| Objective:To observe the effects of Shigan powder on the memory and learning ability,the morphology and structure of hippocampal neurons,the ability of anti-oxidative stress,and the changes of apoptosis-related gene protein and mRNA levels in pentylenetetrazol-induced epilepsy rats,to explore Shigansan's antiepileptic mechanism.Methods:A total of 120 healthy female SD rats were selected and 15 were randomly selected as blank group.The other rats were lighted with pentylenetetrazole(PTZ)by intraperitoneal injection to make model.The successful model rats were randomly assigned to the model group,the Valproate group,the Acorus gramineus soland group,the Gansong group,and the Shigan powder group,gavage was performed 1 day after the completion of the group.After four weeks of gavage treatment,the Morris water maze test(location cruising experiment and space exploration experiment)were given,and the materials were obtained after the end of the treatment.The contents of SOD,MDA and GSH-Px of the cerebral hippocampal tissue were determined after the samples were completely taken.Real-time PCR was used to detect the mRNA expression of AIF,Caspase-3,Cyt-C and Akt in rats'hippocampus.Western blot was used to detect the protein expression of AIF,Caspase-3,CytC,p-Akt and Akt in rats' hippocampus.The pathological changes of rat hippocampal neurons were observed by HE staining and Nissl staining.Results:1.The results of the Morris water maze navigation experiment show that:Shigan powder group,Gansong group and Acorus gramineus soland group can all shorten the escape latency of rats in each group.Among them,Shigan powder group works best(P<0.05),and there was no significant difference between Shigan powder group and Valproate group(P>0.05).In the space exploration experiment,compared with the model group,Shigan powder group,Gansong group and Acorus gramineus soland group can significantly increase the number of safely crossing the safe platform area in pentylenetetrazole induced epileptic rats(P<0.05).Shigan powder group rats crossed the platform more frequently than Acorus gramineus soland group and Gansong group rats(P<0.05).There was no significant difference in the number of crossing platform between the Valproate group and the Shigan powder group(P>0.05).2.Ability of antiepileptic:Compared with the model group,the activity of SOD and GSH-Px in the cerebral hippocampal tissue of Shigan powder group,Acorus gramineus soland group,Valproate group rats increased,and the production of MDA decreased(P<0.05).The activities of SOD,GSH-Px and MDA in the hippocampus of rats in the Gansong group were not significantly different from those in the model group(P>0.05).The effect of Shigan powder group is superior to Acorus gramineus soland group and Gansong group(P<0.05).There was no significant difference between Shigan powder group and Valproate group(P>0.05).3.Pathomorphology:?Results of HE staining showed:Compared with the blank group,the pyramidal cell structure and levels of model group were disordered,the number of cells was significantly reduced,the cell body was swollen and deformed,the outline was blurred,the cytoplasm was stained deeply and the nucleus was crushed and solidified.Compared with the model group,the number of pyramidal cells were increased in the Shigan powder group,the gap was significantly reduced,the arrangement was neat,the level was clear,the cytoplasm color was deeper,the nucleolus was slightly clearer,and the phenomenon of deep staining and shattering and fragmentation were improved.?Nissl staining results:Comparison with model group and blank group,cytoplasm Nissl bodies were absent or reduced,a small amount were scattered sporadicly.Compared with the model group,the Shigan powder group had a rich distribution of Nissl bodies in the cytoplasm.4.mRNA expression of Hippocampal tissue:Real-time PCR assay showed showed no significant difference in AktmRNA expression in hippocampus of rats in each group(P>0.05).Besides,comparing with model group:Shigan powder group,Gansong group and Acorus gramineus soland group could inhibit the expression of AIF,Cyt-C and Caspase-3mRNA in hippocampus of epileptic rats induced by PTZ(P<0.05).The expression of Shigan powder group was stronger than Acorus gramineus soland group and Gansong group(P<0.05).There was no significant difference between Shigan powder group and Valproate group(P>0.05).5.Gene protein expression of Hippocampal tissue:Western blot showed no significant difference in expression of Aktprotein in hippocampus of rats in each group(P>0.05).Compared with model group,Shigan powder,Gansong group and Acorus gramineus soland group could all up-regulate the expression of p-Akt protein in hippocampus of PTZ-induced rats and inhibit the expression of AIF,Cyt-C and Caspase-3 protein(P<0.05).The expression of Shigansan group was stronger than Acorus gramineus soland group and Gansong group(P<0.05).There was no significant difference between Shigan powder group and Valproate group(P>0.05).Conclusion:1.Shigan powder can significantly improve the cognitive function of PTZ-induced epilepsy rats and improve their ability to learn and remember.2.Shigan powder can improve the pathological morphology of hippocampus in PTZ induced epilepsy rats.It can increase SOD and GSH-Px activity in rat hippocampus,inhibit MDA production,to increase the ability of anti-oxidation of neurons.3.Shigan powder can increase the expression of p-Akt mRNA and protein in the hippocampus of PTZ induced epilepsy rats and inhibit the expression of AIF,Cyt-C,Caspase-3 mRNA and protein,so as to exert anti-apoptosis effect on neurons.This study confirmed that Shigan powder's antiepileptic mechanism is closely related to the protection of hippocampal neurons,increase of antioxidant capacity and decrease of neuronal apoptosis. |
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