Brustovetsky Nickolay, Brustovetsky Tatiana, Jemmerson Ronald, Dubinsky Janet M
Department of Neuroscience, University of Minnesota, Minneapolis 55455, USA.
J Neurochem. 2002 Jan;80(2):207-18. doi: 10.1046/j.0022-3042.2001.00671.x.
The mechanisms of Ca2+-induced release of Cytochrome c (Cyt c) from rat brain mitochondria were examined quantitatively using a capture ELISA. In 75 or 125 mm KCl-based media 1.4 micromol Ca2+/mg protein caused depolarization and mitochondrial swelling. However, this resulted in partial Cyt c release only in 75 mm KCl. The release was inhibited by Ru360, an inhibitor of the Ca2+ uniporter, and by cyclosporin A plus ADP, a combination of mitochondrial permeability transition inhibitors. Transmission electron microscopy (TEM) revealed that Ca2+-induced swelling caused rupture of the outer membrane only in 75 mm KCl. Koenig's polyanion, an inhibitor of mitochondrial porin (VDAC), enhanced swelling and amplified Cyt c release. Dextran T70 that is known to enhance mitochondrial contact site formation did not prevent Cyt c release. Exposure of cultured cortical neurons to 500 microM glutamate for 5 min caused Cyt c release into the cytosol 30 min after glutamate removal. MK-801 or CsA inhibited this release. Thus, the release of Cyt c from CNS mitochondria induced by Ca2+ in vitro as well as in situ involved the mPT and appeared to require the rupture of the outer membrane.
利用捕获酶联免疫吸附测定法（ELISA）对钙离子诱导大鼠脑线粒体释放细胞色素c（Cyt c）的机制进行了定量研究。在以75或125 mM氯化钾为基础的培养基中，1.4 μmol Ca²⁺/mg蛋白质会导致去极化和线粒体肿胀。然而，这仅在75 mM氯化钾中导致部分Cyt c释放。钙离子单向转运体抑制剂钌红（Ru360）以及线粒体通透性转换抑制剂环孢素A加二磷酸腺苷（ADP）组合可抑制这种释放。透射电子显微镜（TEM）显示，钙离子诱导的肿胀仅在75 mM氯化钾中导致外膜破裂。线粒体孔蛋白（电压依赖性阴离子通道，VDAC）抑制剂柯尼格多聚阴离子可增强肿胀并放大Cyt c释放。已知可增强线粒体接触位点形成的葡聚糖T70并不能阻止Cyt c释放。将培养的皮层神经元暴露于500 μM谷氨酸5分钟，在去除谷氨酸30分钟后会导致Cyt c释放到细胞质中。MK-801或环孢素A可抑制这种释放。因此，体外以及原位由钙离子诱导的中枢神经系统线粒体释放Cyt c涉及线粒体通透性转换（mPT），并且似乎需要外膜破裂。
