The Effect Of Pinellia Total Alkaloids On EcoG,CEP And Cerebral GABA,Glu Receptor In Epileptic Rats

Objective To establish rat cortical evoked potential(CEP)and epileptiform discharge(ED) model and testify that the change of CEP can be a new electrophysiological index which may assess epileptogenic animal and the pharmacodynamics of antiepileptic drugs.Then,we investigate the effect of pinellia total alkaloids(PTA)on rat electrocorticogram(EcoG)and CEP,analyse the influence of PTA on GABA,Glu receptor and expression of cerebral GABA_A receptor mRNA,and explore the receptor mechanism of antiepileptic ation of PTA.Methods(1)Electrical stimulation of sciatic nerve and penicillin-evoked,glutamate-induced rat epileptic model were respectively applied to analyse the effect of PTA 400mg·kg^-1given by sublingual iv(siv)on latency,frequency and amplitude of ED and amplitude of CEP,simultaneously recording EcoG and CEP of epileptic rats with RM6240C multichannel biological signal processing system.(2)Correlation analysis between frequency of ED and amplitude of CEP in PNC,Glu,PTA+PNC and PTA+Glu groups were fulfilled by Pearson correlation analysis.(3) GABA and AP₅ were positive control;Glu and PNC were tool drugs;Amplitude of CEP was obs erving index.Rat CEP model was used to draw dose-effect curve by siv cumulatingly different doses of GABA,AP₅ and PTA.Observe the impact of Glu and PNC on dose-effect curve of each drug,compute their IC₅₀respectively and compare inhibitory intensity of the three drugs on amplitude of CEP.Evaluate the action of PTA on GABA,Glu receptor by analyzing doseeffect curve.(4)PNC-chronic kindling rat model and reverse transcriptase-polymerase chain reaction(RT-PCR)were adopted to observe the effect of two doses of PTA on seizure latency,degree and expression of cerebral GABA_A receptor mRNA.Results(1)After electrical stimulation of sciatic nerve,typical CEP was recorded in sensory area of contralateral cerebral cortex.EcoG appeared pronounced ED when given PNC,Glu by lateral cerebral ventricle.When frequency of ED increased,the amplitude of CEP augmented correspondingly.PTA could prolong latency of ED,decrease frequency,amplitude of ED and amplitude of CEP synergistically induced by electrical stimulation of sciatic nerve and PNC,Glu.Compared to PNC,Glu group,there were significantly statistical difference(P＜0.01).(2) Correlation coefficient r between frequency of ED and amplitude of CEP in PNC group was 0.947(P＜0.01),and r value of PTA+PNC,Glu,PTA+Glu group were 0.939(P＜0.01),0.930 (P＜0.01),0.864(P＜0.05),respectively.(3)GABA,AP₅ and PTA could all dose-dependently inhibit amplitude of CEP.Dose-effect curve of GABA and PTA moved towards right when given PNC,GABA receptor antagonist;But,the efficacy of GABA stabilized and that of PTA descended.Dose-effect curve of AP₅ and PTA moved towards right when given Glu;But,the efficacy of AP₅ didn't change and that of PTA dropped.(4)Compared with noraml rats, expression of cerebral GABA_A receptor mRNA in PNC chronic kindling rat decreased;there was markedly statistical difference between them(P＜0.01).Pre-treatment of PTA could both prolong seizure latency,and there was respectively statistical difference compared with model group(P＜0.01).PTA could also lessen seizure degree and significantly increase expression of GABA_A receptor mRNA.There were statistical difference compared to model group(P＜0.05, P＜0.01),no difference with normal group.Conclusions(1)Frequency of ED and amplitude of CEP induced by PNC,Glu and affected by PTA had very good correlation by Pearson correlation analysis.Therefore,it suggested that the change of CEP amplitude can not only be a new electrical index which may assess epileptogenic animal and seizure degree,but also be a new index of evaluating pharmacodynamics in animal experimental research of antiepileptic drugs.(2)PTA could markedly prolong latency of PNC-induced ED.Meanwhile,PTA decreased frequency and amplitude of spike wave,and amplitude of CEP producing inhibitory action on paroxymal depolarization shift(PDS).Its mechanism may be associated with enhancement of GABAergic inhibitory function.(3)PTA could significantly oppose Glu-evoked seizure.It prolonged latency of ED,decreased frequency and amplitude of ED and amplitude of CEP.All these effects could be the evidence that PTA lowered the function of Glu excitatory system in animal cerebrum, regulated the balance of inhibitory and excitatory function in CNS,and limited the development and diffusion of ED.(4)GABA,AP₅ and PTA could all inhibit amplitude of CEP in dose-dependent way.GABA and PNC competitively antagonized GABA receptor.Compared to GABA,PNC could decrease the maximal efficacy of PTA.It suggested that PTA and PNC non-competitively antagonize GABA receptor.AP₅ and Glu competitively antagonized Glu receptor,but PTA and Glu non-competitively antagonized Glu receptor.Inhibitory intensity of the three drugs were:AP₅＞GABA＞PTA.The non-competitively antagonizing mechanism may be associated with:â‘ Binding site of PTA and PNC/Glu in receptor is different.When PNC/Glu binds with corresponding receptor,its conformation changes,which hinders binding of PTA with receptor.So,the maximal efficacy of PTA drops.â‘¡PTA affects other target during receptor signal transduction.â‘¢The antiepileptic action of PTA has no direct relationship with GABA receptor and Glu receptor.It brings about antiepileptic effect through other unknown mechanisms.(5)PTA could prolong seizure latency,decrease seizure degree and antagonize PNC chronic kindling rat seizure.It is postulated that its antiepileptic mechanism is associated with expression increase of cerebral GABA_A receptor mRNA,upregulation of GABA_A receptor numbers and enhancement of cerebral inhibitory function.