Action of ionophore A23187 at the cellular level. Separation of effects at the plasma and mitochondrial membranes.
Action of ionophore A23187 at the cellular level. Separation of effects at the plasma and mitochondrial membranes.
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离子载体 A23187 在细胞水平的作用。
DOI：
10.1016/s0021-9258(17)33330-6
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发表时间：
1976
期刊：
影响因子：
--
通讯作者：
H. Lardy
中科院分区：
文献类型：
--
作者：
D. Babcock;N. First;H. Lardy
牛附睾精子在0.1 - 0.2 mM CaCl 2存在下体外有氧孵育，可累积25 - 50 nmol钙/10（8）个细胞。添加低浓度的离子载体A23187（0.01至0.5 nmol/mg精子蛋白）可诱导这种积累的钙外流。在高离子载体浓度（0.5至5.0 nmol/mg精子蛋白）下，钙释放后，不依赖于线粒体膜电位的钙流入量高达25 nmol/10（8）个细胞。用多烯抗生素filipin处理产生的精子质膜渗透性的选择性增加导致在高浓度A23187存在下积累的钙的释放。首先用菲律宾处理的精子具有积累和保留钙的能力（在可氧化底物的存在下），但在添加3 nmol A23187/mg蛋白质后释放Ca 2+，随后没有再积累。这些观察结果解释了竞争性钙泵的存在下，线粒体和质膜的精子内运作。用高浓度A23187处理允许钙流入精子细胞的非线粒体区室，这是该阳离子在线粒体和质膜上平衡的结果。钙摄取量及其对菲律宾肽的敏感性表明钙与可溶性细胞内组分的结合也参与其中。低浓度的A23187诱导钙外流的能力被解释为质膜泵的持续操作与离子载体诱导的线粒体膜上的浓度梯度衰减一致的结果。低水平的线粒体上的离子载体的这种假设的行动是由菲律宾处理的细胞和完整的精子的磷酸盐和膜相关的钙的呼吸反应和运动的钙的净运动的观察支持。有人建议，这种明显的选择性的离子载体作用的基础在于在这些细胞的质膜和线粒体膜中的竞争钙泵的相对活动和动力学性质。离子载体诱导的钙离子流入线粒体外空间导致精子呼吸和动力学活动的刺激。这种运动性的激活也被观察到与细胞完全依赖于糖酵解（通过与呼吸抑制剂的治疗），并表明在鞭毛功能的调节钙的直接参与。
Bovine epididymal spermatozoa incubated aerobically in vitro in the presence of 0.1 to 0.2 mM CaCl2 accumulate 25 to 50 nmol of calcium/10(8) cells. The addition of low concentrations of the ionophore A23187 (0.01 to 0.5 nmol/mg of sperm protein) induces efflux of this accumulated calcium. At high ionophore concentrations (0.5 to 5.0 nmol/mg of sperm protein), calcium release is followed by an influx of up to 25 nmol of calcium/10(8) cells that is not dependent on mitochondrial energization. A selective increase in the permeability of the sperm plasma membrane produced by treatment with the polyene antibiotic, filipin, results in the release of that calcium which is accumulated in the presence of high concentrations of A23187. Sperm first treated with filipin possess the ability to accumulate and retain calcium (in the presence of an oxidizable substrate) but release Ca2+ without subsequent reaccumulation after the addition of 3 nmol of A23187/mg of protein. These observations are explained by the existence of competing calcium pumps operating within the mitochondrial and plasma membranes of the spermatozoan. Treatment with high concentrations of A23187 allows calcium influx into a non-mitochondrial compartment of the sperm cell as a consequence of the equilibration of this cation across both mitochondrial and plasma membranes. The amount of calcium uptake and its sensitivity to filipin indicate that calcium binding to soluble, intracellular components is also involved. The ability of low concentrations of A23187 to induce calcium efflux is explained as a result of the continued operation of the plasma membrane pump coincident with ionophore-induced decay of the concentration gradient across the mitochondrial membrane. This hypothetical action of low levels of the ionophore on the mitochondria is supported by the observation of net movements of calcium with filipin-treated cells and the respiratory responses and movements of phosphate and membrane-associated calcium with intact sperm. It is suggested that the basis of this apparent selectivity of ionophore action lies in the relative activities and kinetic properties of the competing calcium pumps in the plasma and mitochondrial membranes of these cells. Ionophore-induced influx of calcium into the extramitochondrial space results in a stimulation of respiration and kinetic activity of the sperm. This activation of motility is observed also with cells made entirely dependent upon glycolysis (by treatment with respiratory inhibitors) and suggests a direct involvement of calcium in the regulation of flagellar function.