Reed D J, Savage M K
Department of Biochemistry and Biophysics, Oregon State University, Corvallis 97331-7305, USA.
Biochim Biophys Acta. 1995 May 24;1271(1):43-50. doi: 10.1016/0925-4439(95)00008-r.
Treatment of isolated mitochondria with Ca2+ and inorganic phosphate (Pi) induces an inner membrane permeability that appears to be mediated through a cyclosporin A (CsA)-inhibitable Ca(2+)-dependent pore. Isolated mitochondria during inner membrane permeability undergo rapid efflux of matrix solutes such as glutathione as GSH and Ca2+, loss of coupled functions, and large amplitude swelling. Permeability transition without large amplitude swelling, a parameter often used to assess inner membrane permeability, has been observed. The addition of either oligomycin, antimycin, or sulfide to incubation buffer containing Ca2+ and Pi abolished large amplitude swelling of mitochondria. The GSH status during a Ca(2+)- and Pi-dependent mechanism of mitochondrial GSH release in isolated mitochondria was influenced significantly by metabolic inhibitors of the respiratory chain but did not prevent inner membrane permeability as demonstrated by the release of mitochondrial GSH and Ca2+. The release of GSH was inhibited by the addition of CsA, a potent inhibitor of permeability transition. Under these conditions we did not find GSSG; however, rapid oxidation of pyridine nucleotides and depletion of ATP and ADP with conversion to AMP occurred. The addition of CsA, prevented the oxidation of pyridine nucleotides and depletion of ATP and ADP. Since NADH and NADPH were extensively oxidized, protection against oxidative stress is reflected in maintenance of GSH and not observable lipid peroxidation. Evidence from transmission electron microscopy analysis, combined with the GSH release data, indicate that permeability transition can be observed in the absence of large amplitude swelling.
用Ca2+和无机磷酸盐（Pi）处理分离的线粒体可诱导内膜通透性增加，这似乎是通过环孢菌素A（CsA）抑制的Ca(2+)依赖性孔介导的。在内膜通透性增加过程中，分离的线粒体基质溶质如谷胱甘肽（GSH）和Ca2+会快速外流，偶联功能丧失，并出现大幅度肿胀。已经观察到没有大幅度肿胀的通透性转变，这是一个常用于评估内膜通透性的参数。向含有Ca2+和Pi的孵育缓冲液中添加寡霉素、抗霉素或硫化物可消除线粒体的大幅度肿胀。在分离的线粒体中，Ca(2+)和Pi依赖性线粒体GSH释放机制过程中的GSH状态受到呼吸链代谢抑制剂的显著影响，但如线粒体GSH和Ca2+的释放所示，并未阻止内膜通透性增加。添加CsA可抑制GSH的释放，CsA是通透性转变的有效抑制剂。在这些条件下，我们未发现氧化型谷胱甘肽（GSSG）；然而，吡啶核苷酸快速氧化，ATP和ADP耗竭并转化为AMP。添加CsA可防止吡啶核苷酸氧化以及ATP和ADP耗竭。由于NADH和NADPH被广泛氧化，对氧化应激的保护作用体现在GSH的维持上，而未观察到脂质过氧化。透射电子显微镜分析的证据与GSH释放数据相结合，表明在没有大幅度肿胀的情况下也可观察到通透性转变。
