Xu W, Willis J S
Department of Physiology, University of Illinois, Urbana 61801.
J Membr Biol. 1994 Sep;141(3):277-87. doi: 10.1007/BF00235137.
Previous work showed that in hamster red cells the amiloride-sensitive (AS) Na+ influx of 0.8 mmol/liter cells/hr is not mediated by Na-H exchange as in other red cells, but depends upon intracellular Mg2+ and can be increased by 40-fold by loading cells with Mg2+ to 10 mM. The purpose of this study was to verify the connection of AS Na+ influx with Na-dependent, amiloride-sensitive Mg2+ efflux and to utilize AS Na+ influx to explore that pathway. Determination of unidirectional influx of Na+ and net loss of Mg2+ in parallel sets of cells showed that activation by extracellular [Na+] follows a simple Michaelis-Menten relationship for both processes with a Km of 105-107 mM and that activation of both processes is sigmoidally dependent upon cytoplasmic [Mg2+] with a [Mg2+]0.5 of 2.1-2.3 mM and a Hill coefficient of 1.8. Comparison of Vmax for both sets of experiments indicated a stoichiometry of 2 Na:1 Mg. Amiloride inhibits Na+ influx and Mg2+ extrusion in parallel (Ki = 0.3 mM). Like Mg2+ extrusion, amiloride-sensitive Na+ influx shows an absolute requirement for cytoplasmic ATP and is increased by cell swelling. Hence, amiloride-sensitive Na+ influx in hamster red cells appears to be through the Na-Mg exchange pathway. There was no amiloride-sensitive Na+ efflux in hamster red cells loaded with Na+ and incubated with high [Mg2+] in the medium with or without external Na+, nor with ATP depletion. Hence, this is not a simple Na-Mg exchange carrier.
先前的研究表明，仓鼠红细胞中对氨氯吡咪敏感（AS）的钠离子内流速度为0.8 mmol/升细胞/小时，与其他红细胞不同，它不是由钠-氢交换介导的，而是依赖于细胞内镁离子，并且通过将细胞内镁离子加载到10 mM可使其增加40倍。本研究的目的是验证AS钠离子内流与钠依赖性、氨氯吡咪敏感的镁离子外流之间的联系，并利用AS钠离子内流来探索该途径。对平行细胞组中钠离子的单向内流和镁离子的净损失进行测定，结果表明，细胞外[Na⁺]对这两个过程的激活均遵循简单的米氏关系，Km为105 - 107 mM，并且这两个过程的激活均呈S形依赖于细胞质[Mg²⁺]，[Mg²⁺]0.5为2.1 - 2.3 mM，希尔系数为1.8。两组实验的Vmax比较表明化学计量比为2 Na:1 Mg。氨氯吡咪同时抑制钠离子内流和镁离子外排（Ki = 0.3 mM）。与镁离子外排一样，氨氯吡咪敏感的钠离子内流对细胞质ATP有绝对需求，并且会因细胞肿胀而增加。因此，仓鼠红细胞中氨氯吡咪敏感的钠离子内流似乎是通过钠-镁交换途径进行的。在用高[Mg²⁺]培养基加载钠离子并孵育的仓鼠红细胞中，无论有无外部钠离子，也无论ATP是否耗尽，均未发现氨氯吡咪敏感的钠离子外流。因此，这不是一个简单的钠-镁交换载体。
