Low-frequency Hippocampal Stimulation Decreased The Lamininβ1 Level And Inhibited The Mossy Fiber Sprouting In Pharmacoresistant Temporal Lobe Epileptic Rats

Objective:To establish the lithium-pilocarpine induced pharmacoresistant model of epilepsy.The low-frequency hippocampal stimulation was performed and the pathologic changes of hippocampal mossy fiber sprouting,as well as the lamininβ1 expression was abserred.So as to explore,the possible mechanism of the low-frequency hippocampal stimulation.Methods:A total of 135 healthy male SD rats were selected to prepare epileptic model by lithium-pilocarpine intraperitoneal injection.Fourteen days after the successful preparation of the model,the rats with spontaneous seizures were respectively received intraperitoneal injection with phenobarbital sodium and intragastric administration with carbamazepine. Morphological change was taken as the index. Based on the epileptic seizure level, duration, electroencephalogram(EEG) changes, and frequency, the pharmacoresistant epileptic rats were selected out according their responses to phenobarbital and carbamazepine.The pharmacoresistant epileptic rats were assigned to a drug-resistant group or a hippocampus stimulation group. The behavioral changes and EEG changes of the rats were observed to evaluate the therapeutic effect of the hippocampus stimulation at hippocampus. After the treatment was completed, the hippocampi of the rats were extracted and examined through HE staining, Timm staining, and the ultrastructures were observed under the electron microscope, and the average optical density. Expression of laminin beta 1 were detected by Western blotting and a immunohistochemical method.Results:1. Among the 135 male SD rats,the process of kindling was completed in 102 ones, with the degree of seisures at V stage.Thirty-three rats died during the experiment.Fourteen days later, spontaneous seizures occured in 81 rats, with a successful kindling rate of 79.4%. Through the phenobarbital sodium and carbamazepine selection,18 drug resistent rats were selected out, with a drug resistance rate of 22.2%. With the rats chronically resistent to phenobarbital sodium and carbamazepine, the lithium-pilocarpine model of epilepsy was successfully established.2.After two weeks of hippocampus stimulation,the seizure degree and the duration of the drag resistent rats were decreased significantly compared to the those before the therapy. The differences were of statistic significance(P< 0.05). Significant complication didn’t occur in the treatment, and treating the drug resistent epilepsy by stimulation at hippocampus was safe and effective.3.Through the HE staining, electron microscopy observation, and the pathological observation of the CA3 area of hippocampus of rats in the drug resistent group and the drug sensitive group, it was shown that in the drug sensitive group, the cells were with sharp outline, in alignment; in the drug resistent group, the cells were with degeneration and necrosis, swelling, and disorganized, and cells in the hippocampus were arranged with loose structure, with nucleus deformation.4. Through Timm staining, the mossy fiber sprouting in the CA3 area of hippocampus of rats in the drug resistent group and the drug sensitive group was observed. In the drug resistent group, the mossy fiber sprouting in the inner molecular layer of the dentate gyrus and the lower cone layer of the CA3 area was increased. In the hippocampus stimulation group, it was shown that electric stimulation can prevent the mossy fiber sprouting to a certain degree.5. Through the immunohistochemical method,the changes of laminin β1 expression of hippocampus of rats in the drug resistent group under the electric stimulation were observed. By comparing the drug resistent group with the drug sensitive group, laminin β1 expression increased significantly; by comparing the hippocampus stimulation group with the drug resistent group, laminin β1 expression at the body surface was decreased significantly(P< 0.05).6. Through the Western blotting test, the lamininβ1 protein in the drug resistent group was increased compared to that in the drug sensitive group; the difference was of statistic significance(P<0.05).Conclusion:Low-frequency hippocampal stimulation could inhibit the seizure frequency and the seizure degree; decreas the laminin β1 expression as well as the mossy fiber sprouting These changes would contribute to the mechanism of the electric stimulation at hippocampus.