Effect Of M Channel Opener Retigabine On The Firing Of Hippocampal Dentate Gyrus Granule Cells In Mice With Acute Epilepsy And Its Anti-epileptic Effect

Objective:To investigate the effect of M channel activator retigabine on the electrical activity of granulosa cells in the dentate gyrus of the hippocampus;to explore the antiepileptic effect and protective effect of M channel activator retigabine on hippocampal injury.Methods:1.The brains of ICR mice were taken for electrophysiological experiments after acute epileptic seizure.The discharge of granulosa cells in the dentate gyrus of the hippocampus was recorded by whole-cell patch-clamp current clamp;the M current changes of granulosa cells were recorded by whole-cell patch-clamp voltage clamp;Tissue,Western Blot and immunofluorescence were used to detect the expression of KCNQ 2/3.2.After ICR mice experienced acute epileptic seizures,the brains were taken for electrophysiological experiments,and the effects of retigabine on the firing frequency of granulosa cells in the dentate gyrus of the hippocampus were recorded by whole-cell patch-clamp current clamp.3.Observation of the antiepileptic effect of M channel activator retigabine in ICR mice: ICR mice were randomly divided in blank group（Control）,epilepsy group（epilepsy,EP）,low（2 mg/kg）,high（10 mg/kg）dose retigabine group（Retigabine,RGB）,phenobarbital sodium group（Phenobarbital）.The above treatments were given during the acute attack,and the anti-epileptic effect of retigabine was observed by analyzing the seizure state and EEG recording.One week after the acute epileptic seizure,the cognitive changes of mice were detected by novel object recognition experiment,the excitation state of neurons in each region of the hippocampus was observed by c Fos staining,the damage of neurons in each region of the hippocampus was observed by HE staining,and the survival rate was analyzed to evaluate the safety of the drug.Results:1.Whole-cell mode current-clamp recording: compared with the blank group,the firing frequency of granulosa cells in the hippocampal dentate gyrus increased in the epilepsy group,and the difference was statistically significant（P<0.05）;whole-cell mode voltage-clamp recording: compared with the blank group group,the M current of granulosa cells in the hippocampal dentate gyrus decreased during acute epileptic seizure,and the difference was statistically significant（P<0.0001）.The expression decreased,and the difference was statistically significant（all P<0.01）.2.Current-clamp recording in whole-cell mode: Compared with the epilepsy group,the firing frequency of granulosa cells in the dentate gyrus of the hippocampus was decreased after the application of retigabine,and the difference was statistically significant（P<0.05）.3.Analysis of seizure behavior: low-dose retigabine has a better anticonvulsant effect;EEG results show that: the hippocampal discharge frequency increases during epilepsy,and the effect of low-dose retigabine on central electrical activity is significantly improved;4.The results of c Fos showed that: after acute epileptic seizure,the neurons in the CA1 area of the hippocampus of mice continued to be excited,mainly concentrated in the CA1 area.Application of retigabine in acute attack has a certain anti-epileptic effect and improves the continuous activation of neurons in CA1 area.5.Histological HE results showed that low-dose retigabine significantly improved neuronal damage in CA1 area.6.The results of new object recognition showed that: the mice in the epilepsy group had cognitive dysfunction,it was difficult to recognize and recall familiar objects,and they did not have a preference for new objects during the test.The preference index（DI）for novel objects was lower than that of the blank group,and retigabine could improve cognition in mice,and the low-dose retigabine group was better than the high-dose group.7.The 7-day survival rate of each group showed that the survival rate of the low-dose retigabine group was higher than that of the high-dose retigabine group.Conclusions:1.After acute epileptic seizures in mice,the firing frequency of granulosa cells in the hippocampal dentate gyrus increased;the expression of KCNQ2/3 channel proteins decreased,and the M current decreased.This change may be caused by excessive discharge of granulosa cells in the hippocampal dentate gyrus during epilepsy.reason;2.The M channel activator retigabine can reduce the excitability of granulosa cells in the hippocampal dentate gyrus;3.Low-dose application of M channel activator retigabine can play a good antiepileptic effect,reduce neuronal excitability,reduce hippocampal neuronal cell damage,and then improve cognition and play a role in brain protection.The anti-epileptic effect and the protective effect on hippocampal neurons of high-dose retigabine were not significant.