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背景:α-氨基-3-羟基-5-甲基-4-异恶唑丙酸(AMPA)受体的过度激活在继发性脑损害的发生过程中起重要作用,研究表明,AMPA受体第二亚单位(GluR2)的减少是引起Ca2+快速内流主要原因,但缺血损伤后神经元膜表面AMPA受体数量及结构(亚单位组成比例)变化特征尚不清楚。目的:探讨缺血后神经元膜表面AMPA受体亚单位(GluRs)含量组成变化及其对Ca2+内流和细胞死亡的影响,为脑缺血治疗的特异性干预途径提供理论依据。设计:随机对照的实验研究。单位:解放军第三军医大学大坪医院野战外科研究所。材料:孕18dSD大鼠8只(解放军第三军医大学野战外科研究所动物所提供),进行胚胎海马神经元分离和培养。干预:缺血损伤组更换无糖细胞外液后置专用缺氧箱内缺氧30min,然后重新添加维持培养基继续培养,对照组在相同时刻更换正常培养基。采用PI染色、双重免疫荧光标记和细胞比色分析技术定量观察神经元死亡和膜表面GluRs含量变化,采用Fura-2法测定胞内Ca2+含量。主要观察指标:①伤后神经元死亡数目。②突触内和膜表面GluRs含量。③胞内Ca2+含量。结果:缺氧损伤后24hGluR2阳性突触数目构成比为0.42±0.16,与对照组(0.58±0.15)相比显著降低,差异有显著性意义(t=2.348,P=0.023),膜表面GluR2含量也显著减少,差异有显著性意义(t=2.427
BACKGROUND: Over-activation of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor plays an important role in the pathogenesis of secondary brain damage. Studies have shown that AMPA receptor The decrease of the second subunit (GluR2) is the main cause of rapid Ca2 + influx. However, the characteristics of the number and structure of AMPA receptors on the surface of neuronal membranes after ischemic injury are still unclear. Objective: To investigate the changes of the content of AMPA receptor subunit (GluRs) and its effect on Ca2 + influx and cell death in ischemic neuronal membrane, and to provide a theoretical basis for the specific intervention of cerebral ischemia. Design: Randomized controlled experimental study. Unit: Daping Hospital of PLA Third Military Medical University Field Surgery Research Institute. MATERIALS: Eight pregnant SD rats (provided by the Institute of Field Surgery, Third Military Medical University of PLA) were used to isolate and culture the embryonic hippocampal neurons. Intervention: The ischemic injury group was replaced with sugar-free extracellular fluid and placed in hypoxia for 30 minutes in hypoxia chamber. Then, the culture medium was added again to continue the culture, while the control group was changed to normal medium at the same time. The changes of neuronal death and GluRs on membrane surface were quantitatively determined by PI staining, double immunofluorescence labeling and cell colorimetric assay. The intracellular Ca2 + content was determined by Fura-2 assay. MAIN OUTCOME MEASURES: ① the number of neuronal death after injury. ② synaptic and membrane surface GluRs content. ③ intracellular Ca2 + content. Results: The ratio of the number of GluR2-positive synapses at 24 h after hypoxia injury was 0.42 ± 0.16, which was significantly lower than that of control group (0.58 ± 0.15) (t = 2.348, P = 0.023) Also significantly reduced, the difference was significant (t = 2.427