| Objective Epilepsy(EP)is a common central nervous system disease,which is a chronic brain disease characterized by a persistent tendency to produce seizures.In China,the overall prevalence of epilepsy is 7 ‰,most of which have onset in childhood,and the occurrence of epilepsy has a huge impact on the quality of life and learning activities of children,but also causes a serious burden on their families and society.As a multi-etiological disease,genetic factors,brain trauma,tumor growth,infection,and malformations can cause epilepsy.Despite the great variability in etiology,mitochondrial dysfunction and kinetic changes have attracted more and more attention of domestic and foreign scholars and become a new hotspot in epilepsy research.During epileptogenesis,cells can produce large amounts of free radicals,leading to oxidative stress damage and neuronal apoptosis,affecting mitochondrial function.On the other hand,mitochondria act as highly dynamically changing organelles by constantly dividing and fusing in order to maintain the normal function of the cell.Among them,inhibition of dynamin-related protein 1(Drp1)and Fis1 regulate mitochondrial division and play a key role in mitochondria-dependent apoptotic pathways,while mitochondrial split protein inhibitor(Mdivi-1)affects mitochondrial division by inhibiting the action of Drp1.At present,there are few reports on the role and mechanism of mitochondrial dynamics in neuronal cell injury during seizures at home and abroad.Are there age-dependent effects of mitochondrial division on epileptic neural cells? Does Mitochondrial Mitogen-Inhibitor Protect Neuronal Cells in Seizures?And the mechanism of the protective effect is still not fully elucidated.Therefore,in this study,we established a juvenile rat model of pentylenetetrazole(PTZ)chronic kindling epilepsy,observed the effects of Mdivi-1 on the behavior of juvenile epileptic rats,and detected the expression of Drp1 and caspase-3 genes and proteins in the hippocampus,so as to investigate the protective effect of Mdivi-1 on hippocampal neuronal injury in juvenile pentylenetetrazole-induced epileptic rats.Methods 90 clean level Wistar male rats(3-4 weeks,weight 50-70g)were randomly divided into group A(normal control group,n=30),group B(epilepsy group,n=30)and group C(Mdivi-1 group,n=30).The experimental groups(groups B and C)were intraperitoneally injected with PTZ at a body weight of 40 mg/kg,and the group A was given the same amount of normal saline.After successful modeling of 2 weeks of chronic kindle,the intervention group was intraperitoneally injected with Mdivi-1(1.2mg/kg)30 minutes before PTZ injection;the control group and the non-intervention epilepsy group were intraperitoneally injected with an equal volume of saline at the corresponding time points.The behavior of the rats was observed for 2 hours after injection,and their behavioral performance was recorded.After the completion of the intervention,pentobarbital was intraperitoneally injected for anesthesia at 3 h,24 h,and72 h.The hippocampus was isolated by decapitation after taking trunk blood.Drp1 and caspase-3 protein expression levels were detected by Western-blot method,Drp1 and caspase-3 gene level expression was detected by RT-PCR method,and immunohistochemistry of hippocampal tissues for rats in each group.Results1.Behavior of young rats: the rats in normal control group moved freely after normal saline injection,and there was no seizure behavior appeared,different degrees of seizures were occurred in rats with epilepsy after PTZ injection.With the extension of modeling time,the level of attack gradually increased,eventually reaching the standard of chronic ignition.There was no significant difference in seizure latency and seizure time between group B and group C after administration of Mdivi-1.2.RT-PCR results: Compared with the control group,the expressions of Drp1 and caspase-3 m RNA in the cells of group B and C were increased after PTZ kindling epilepsy model(P < 0.05),and the expressions of Drp1 and caspase-3 m RNA in group C were decreased compared with those in group B at each time point after Mdivi-1intervention(P < 0.05),with no significant difference compared with group A(P > 0.05).There were no significant differences in Drp1 and caspase-3 m RNA expression between the time periods of 3 h,24 h,and 72 h within group A(P > 0.05),Drp1 and caspase-3m RNA expression at 24 h in group B was significantly higher than that at 3 h and 72 h(P < 0.05),and Drp1 m RNA expression in group C peaked at 24 h,but caspase-3m RNA expression was not significantly different at each time period(P > 0.05).3.Western-blot results: Compared with group A,the expression levels of Drp1 protein in the hippocampus of group B were significantly increased at each time point,and the difference was statistically significant(P < 0.05);the protein expression levels of group A were not statistically different at each time point(P > 0.05);the protein expression levels of group C were significantly lower than those of group B at each time point(P < 0.01),and there was no significant statistical difference with group A(P >0.05).The results of caspase-3 protein expression between the groups were similar here.4.Immunohistochemical results: Compared with the control group,the number of Drp1 positive cells in the hippocampus of rats in group B was significantly increased at each time period(P < 0.01),while the number of positive cells in group C was less than that in group B(P < 0.05).The number of caspase-3 positive cells in the hippocampus of rats in group B was more than that in group A(P < 0.05),while the number of positive cells in group C was less than that in group B(P < 0.05).Conclusions1.The levels of Drp1 and Drp1 m RNA of hippocampal were increased after epileptic seizures in young rats,which was demonstrating the involvement of mitochondrial division in the process of pathological injury in chronic epilepsy,so the change of mitochondrial dynamics may be related to epileptogenesis in children;2.Mdivi-1 had a neuroprotective effect on hippocampal neurons in epileptic young rats;The mechanism may be related to the Cytochrome C signal transduction pathway and caspase-3 activation;3.Inhibition of mitochondrial division could become a new thought and method for the treatment of refractory epilepsy in children. |
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