Changes Of P53 Expression In Hippocampus Of Pentylenetetrazole-kindled Rats

Objectives: Epilepsy, which is essentially an abnormality of cerebral function resulting from the discharges of some cerebral neuron in over high excitement state, has been a common clinical syndrome. The fast change of chemical substances in brain, such as enzymes, neurotransmitters and amino acids, and the apoptosis correlating genes can lead to brain injured after seizure. The mechanics of cell apoptosis after epileptic seizure is not known yet. P53 gene is an important anti-oncogene. In recent studies, p53 gene has a close connection with cell apoptosis. Rats were kindled by PTZ, and induced status epilepticus(SE), changes of behavior were observed. Meanwhile, the expression of p53 in the hippocampus was determined by reverse transcriptase polymerase chain reaction(RT-PCR),western blot and immunohistochemistry method in SE group and normal control group. To approach the apoptosis correlating pathway mediated by p53, can explore the control mechanism of p53 gene in epilepsy.Methods: Use status epilepticus(SE) model induced by PTZ intraperitoneal injection, 120 mature male Sprague- Dawley(SD) rats (weight 200±20g) were separated into two random groups: status epilepticus group and normal control group. Status epilepticus group is divided into 1, 6, 12,24,72h subset groups. Each group includes 20 rats. Rats were injected by 1%PTZ 40mg/kg, 20mg/kg after 10 mins, 10mg/kg after 10 mins. According to Racine criterion, seizure intensity was divided into 6 grades: grade 0: any seizure was not observe grade I: a little tingling of beards and twitching face; grade II: nodding, chewing accompanying with twitching face or wet dog shakes; grade III: raising one of a forelimb and clonus; grade IV: standing accompanying with bilateral forelimbs; grade V: standing accompanying with falling down, generalized tonic-clonic seizure. If 3 successive seizures of grade IV or V were observed, and SE lasted till 30mins, the lighting was to be successful. while the NC rats were injected by normal sodium. Changes of behavior were observed. Meanwhile,the expression of neurotrophin receptor p53 in the hippocampus was determined by RT-PCR and western blot of SE group and normal control group. The changes of p53 at CA1 and CA3 of hippocampus were observed by immunohistochemistry method.Results:1 results of epilepticus mode: SE group including 100 rats, 16 rats use 40mg/kg, 54 use 60mg/kg,25 use 70 mg/kg,5 use 80mg/kg, began to stare, nod, wash their face and twitching face. Accompanied by injecting every 10mins, their muscle began to clonus, their forelimbs began to uplift and tonic clonic seizure at last. The SE group rats recurrently attacking reached to over 30mins, and comforted to the SE criterion. 34 rats died resulting of tonic clonic seizure. The death rate was 34%. The NC group rats did not appear epileptic seizure.2 The expression of p53 mRNA in the hippocampus: The expression of p53 began to increase at 6 hours following SE(P<0.01), and reached peak at 24 hours(P<0.001),and began to decrease slowly at 48 hours(P<0.05),and decreased significantly after 72 hours(P<0.01), but still higher than normal control group.3 The expression of p53 protein in the hippocampus: The expression of p53 protein began to increase at 6 hours following SE(P<0.05), and reached peak at 24 hours(P<0.001), and began to decrease slowly at 48 hours(P<0.01), The changes of p53 protein is according with p53 mRNA in the hippocampus.4 Changes of immunological reaction at CA1 and CA3 of hippocampus: the immunological reaction of p53 protein is very weak at 6 hours following SE(P<0.05), and reached peak at 24 hours(P<0.001), expressing both at CA1 and CA3 of hippocampus. The immunoreactant was brown and most of them were in the nucleus of the neuron. The immunological reaction of p53 is insignificant in NC group.Conclusion:1 PTZ kinding by intraventriculatal injection provides a validated model of postseizure dysfunctioning, which is similar to human being of tonic clonic seizure.2 The expression of p53 began to increase at 6 hours following SE, and reached peak at 24 hours,and began to decrease slowly. p53 gene and protein probably take an important part of neuronal apoptosis in PTZ-induced seizures.