Landry Y, Lehninger A L
Biochem J. 1976 Aug 15;158(2):427-38. doi: 10.1042/bj1580427.
Ehrlich ascites-tumour cells accumulate Ca2+ when incubated aerobically with succinate, phosphate and rotenone, as revealed by isotopic and atomic-absorption measurements. Ca2+ does not stimulate oxygen consumption by carefully prepared Ehrlich cells, but des so when the cells are placed in a hypo-osmotic medium. Neither glutamate nor malate support Ca2+ uptake in 'intact' Ehrlich cells, nor does the endogenous NAD-linked respiration. Ca2+ uptake is completely dependent on mitochondrial energy-coupling mechansims. It was an unexpected finding that maximal Ca2+ uptake supported by succinate requires rotenone, which blocks oxidation of enogenous NAD-linked substrates. Phosphate functions as co-anion for entry of Ca2+. Ca2+ uptake is also supported by extra-cellular ATP; no other nucleoside 5'-di- or tri-phosphate was active. The accumulation of Ca2+ apparently takes place in the mitochondria, since oligomycin and atractyloside inhibit ATP-supported Ca2+ uptake. Glycolysis does not support Ca2+ uptake. Neither free mitochondria released from disrupted cells nor permeability-damaged cells capable of absorbing Trypan Blue were responsible for any large fraction of the total observed energy-coupled Ca2+ uptake. The observations reported also indicate that electron flow through energy-conserving site 1 promotes Ca2+ release from Ehrlich cells and that extra-cellular ATP increase permeability of the cell membrane, allowing both ATP and Ca2+ to enter the cells more readily.
同位素和原子吸收测量结果显示，艾氏腹水癌细胞在与琥珀酸盐、磷酸盐和鱼藤酮进行需氧孵育时会积累钙离子。精心制备的艾氏细胞中，钙离子不会刺激氧气消耗，但当细胞置于低渗培养基中时则会刺激氧气消耗。无论是谷氨酸还是苹果酸都不能支持“完整”的艾氏细胞摄取钙离子，内源性烟酰胺腺嘌呤二核苷酸（NAD）相关的呼吸作用也不能支持。钙离子摄取完全依赖于线粒体能量偶联机制。一个意外的发现是，琥珀酸盐支持的最大钙离子摄取需要鱼藤酮，而鱼藤酮会阻断内源性NAD相关底物的氧化。磷酸盐作为钙离子进入细胞的共阴离子发挥作用。细胞外ATP也能支持钙离子摄取；其他核苷5'-二磷酸或三磷酸均无活性。钙离子的积累显然发生在线粒体中，因为寡霉素和苍术苷会抑制ATP支持的钙离子摄取。糖酵解不支持钙离子摄取。无论是破碎细胞释放的游离线粒体，还是能够吸收台盼蓝的通透性受损细胞，都不能对观察到的总能量偶联钙离子摄取量的很大一部分负责。所报告的观察结果还表明，通过能量保存位点1的电子流会促进钙离子从艾氏细胞中释放，并且细胞外ATP会增加细胞膜的通透性，使ATP和钙离子都更容易进入细胞。
