摘要
应用红藻氨酸(kainicacid,KA)诱导的大鼠边缘叶发作癫痫模型,检测Bad(Bcl-2-associateddeathprotein)、14-3-3、磷酸化Bad、Bcl-XL和Bcl-2在癫痫大鼠海马神经元的表达。单侧杏仁核内注射KA诱导癫痫发作,持续记录脑电和局部脑血流(regionalcerebralbloodflow,r-CBF),发作1h后静脉注射30mg/kg安定终止发作,然后分别用cresylviolet染色和TUNEL染色观察海马神经元存活和凋亡的变化;用免疫荧光、Westernblot和免疫沉淀检测海马Bad、14-3-3、磷酸化Bad、Bcl-XL和Bcl-2的表达。结果表明,发作终止8h时出现TUNEL阳性细胞,24h时达高峰;发作诱导Bad去磷酸化,去磷酸化的Bad与分子伴侣蛋白14-3-3解离,然后Bad与Bcl-XL结合;磷酸化Bad表达减少而Bcl-2表达增加;发作前后r-CBF无明显变化。以上结果提示,癫痫发作诱导Bad的去磷酸化和Bcl-2表达上调,Bad的去磷酸化可能具有损伤作用,而Bcl-2的表达上调则对癫痫神经元损伤具有保护作用,但与脑缺血无关。
The purpose of the present study was to explore the seizure-induced changes in Bad (Bcl-2-associated death protein), 14-3-3, phosphoBad, Bcl-2 and Bcl-XL expression in the rat model of focal limbic seizure. Unilateral intra-amygdaloid injection of kainicacid (KA) was made to induce seizure. Electroencephalogram (EEG) and regional cerebral flow (r-CBF) were monitored continuously.Diazepam (30 mg/kg) was administered to terminate the seizure. The apoptotic and surviving neurons in the hippocampus wereobserved by terminal deoxynucleotidyl transferrase-mediated dUTP nick end labeling (TUNEL) and cresyl violet staining, the expres-sion of Bad, 14-3-3, phosphoBad, Bcl-2 and Bcl-XL were detected with immunofluoresence, Western blot and immunoprecipitation.The results showed that TUNEL-positive neurons appeared at 8 h and reached maximum at 24 h following seizure cessation within theipsilateral CA3 subfield of the hippocampus. Seizure induced the dephosphorylation of Bad and the dissociation of Bad from itschaperone protein 14-3-3 and subsequent dimerization of Bad with Bcl-XL. The expression of phosphoBad decreased and Bcl-2 increased.There was little change in r-CBF after the seizure. These results suggest that seizure leads to a dephosphorylation of Bad and anupregulation of Bcl-2. Dephosphorylation of Bad may be injurious while the upregulation of Bcl-2 may be protective to the brain damageinduced by seizures, but not related with r-CBF.
出处
《生理学报》
CAS
CSCD
北大核心
2005年第3期310-318,共9页
Acta Physiologica Sinica
