Ischemic injury to rat forebrain mitochondria and cellular calcium homeostasis.
Ischemic injury to rat forebrain mitochondria and cellular calcium homeostasis.
复制标题
大鼠前脑线粒体缺血性损伤和细胞钙稳态。
DOI：
10.1016/0167-4889(92)90180-j
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Lee,CP
中科院分区：
文献类型：
--
作者：
Sciamanna,MA;Zinkel,J;Fabi,AY;Lee,CP
Kameyama 等人的三血管闭塞模型。 （Kameyama, M.、Suzuki, J.、Shirane, R. 和 Okawa, A. (1985) Stroke 16, 489–493）进行修改以诱导完全可逆的大鼠前脑缺血。描述了一种快速、简单的大鼠脑线粒体分离和纯化方法，该方法可提供高产量。从缺血性大脑（缺血 12-30 分钟）中分离出的线粒体表现出状态 3 呼吸频率降低约 10%。 70% 与 NAD 连接的呼吸底物有关。琥珀酸盐和鱼藤酮的效果较小。状态 4 呼吸活动仍接近对照水平，除了在 NAD 连接底物缺血 15 分钟时（增加 25%）。同样，对于琥珀酸盐和鱼藤酮，大约为。缺血 20 分钟时观察到状态 4 活性增加 30%。因此，呼吸控制指数（RCIs）下降。在检测混合物中添加 EGTA(EDTA) 或钌红后，呼吸速率和 RCI 都可以恢复到接近控制水平。使用fura-2作为Ca2+探针的分析表明，缺血线粒体的Ca2+摄取的一级速率常数大大降低，而位于线粒体内膜外部的Ca2+显着增加。这些数据表明，缺血会破坏细胞 Ca2+ 稳态，导致胞质 Ca2+ 浓度增加，导致线粒体膜上 Ca2+ 过度结合，并抑制呼吸链连接的氧化磷酸化和前脑线粒体 Ca2+ 转运活性。这些缺陷与缺血的持续时间成正比。
The three-vessel occlusion model of Kameyama et al. (Kameyama, M., Suzuki, J., Shirane, R. and Ogawa, A. (1985) Stroke 16, 489–493) was adapted with modifications to induce complete reversible rat forebrain ischemia. A fast and simple procedure for the isolation and purification of rat brain mitochondria, which provides high yield, is described. Mitochondria isolated from ischemic brain (12–30 min ischemia) exhibited decreases in State 3 respiratory rates of approx. 70% with NAD-linked respiratory substrates. Less effect was observed with succinate and rotenone. The State 4 respiratory activity remained near control levels except at 15 min of ischemia (25% increase) with NAD-linked substrates. Similarly, with succinate and rotenone, an approx. 30% increase in State 4 activity was observed at 20 min of ischemia. Consequently, the respiratory control indices (RCIs) were decreased. Both the respiratory rates and RCIs could be restored to near control levels upon the addition of EGTA(EDTA) or ruthenium red to the assay mixture. Analysis employing fura-2 as a Ca2+probe, indicated a great decrease in the first order rate constant for Ca2+uptake of ischemic mitochondria and a significant increase in Ca2+located externally to the inner mitochondrial membrane. These data suggest that ischemia disrupts cellular Ca2+homeostasis with an increase in the cytosolic Ca2+concentration which results in excessive association of Ca2+on the mitochondrial membrane and an inhibition of the respiratory chain-linked oxidative phosphorylation and Ca2+-transport activity of forebrain mitochondria. These deficits are proportional to the duration of ischemia.