Clarification Of The Role Of Glucocorticoids On Hippocampal Synaptic Actin Cytoskeleton Stablization In Epileptic Brain

In recent years,the important role of glucocorticoids in the clinical treatment of epilepsy has become increasingly prominent.Several studies have confirmed the antiepileptic and neuroprotective effects of glucocorticoids.However,the mechanism of glucocorticoid treatment in epilepsy has not been fully elucidated.Filamentous actin(F-actin)is a stable cytoskeleton component composed of actin monomers.It is also the main structure of presynaptic axon terminals and postsynaptic dendritic spines.Previous studies have confirmed that F-actin remodeling is closely related to nerve excitability abnormalities caused by synaptic remodeling in epileptic brain.Glucocorticoids have also been shown to act directly on synaptic and dendritic spines,regulating the remodeling of dendritic spines and synaptic structures.Therefore,F-actin may be a key target of glucocorticoid antiepileptic therapy.The study of the relationship between glucocorticoids and actin cytoskeleton stability,and the discussion of the possibility of the stable F-actin cytoskeleton blocking the formation and development of epilepsy,is expected to provide a theoretical basis for the treatment of epilepsy with glucocorticoids,and also provide a new idea for the development of new antiepileptic drugs based on the maintenance of the stability of the hippocampus actin cytoskeleton.Objective:The purpose of our experiment was to explore the protective effect of glucocorticoids on hippocampal neuronal synaptic actin cytoskeleton of epileptic brain,so as to provide a theoretical basis for the treatment of epilepsy with glucocorticoids and a new target for the treatment of epilepsy.Methods: In this study,healthy adult male ICR mice were selected to investigate the effect of dexamethasone on the remodeling of F-actin in the hippocampal synapses during the chronic and acute phases of pilocarpine induced status epilepticus(SE)model.(1)Chronic model: The mice were divided into Control group,PILO group and PILO+DEX group.Mice in PILO + DEX group were intraperitoneally injected with dexamethasone 10 mg /kg on day 1,2 and 3 after SE.The behavioral changes of spontaneous seizure in mice were observed in 28 days after SE,as the time point of chronic model.Immunofluorescence staining and laser confocal scanning were used to detect the changes of F-actin,neuron number and presynaptic/postsynaptic markers in each hippocampal region.This part of our study was to investigate the effect of dexamethasone on the remodeling of F-actin in hippocampal synapses during chronic phase of pilocarpine induced epileptic model.(2)Acute model: The mice were divided into Control group,PILO group and PILO+DEX group.Mice in PILO + DEX group were intraperitoneally injected with pilucapine and given dexamethasone 10mg/kg.The changes of F-actin,number of neurons,and presynaptic/postsynaptic markers in the hippocampus were observed in 1 hour after SE,as the time point of the acute phase,to study the relationship between glucocorticoids and hippocampal cytoskeletal stability in the acute phase of epilepsy.The RU486 group mice were pretreated by Mifepristone(RU486,a glucocorticoid receptor antagonist)to determine the F/G ratio and analyze the protein of p Cofilin.This part of our study was to investigate the effects of glucocorticoid receptor inhibition on the cytoskeletal dynamics of F-actin in epileptic brain,and to determine the pathway of glucocorticoid in brain.Results:(1)Effects of dexamethasone on the remodeling of F-actin in hippocampus during chronic phase of pilocarpine induced epileptic model:(1)Pilocarpin-induced spontaneous recurrent seizures occurred in the chronic epileptic model mice,and the duration of seizures in PILO+DEX group was significantly shortened than that in PILO group(P < 0.05).(2)Dexamethasone treatment can effectively alleviate F-actin injury of hippocampal neurons in pilocarpine induced epileptic model in the chronic phase.(3)Dexamethasone treatment could alleviate the loss of pyramidal cells in the hippocampal CA1 and CA3 regions in the chronic phase of pilocarpine induced epileptic model(p < 0.05),but had no significant effect on DGCs proliferation(p > 0.05).(4)Dexamethasone treatment can effectively prevent the reduction of PSD95 and Zn T3,but does not prevent the generation of new synaptic connections and the construction of abnormal synaptic loops.(2)Effects of dexamethasone on the remodeling of hippocampal F-actin in pilocarpin-induced SE model in acute stage:(1)Dexamethasone treatment significantly increased the incubation period of grade 3 seizures(P < 0.01);However,there was no difference between PILO group and PILO+DEX group in the number and level of seizures at or above grade 3(P=0.40,0.23).(2)Dexamethasone treatment significantly alleviated F-actin damage in the hippocampal neurons after SE(P<0.01),and this effect was antagonized by the glucocorticoid receptor antagonist RU486.(3)The number of pyramidal neurons in PILO group and PILO+DEX group did not change significantly after SE.(4)Dexamethasone inhibited the decline of PSD95 in hippocampal CA3 and CA1 regions,and stabilized the postsynaptic structure.Conclusion:(1)Dexamethasone dose of 10 mg/kg could prolong the incubation period of grade 3 epileptic seizure in mice during the pilocarpin-induced SE model.The duration of spontaneous recurrent seizures could be reduced in the chronic epileptic period.(2)Dexamethasone treatment could alleviate the death of hippocampal neurons in the chronic phase of epileptic model.(3)Dexamethasone treatment had a protective effect on the hippocampal neuronal synaptic actin cytoskeleton in the acute and chronic epileptic models.And dexamethasone treatment might contribute to maintaining the stability of the synaptic structure.(4)Dexamethasone might regulate the dynamic changes of F-actin in hippocampal neuron synapses by acting on the postsynaptic membrane glucocorticoid receptor.