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背景:海马CA1区锥体细胞是对缺血缺氧性损伤最为敏感的神经元,缺血缺氧后海马CA1区锥体细胞膜电位表现为缺氧早期细胞膜的超极化,随着缺氧时间的延长,细胞膜发生缓慢去极化及快速去极化,引起神经元不可逆性损伤。目的:应用细胞内记录技术,观察N-甲基-D-天冬氨酸受体阻断剂MK801对离体海马脑片CA1区锥体细胞缺氧期间电生理指标的变化。设计:观察对比实验。单位:解放军第九七医院,徐州医学院江苏省麻醉学重点实验室,Healthy-ScienceCenter,StateUniversityofNewYork。材料:实验于2002-09/2003-02在美国纽约州立大学医学中心完成。选择成年雄性SD大鼠5只,预吸纯氧3min后以体积分数为0.02的异氟醚麻醉,快速断头取脑,制备离体海马脑片。方法:大鼠海马脑片随机分为单纯缺氧组和MK801组,每组10个。单纯缺氧组海马脑片给予10min缺氧;而MK801组的海马脑片在缺氧前10min及10min的缺氧期间分别应用100μmol/L的MK801。所有脑片均给予60min的复氧。应用细胞内记录技术记录海马CA1区神经元缓慢去极化及快速去极化的时间、快速去极化的幅度。在复氧末,应用细胞内注入电流及经Schaffer通路刺激,观察神经元对刺激的反应。主要观察指标:①两组海马CA1区锥体神经元缓慢去极化速率。②两组海马CA1区神经元的快速去极化时间。③两组海马CA1区神经元的快速去极化幅度。④MK801对海马CA1区神经元功能恢复的影响。结果:①海马CA1区锥体神经元缓慢去极化速率:单纯缺氧组显著高于MK801组[(0.20±0.05)mV/s,(0.08±0.03)mV/s,(P<0.05)]。②海马CA1区神经元的快速去极化时间:MK801组显著高于单纯缺氧组[(537±139)s,(261±26)s,(P<0.05)]。③海马CA1区神经元的快速去极化的幅度:MK801组显著小于单纯缺氧组[(4±13)mV,(53±7)mV,(P<0.05)。④在复氧末期,MK801组的10个神经元中,9个神经元恢复对刺激的反应。结论:N-甲基-D-天冬氨酸受体阻断剂MK801可以显著降低缺氧引起的神经元缓慢去极化速率,延缓神经元快速去极化的发生及降低快速去极化的幅度,说明N-甲基-D-天冬氨酸受体阻断剂可显著减轻神经元的缺氧性损伤,促进复氧后神经元功能的恢复。
BACKGROUND: Pyramidal cells in the hippocampal CA1 region are the most sensitive neurons to hypoxic-ischemic injury. After hypoxia and hypoxia, the membrane potential of hippocampal CA1 pyramidal cells is hyperpolarized in early hypoxia. Prolonged cell membrane slow depolarization and rapid depolarization, causing irreversible neuronal damage. OBJECTIVE: To observe the changes of electrophysiological indexes of hippocampal CA1 pyramidal cells during hypoxia by using intracellular recording technique to observe the effects of N-methyl-D-aspartate receptor blocker MK801 on hippocampal slices. Design: observe the contrast experiment. Unit: 97th People’s Liberation Army Hospital, Jiangsu Province Key Laboratory of Anesthesiology, Xuzhou Medical College, Healthy-ScienceCenter, StateUniversityofNewYork. MATERIALS: Experiments were performed at the New York State University Medical Center from September 2002 to February 2003. Five adult male Sprague-Dawley rats were pretreated with pure oxygen for 3 minutes and then anesthetized with isoflurane at a volume fraction of 0.02. The brain was rapidly decapitated to prepare isolated hippocampal slices. Methods: Rat hippocampal slices were randomly divided into simple hypoxia group and MK801 group, 10 in each group. The hippocampal slices of hypoxia group were given hypoxia for 10 minutes while the hippocampal slices of MK801 group were treated with 100 μmol / L MK801 for 10 minutes before hypoxia and 10 minutes hypoxia respectively. All brain slices are given reoxygenation 60min. Intracellular recording technique was used to record the slow depolarization and rapid depolarization time of neurons in hippocampal CA1 region and the amplitude of rapid depolarization. At the end of reoxygenation, cells were injected with current and stimulated with Schaffer’s pathway to observe the response of neurons to the stimulus. MAIN OUTCOME MEASURES: ① Slow depolarization rate of pyramidal neurons in CA1 area of hippocampus in both groups. ② rapid depolarization time of hippocampal CA1 neurons in both groups. ③ rapid depolarization of neurons in CA1 area of hippocampus in both groups. Effect of MK801 on functional recovery of neurons in CA1 area of hippocampus. Results: ① The slow depolarization rate of pyramidal neurons in CA1 hippocampus was significantly higher than that in MK801 group [(0.20 ± 0.05) mV / s, (0.08 ± 0.03) mV / s, P <0.05] . ② The rapid depolarization time of hippocampal CA1 neurons in MK801 group was significantly higher than that in hypoxia group [(537 ± 139) s, (261 ± 26) s, P <0.05]. (3) The amplitude of rapid depolarization in hippocampal CA1 neurons was significantly lower in MK801 group than in hypoxia group [(4 ± 13) mV, (53 ± 7) mV, P <0.05). ④ At the end of reoxygenation, of the 10 neurons in the MK801 group, 9 neurons recovered their response to stimulation. Conclusion: N-methyl-D-aspartate receptor blocker MK801 can significantly reduce the slow depolarization rate of neurons caused by hypoxia, delay the depolarization of neurons and reduce the depolarization Amplitude, indicating that N-methyl-D-aspartate receptor blockers can significantly reduce neuronal hypoxia damage and promote neuronal recovery after reoxygenation.