Haynes R C, Picking R A, Zaks W J
J Biol Chem. 1986 Dec 5;261(34):16121-5.
Rat liver mitochondria were incubated at 30 degrees C with 4 mM ATP in a medium similar in electrolyte composition to that of hepatic cytosol. Under these conditions, a net increase in mitochondrial adenine nucleotides was observed that was dependent on the concentration of free Ca2+ [( Ca2+]) in the incubation medium. At 0.2 microM [Ca2+] or less, there was no demonstrable uptake of adenine nucleotides; at 0.4 microM [Ca2+], or greater, net uptake occurred. The calcium-dependent accumulation of nucleotides by mitochondria required Mg2+ in the incubation medium and was insensitive to carboxyatractyloside. The uptake of adenine nucleotides was enhanced by the addition of antimycin A or antimycin A together with oligomycin. Accumulation of nucleotides appeared to be associated with a small increase in mean mitochondrial volume, but the membrane potential was not affected. No uptake or loss of NAD-NADH by mitochondria was detected. Ruthenium red failed to inhibit the calcium-dependent uptake of adenine nucleotides by the mitochondria, indicating that stimulation of this process by Ca2+ does not involve transport of the cation into mitochondria by the Ca2+ uniporter. Because glucagon acts to elevate cytosolic [Ca2+] from approximately 0.2 microM to 0.6 microM, the same range affecting nucleotide uptake, it is proposed that the increase in mitochondrial adenine nucleotides that follows treatment with glucagon is mediated by the rise in cytosolic [Ca2+] produced by the hormone. This hypothesis was supported by the observation that epinephrine and A23187, agents that raise cytosolic [Ca2+], increased the content of mitochondrial adenine nucleotides in isolated hepatocytes. Furthermore, cells, incubated under calcium-depleting conditions, had a diminished response to glucagon.
大鼠肝脏线粒体在30℃下于一种电解质组成与肝细胞溶质相似的培养基中与4 mM ATP一起孵育。在这些条件下，观察到线粒体腺嘌呤核苷酸有净增加，这取决于孵育培养基中游离Ca2+[(Ca2+)]的浓度。在0.2 microM [Ca2+]或更低时，没有可证明的腺嘌呤核苷酸摄取；在0.4 microM [Ca2+]或更高时，发生净摄取。线粒体对核苷酸的钙依赖性积累需要培养基中有Mg2+，并且对羧基苍术苷不敏感。加入抗霉素A或抗霉素A与寡霉素一起可增强腺嘌呤核苷酸的摄取。核苷酸的积累似乎与线粒体平均体积的小幅增加有关，但膜电位不受影响。未检测到线粒体对NAD - NADH的摄取或损失。钌红未能抑制线粒体对腺嘌呤核苷酸的钙依赖性摄取，表明Ca2+对该过程的刺激不涉及阳离子通过Ca2+单向转运体进入线粒体。因为胰高血糖素的作用是将细胞质[Ca2+]从大约0.2 microM升高到0.6 microM，这也是影响核苷酸摄取的相同范围，所以有人提出，胰高血糖素处理后线粒体腺嘌呤核苷酸的增加是由该激素产生的细胞质[Ca2+]升高介导的。肾上腺素和A23187这两种升高细胞质[Ca2+]的物质增加了分离的肝细胞中线粒体腺嘌呤核苷酸的含量，这一观察结果支持了这一假设。此外，在缺钙条件下孵育的细胞对胰高血糖素的反应减弱。
