Mitochondrial membrane potential and hydroethidine-monitored superoxide generation in cultured cerebellar granule cells
Mitochondrial membrane potential and hydroethidine-monitored superoxide generation in cultured cerebellar granule cells
复制标题
DOI：
10.1016/s0014-5793(97)01088-0
复制
发表时间：
1997-09-22
期刊：
影响因子：
3.5
通讯作者：
Nicholls, DG
中科院分区：
文献类型：
--
作者：
Budd, SL;Castilho, RF;Nicholls, DG
据报道，线粒体去极化可增强超氧阴离子（O-2（.-））的产生。在许多细胞制备物中，而在分离的线粒体中观察到抑制。小脑颗粒细胞用> 1 μ M氢乙啶（二氢乙啶）平衡，氢乙啶被O-2（.-）氧化成荧光乙啶阳离子。然而，对照显示荧光增强是未结合的预先形成的乙锭从细胞内的线粒体基质释放的结果，产生荧光增强。通过使用低浓度（1 μ M）染料消除了这种模糊性，在这种情况下，产生的乙锭保持结合在线粒体内。在这些条件下，抗霉素A，而不是质子载体的加入，产生了荧光的增加，它的结论是，过量的乙锭作为线粒体膜电位的指示剂模糊了监测O-2（.-）并且在细胞中使用该指示剂的某些实验可能需要重新评估，（C）1997年欧洲生物化学学会联合会。
Mitochondrial depolarisation has been reported to enhance the generation of superoxide anion (O-2(.-)) in a number of cell preparations while an inhibition has been observed with isolated mitochondria. Cerebellar granule cells equilibrated with > 1 mu M hydroethidine (dihydroethidium) which is oxidised to the fluorescent ethidium cation by O-2(.-) showed a large increase in fluorescence on protonophore addition, However, controls showed the fluorescent enhancement to be a consequence of release of unbound preformed ethidium from the mitochondrial matrix within the cell with resultant fluorescent enhancement, This ambiguity was removed by the use of low (1 mu M) concentrations of dye in which case generated ethidium remained bound within the mitochondria, Under these conditions antimycin A, but not protonophore addition, produced an increase in fluorescence, It is concluded that excess ethidium acts as an indicator of mitochondrial membrane potential obscuring the monitoring of O-2(.-) and that certain experiments employing this indicator in cells may require re-evaluation, (C) 1997 Federation of European Biochemical Societies.