Brain protection from the neuronal death induced by spreading depression

大脑免受抑郁症蔓延引起的神经元死亡的保护

• 批准号: 13671612
• 负责人: IIJIMA Takehiko
• 金额: $ 0.45万
• 依托单位: Kyorin University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13671612/
• 关键词: Cerebral ischemia; Mitochondria; Menbrane potential; Glutathione; Reactive nitrogen species; reperfusion injury; spreading depression; microdialysis; 虚血

We have accomplished two projects supported by this grant. First, we examined radical formation and radical scavenging during reperfusion period after ischemia. Second, we examined mitochondrial membrane potential during reoxygenation after oxygen-glucose deprivation. The first project suggested physiological protective mechanism of neuron from toxic effect of free radical during early reperfusion phase. Therefore, we moved to mitochondrial mechanism involved in active neuronal suicide, apoptosis because active neuronal death executed by neuron itself seems to be a main cause of neuronal damage after ischemia.1. The role of glutathione during reperfusionA timed profile of glutathione oxidation and reactive nitrogen species during reperfusion after cerebral ischemia in rat was obtained. GSH and GSSG increased and reached a peak; 3408 ^^+__-1710% (mean ^^+__- SE) at 25 min of reperfusion (P<0.0001); and 329 ^^+__- 104% at 50 min of reperfusion (p=0.06), respectively. NO_2 levels did not significantly change. These data suggest that GSH releases during early phase of reperfusion and its rapid oxidation contributes to prevent increase in reactive nitrogen species.2. Involvement of mitochondrial mechanism for ischemic neuronal deathA primary hippocampal culture seeded in a 35mm fenestrated dish for fluorescence microscopy was mounted in a sealed chamber for an anaerobic incubation (oxygen-glucose deprivation (OGD)). During OGD, MMP decreased to 0.72 ^^+__- 0.03 (normalized JC-1 fluorescence), then increased to the hyperpolarized level 1.99 ^^+__- 0.12 during 60min reoxygenation after 30 min OGD. After 90 min OGD and reoxygenation, MMP was reduced and never recovered. The intracellular ATP content was 8.1 ^^+__- 6.6% and 3.2 ^^+__- 1.9% after 30 mm OGD and 30 min reoxygenation following 30 min OGD, respectively. This observation suggests the inhibition of electron reentry into an inner membrane during reoxygenation and the disturbance of F_0F_1-ATP synthase.

我们已经完成了这笔赠款支持的两个项目。首先，我们检查了缺血后再灌注期间的自由基形成和自由基清除。其次，我们检查了氧糖剥夺后复氧过程中的线粒体膜电位。第一个项目提出了神经元在再灌注早期免受自由基毒性作用的生理保护机制。因此，我们转向参与主动神经元自杀、凋亡的线粒体机制，因为神经元本身执行的主动神经元死亡似乎是缺血后神经元损伤的主要原因。 1．谷胱甘肽在再灌注过程中的作用获得了大鼠脑缺血后再灌注过程中谷胱甘肽氧化和活性氮的定时曲线。 GSH和GSSG升高并达到峰值；再灌注 25 分钟时 3408 ^^+__-1710%（平均 ^^+__- SE）（P<0.0001）；再灌注 50 分钟时分别为 329 ^^+__- 104% (p=0.06)。 NO_2水平没有显着变化。这些数据表明，GSH在再灌注早期释放，其快速氧化有助于防止活性氮的增加。2．缺血性神经元死亡的线粒体机制的参与将原代海马培养物接种在用于荧光显微镜的35mm有窗培养皿中，然后安装在密封室中进行厌氧培养（氧-葡萄糖剥夺（OGD））。 OGD期间，MMP下降至0.72 ^^+__- 0.03（标准化JC-1荧光），然后在30分钟OGD后60分钟复氧期间增加至超极化水平1.99 ^^+__- 0.12。 90 分钟 OGD 和复氧后，MMP 减少且从未恢复。 30 mm OGD 后和 30 分钟 OGD 后 30 分钟复氧后，细胞内 ATP 含量分别为 8.1 ^^+__- 6.6% 和 3.2 ^^+__- 1.9%。这一观察结果表明，在再氧合过程中电子重新进入内膜受到抑制，并且 F_0F_1-ATP 合酶受到干扰。

项目成果

飯島毅彦, 三嶋竜弥, 赤川公朗, 比嘉正祐, 巌康秀: "培養細胞におけるミトコンドリア膜電位の虚血時の変化-プロポフォールの影響-"Pharmacoanesthesiology. 13(2). 112-115 (2001)

Takehiko Iijima、Tatsuya Mishima、Kimiro Akakawa、Masuke Higa、Yasuhide Iwao：“培养细胞缺血期间线粒体膜电位的变化 - 异丙酚的影响 -” Pharmacoanesthesiology 13(2)。

Iijima T., Sakamoto H., Okada C., and Iwao Y.: "Relationship between oxidation of glutathione and reactive nitrogen species during the early-reperfusion phase of cerebral ischemia"Neurochem Res. 27(6). 497-500 (2002)

Iijima T.、Sakamoto H.、Okada C. 和 Iwao Y.：“脑缺血早期再灌注阶段谷胱甘肽氧化与活性氮的关系”Neurochem Res。

飯島 毅彦, 三島竜哉, 赤川公朗, 巌 康秀: "培養神経細胞におけるミトコンドリア膜電位の虚血時の変化"脳循環代謝. 13(3). 156-157 (2001)

Takehiko Iijima、Tatsuya Mishima、Kimiro Akakawa、Yasuhide Iwao：“培养神经元缺血期间线粒体膜电位的变化”《脑循环与代谢》13(3)。

Iijima T, MishimaT, Tohyama M, Akagawa K, Iwao Y: "Mitochondrial membrane potential and intracellular ATP content after transient experimental ischemia in the cultured hippocampal neuron"Neurochemistry International. 43(3). 263-269 (2003)

Iijima T、MishimaT、Tohyama M、Akakawa K、Iwao Y：“培养的海马神经元短暂实验性缺血后的线粒体膜电位和细胞内 ATP 含量”《神经化学国际》。

飯島毅彦, 三嶋竜弥, 赤川公朗, 巌康秀: "培養細胞におけるミトコンドリア膜電位の虚血時の変化"脳循環代謝. 13(3). 156-157 (2001)

Takehiko Iijima、Tatsuya Mishima、Kimiro Akakawa、Yasuhide Iwao：“培养细胞缺血期间线粒体膜电位的变化”《脑循环与代谢》13(3) (2001)。