Role of Mitochondrial Permeability Transition in Ischemic Neuronal Cell Death

本研究旨在研究脑线粒体线粒体通透性转变（mPT）的生化特征，并阐明其在短暂性前脑缺血后神经元损伤中的作用。从大鼠脑和肝脏中分离出线粒体。通过540 nm光吸收测定线粒体体积的变化。通过Western blot分析，我们检测了这些条件下线粒体细胞色素c的释放。沙鼠双侧颈总动脉闭塞5min，引起短暂性前脑缺血。缺血前30分钟和缺血后24小时给予环孢素A （CsA）和/或三氟拉嗪。缺血7天后，计算海马CA1区存活神经元数量。钙（而非促氧化剂）引起脑线粒体光吸收下降，CsA可抑制这一变化。然而，这两种刺激导致肝脏线粒体的光吸收下降幅度更大。在这些条件下，钙诱导脑线粒体释放细胞色素c，而CsA不抑制这种释放。然而，同样的刺激导致肝脏线粒体释放更多的细胞色素c。高剂量CsA治疗可显著改善短暂性前脑缺血所致的选择性神经元损伤。三氟拉嗪的保护作用增强。这些结果表明，mPT不容易在脑线粒体中发生，细胞色素c通过不依赖于mPT的机制从脑线粒体中释放出来。然而，前脑短暂缺血的结果表明，脑内mPT和/或细胞色素c释放的敏感性可能存在区域特异性差异。

The present study was aimed to determine biochemical characteristics of mitochondrial, permeability transition (mPT) in brain mitochondria, and to clarify its role in neuronal injury following transient forebrain ischemia. Mitochondria were isolated from rat brain and liver. Changes in mitochondrial volume were measured via light absorbance at 540 nm. By Western blot analysis, we examined release of mitochondrial cytochrome c under these conditions. Transient forebrain ischemia was induced by 5-min occlusion of the bilateral common carotid artery in the gerbil. Cyclosporin A (CsA) and/or trifluoperazine were given 30 min before and 24 hour after ischemia. The number of surviving neurons in hippocampal CA1 sectors was counted 7 days after ischemia. Calcium, but not pro-oxidants, induced a decrease of light absorbance in brain mitochondria, which was inhibited by CsA. However, both stimuli led to a much larger decrease of light absorbance in liver mitochondria. Under these conditions, calcium induced a release of cytochrome c from brain mitochondria, which was not inhibited by CsA. However, the same stimulus led to a release of larger amount of cytochrome c from liver mitochondoria. Selective neuronal injury due to transient forebrain ischemia was significantly ameliorated by treatment with high-dose CsA. The protective effect was enhanced by trifluoperazine. These results suggest that mPT does not readily occur in brain mitochondria, and that cytochrome c is released from brain mitochondria through mPT-independent mechanism. However, the findings from transient forebrain ischemia indicate the possibility of region-specific difference in sensitivity for mPT and/or cytochrome c release in the brain.