Grinstein S, Rothstein A, Cohen S
J Gen Physiol. 1985 May;85(5):765-87. doi: 10.1085/jgp.85.5.765.
The activity of the Na+/H+ exchange system of rat thymic lymphocytes was determined by means of intracellular (pHi) and extracellular pH (pH0) measurements. In isotonic media, the antiport is virtually quiescent at physiological pHi (7.0-7.1), but is greatly activated by cytoplasmic acidification. At normal pHi, the antiport can also be activated by osmotic shrinking. Osmotic activation occurs after a delay of 20-30 s and is reversed several minutes after iso-osmolarity is restored. The mechanism of activation was analyzed by comparing the kinetic parameters of transport in resting (isotonic) and hyperosmotically stressed cells. The affinities of the external substrate site for Na+ and H+ are not altered in shrunken cells. In contrast, the Hi+ sensitivity of the antiport (which is largely dictated by an allosteric modifier site) was increased, which accounted for the activation. The concentration of free cytoplasmic Ca2+ [( Ca2+]i) increased after osmotic shrinking. This increase was dependent on the presence of extracellular Ca2+ and Na+ and was blocked by inhibitors of Na+/H+ exchange, which suggests that it is a consequence, rather than the cause, of the activation of the antiport. It is concluded that the shift in the pHi dependence of the modifier site of the Na+/H+ antiport is the primary event underlying the regulatory volume increase that follows osmotic shrinkage.
通过测量细胞内（pHi）和细胞外pH（pH0）来测定大鼠胸腺淋巴细胞的Na+/H+交换系统的活性。在等渗介质中，该反向转运体在生理pHi（7.0 - 7.1）时实际上处于静止状态，但会因细胞质酸化而被极大激活。在正常pHi时，该反向转运体也可被渗透性收缩激活。渗透性激活在延迟20 - 30秒后发生，并在恢复等渗状态几分钟后逆转。通过比较静息（等渗）细胞和高渗应激细胞中转运的动力学参数来分析激活机制。在收缩细胞中，外部底物位点对Na+和H+的亲和力没有改变。相反，反向转运体对H+的敏感性（很大程度上由变构调节位点决定）增加，这解释了激活现象。渗透性收缩后，游离细胞质Ca2+浓度[（Ca2+]i）增加。这种增加依赖于细胞外Ca2+和Na+的存在，并被Na+/H+交换抑制剂阻断，这表明它是反向转运体激活的结果而非原因。得出的结论是，Na+/H+反向转运体调节位点的pHi依赖性变化是渗透性收缩后调节性容积增加的主要潜在事件。
