Effects Of Mengshi Guntan Tang On Expression Of Pentylenetetrazole-induced Epileptic Mice

Part One:The influence of Mengshi guntan Tang on epileptic mice 's EEG and expression of NMDAR1 and GABA-ARPurpose:To establish pentylenetetrazole(PTZ)chronic kindling model and observe the influence of Mengshi guntan Tang on mice's behavior,EEG,expression of hippocampus NMDAR1 and GABA-AR.Method:PTZ kindled epileptic model mice were randomly divided into four groups:the model group:group B;low-dose Mengshi guntan Tang group:group C;high-dose Mengshi guntan Tang group:group D;sodium valproate group:group E.Each group has 10 mice.A blank control group of 10 mice,group A,was also set.Group B were given normal saline,while group C,D,E were respectively given low and high dose of Mengshi guntan Tang,sodium valproate and other different drug gavage.After 4 weeks of administration,except the control group,each group of mice were given an intraperitoneal injection of PTZ 45mg/again(kg·d)for observation of the changes in animal behavior.We used stereotaxic apparatus to record the mice 's EEG changes on the right side of frontal cortex.We removed he hippocampus of mice for determination of the content of glutamate(Glu)and ?-amino butyric acid(GABA)by amino acid kit and enzyme linked immunosorbent assay(ELISA)method,and the expression level of NMDAR1 and GABA-AR by western blot.Result:1.Behavioral performanceCompared with group B,mice in Group D and E significantly prolonged the latency of seizures(P<0.05),and reduced the number of 1h tonic clonic seizures(P<0.05)with shorter duration of seizures(P<0.05).Group D and E showed no significant difference(P>0.05)in comparison.Group A had no seizures.2.EEG performanceEEG of group A showed basic brain waves,mainly alpha and beta waves,with stable amplitude and no ob-vious rhythmicity.Group B showed obvious paroxysmal epileptic discharge,mainly sharp peaks of high amplitude and short burst of sharp wave,spike wave,spike and slow wave complex.Compared with group B,group C and D significantly reduced the average amplitude(P<0.05),and group D showed the first significant amplitude after I5min.Group E showed occasional epileptic waves after treatment,all within 1 mV.3.Amino acid detection and western blot resultsGlu content change:Compared with group A,Glu in hippocampus of group B increased significantly(P<0.05).Compared with group B,group C,D and E decreased significantly(P<0.05).Group D and E showed no statistically significant difference(P>0.05).GABA content change:The content of GABA in group D was increased most significantly,and group B,C and E had significant difference(P<0.05).The increase in group D showed statistically significant difference from group E(P<0.01).Through detection of expression of NMDAR1 and GABA-AR by western blot,it was found that NMDARI in group B was significantly higher than that in group A,while GABA-AR in group B was significantly lower than that in group A(P<0.01).NMDAR1 in group C,D and E group was significantly lower than that in group B(P<0.05),while GABA-AR in group C,D and E group was significantly higher than that in group B(P<0.05).There was no significant difference between group D and E.Conclusion:1.Mengshi guntan Tang can prolong the incubation period of mice with seizures,shorten the onset time,reduce the attack level,suppress the epileptic discharge amplitude and suppress epileptiform discharge frequency;2.Mengshi guntan Tang can reduce the content of Glu,inhibit the expression of NMD AR1,increase the content of GABA,promote the expression of GABA-AR,correct the imbalance of excitatory amino acid and inhibitory amino acid neurotransmitter.Part Two:The influence of Mengshi guntan Tang on the expression of GFAP in hippocampus of epileptic micePurpose:To establish PTZ chronic kindling model and observe the influence of Mengshi guntan Tang on the expression of glial fibrillary acidic protein(GFAP)in hippocampus of mice.Method:Modeling and grouping are the same as Part One.We put removed hippocampus of mice in paraformaldehyde overnight,and then removed it after fixation to make frozen sections by frozen section machine for GFAP immunohistochemical staining.Then we observed the form and quantity of GFAP in district CA1 and CA3 of hippocampus,and then detected the expression level of mRNA GFAP using real-time fluorescence quantitative(Q-PCR)method.Result:Compared with group A,group B showed significantly increased GFAP immunoreactivity,significantly increased quantity of hippocampus,and enhanced expression of GFAP mRNA,which was statistically significant(P<0.05).Compared with group B,GFAP immunoreactivity in group C,D and E was weaker and less than the model group.The expression of GFAPmRNA in group C,D and E was significantly reduced(P<0.05),and the group of low-dose Chinese medicine showed better effect than the high-dose group,but showed no significant difference from the positive control group,(P>0.05).Conclusion:Mengshi guntan Tang can reduce the quantity and inhibit the over expression of GFAP in the hippocampus of PTZ kindled mice,which may be one of mechanism of Mengshi guntan Tang in anti-epileptic.