N-甲基-D-天冬氨酸受体阻断对大鼠海马神经元癫痫样放电模型泛连接蛋白-1表达的影响

发布时间：2018-04-15 01:25

本文选题：癫痫 + N-甲基-D-天冬氨酸(NMDA)受体　；参考：《中国老年学杂志》2017年23期

【摘要】：目的观察N-甲基-D-天冬氨酸(NMDA)受体阻断对大鼠海马神经元癫痫样放电模型神经元细胞活力,凋亡及泛连接蛋白(Panx)-1表达的影响。方法取SD新生乳鼠双侧海马,体外培养海马神经元至第9天,经鉴定的海马神经元随机分为正常对照组、模型组和MK-801(地卓西平,NMDA受体拮抗剂)组。正常对照组为正常细胞的细胞培养液培养3 h后恢复维持培养液;模型组为无镁细胞外液培养3 h后恢复维持培养液;MK-801组加入含10μmol/L MK-801维持培养液预保护30 min,更换为无镁细胞培养液培养3 h后恢复维持培养液。模型建立24 h后,采用细胞计数试剂盒(CCK-8)检测神经元细胞活力,分别采用末端脱氧核苷酸转移酶介导的d UTP缺口末端标记测定法(TUNEL)法和流式细胞仪膜联蛋白V-异硫氰酸荧光素/碘化丙啶(Annexin V-FITC/PI)双染法检测神经元凋亡,采用蛋白质印迹法(Western印迹)检测神经元Panx-1表达,采用实时荧光定量PCR(Real Time-PCR)检测神经元Panx-1 mRNA表达。结果与模型组比较,MK-801组神经元细胞活力显著升高(P0.05),神经元凋亡指数和凋亡率、Panx-1表达、Panx-1 mRNA表达显著降低(P0.05)。结论 NMDA受体阻断可抑制大鼠海马神经元癫痫样放电模型神经元损伤,发挥神经元保护作用,并抑制神经元Panx-1表达。
[Abstract]:Objective to investigate the effects of N- methyl-Daspartic acid (NMDA) receptor blockade on neuronal viability, apoptosis and Panx-1 expression of hippocampal neurons in epileptiform discharges model.Methods the hippocampal neurons of neonatal SD rats were cultured in vitro for 9 days. The identified hippocampal neurons were randomly divided into normal control group, model group and MK-801 (didroxepine NMDA receptor antagonist) group.The normal control group was cultured in the normal cell culture medium for 3 hours and then recovered to maintain the culture medium.The model group was treated with Mg-free extracellular medium for 3 h and then restored the maintenance medium (MK-801) for 30 mins with 10 渭 mol/L MK-801. The MK-801 group was replaced with Mg-free cell culture medium for 3 h and then restored to the maintenance medium.After the model was established for 24 hours, the cell viability was detected by the cell count kit CCK-8.Neuronal apoptosis was detected by terminal deoxynucleotidyl transferase-mediated d UTP Nick end labeling (Tunel) assay and flow cytometry (FCM) double staining of fluorescein Visothiocyanate / Annexin V-FITC / Pi.The expression of Panx-1 in neurons was detected by Western blot, and the expression of Panx-1 mRNA in neurons was detected by real-time quantitative PCR(Real (Time-PCR).Results compared with the model group, the neuronal viability of MK-801 group increased significantly (P 0.05), and the expression of Panx-1 mRNA decreased significantly (P 0.05).Conclusion blockade of NMDA receptor can inhibit neuronal injury, play a protective role and inhibit the expression of Panx-1 in hippocampal neurons.
【作者单位】：牡丹江医学院附属红旗医院神经内科;
【基金】：黑龙江省卫生计生委科研课题(2016-382) 黑龙江省中医药科研项目(2HY16-019) 牡丹江市科学技术计划项目(Z2016s0084)
【分类号】：R742.1

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