Kahlert S, Schild L, Reiser G
Institute of Neurobiochemistry, Medical Faculty, Otto-von-Guericke-University Magdeburg, Leipziger Str. 44, 39120 Magdeburg, Germany.
J Neurosci Res. 2001 Dec 1;66(5):1019-27. doi: 10.1002/jnr.10052.
The influence of physiological Ca(2+)-inducing stimuli and agents mimicking ischemic conditions on mitochondrial potential was studied in postnatal (P1) hippocampal astrocytes. Cytosolic Ca(2+) loads with characteristic kinetics of rise and duration, detected by Fura-2, were provoked by extracellular Ca(2+) influx, release from InsP(3)-sensitive intracellular stores, or inhibition of the reloading of endoplasmic reticulum Ca(2+) stores. Inhibitors of mitochondrial respiration caused only moderate release of Ca(2+) from intracellular stores, inducing a rise of less than 60 nM. The maximal Ca(2+) rise was found with InsP(3)-mediated responses (500 nM; via ATP) or with ionophore (4-Br-A23187)-mediated Ca(2+) influx from extracellular medium (770 nM). Remarkably, all these agents causing significant rise of cytosolic Ca(2+), only minimally depolarized the mitochondria. Membrane potential of mitochondria was monitored by Rh123 or TMRE. Depolarization was only found with very high cytosolic Ca(2+) levels (above 60 microM; measured by fura FF). These were achieved with external Ca(2+) influx by ionophore in combination with inhibition of glycolysis. Thus, mitochondria in the astrocytes are obviously not sensitive to moderate cytosolic Ca(2+) loads, irrespective of the source of Ca(2+). Furthermore, isolated rat brain mitochondria display a low sensitivity of respiratory activity to Ca(2+), which is consistent with the data obtained with the astrocytes in vitro. The capacity of isolated mitochondria to build up a potential was gradually reduced at low micromolar Ca(2+) and totally compromised only at Ca(2+) concentrations in the 100 microM range.
在出生后（P1）海马星形胶质细胞中研究了生理性钙诱导刺激和模拟缺血条件的试剂对线粒体电位的影响。通过Fura-2检测到的具有特征性上升和持续时间动力学的胞质钙负荷，是由细胞外钙内流、从肌醇三磷酸（InsP3）敏感的细胞内储存释放或内质网钙储存再装载的抑制所引发的。线粒体呼吸抑制剂仅引起细胞内储存中钙的适度释放，导致钙升高小于60 nM。在InsP3介导的反应（500 nM；通过ATP）或离子载体（4-溴-A23187）介导的从细胞外介质的钙内流（770 nM）中发现了最大的钙升高。值得注意的是，所有这些导致胞质钙显著升高的试剂，仅使线粒体轻微去极化。线粒体膜电位通过罗丹明123（Rh123）或四甲基罗丹明乙酯（TMRE）监测。仅在非常高的胞质钙水平（高于60 μM；通过fura FF测量）时才发现去极化。这是通过离子载体介导的外部钙内流并结合糖酵解抑制来实现的。因此，星形胶质细胞中的线粒体显然对适度的胞质钙负荷不敏感，无论钙的来源如何。此外，分离的大鼠脑线粒体对呼吸活性对钙的敏感性较低，这与体外星形胶质细胞获得的数据一致。分离的线粒体建立电位的能力在低微摩尔钙浓度下逐渐降低，仅在100 μM范围内的钙浓度时才完全受损。
