) was treated by the way of intraperitoneal injection followed by atropine sulfate (1 mg/kg) after 18 hours to reduce the cholinergic peripheral reaction and received pilocarpine (30 mg/kg) 30 min later. The rats of blank control group were injected intraperitoneal with the isometric normal saline same as pilocarpine. We can inject atropine sulfate (1 mg/kg) 30 min earlier that it is could reduce the cholinergic peripheral reaction. Observing the behavior changes of rats after intrapentioneal injection with pilocarpine. According to Racine grading standard, reaching level III or more can be considered as a seizure. Injected intraperitoneal with diazepam (10 mg/kg) terminates the attack after seizures lasted 90 min and taked care of rats to reduce the death.All rats were anesthetized with 10% chloral hydrate at the dosage of 350 mg/kg by the way of intraperitoneal injection following successfully established epileptic models after 72 hours. Some of rats cut head to take brain, and isolate the hippocampal tissue. Samples placed in-80℃ refrigerator for further study. The proteins expression of Bcl-2 and Bax were detected with western blotting method. Some of rats were transfused with 4% paraformaldehyde solution for fixing the organizations of rats. The brain tissue of rats was removed, and it was treated with 30% sucrose solution for dehydration and with OCT for embedding. Then cutting 15 μm thick slices by freezing microtome were prepared for further study. The pathological changes of hippocampal neuron were observed and Bcl-2, Bax proteins were detected with immunohistochemical and immunofluorescent method.Results:Control group do not appear seizure and death. Compared with pilocarpine-induced epileptic group, seizure latentperiod was increased obviously after treatment of G-Rgl (P<0.05). After seizure induced by pilocarpine combine with lithium chloride, immunohistochemistry and immunofluorescence results show, the expression of Bax protein was increased in the cytoplasm of neurons, especially hippocampal neurons, and the expression of Bcl-2 protein was decreased. Compared with pilocarpine-induced epileptic group, the expression of Bax protein was lower and the expression of Bcl-2 protein was upper in G-Rgl group. The results of Western Blot was consistent with the immunohistochemistry and immunofluorescence and antiapoptotic effect was obvious at the concentration of 100 mg/kg.Conclusions:Seizures can cause the activation of apoptosis factors. It was found that the expression of Bcl-2 was up-regulate and Bax downregulated, and hippocampus neurons apoptosis in rats model of epilepsy induced by intraperitoneal injection of pilocarpine combined with lithium chloride. G-Rgl can obviously prolong seizure latent period and reduce mortality after rats induced by pilocarpine combine with lithium chloride. It could be reduce hippocampal neurons apoptosis through up-regulating the expression of Bcl-2 and down-regulating the expression of Bax. In conclusion, this study shows G-Rgl may play an important role of protecting effect on nervous system through up-regulating the expression of Bcl-2 and down-regulating the expression of Bax. |
