Grinstein S, Goetz J D, Rothstein A
J Gen Physiol. 1984 Oct;84(4):585-600. doi: 10.1085/jgp.84.4.585.
22Na+ flux and cytoplasmic pH (pHi) determinations were used to study the reversibility, symmetry, and mechanism of activation of the Na+/H+ exchange system in rat thymic lymphocytes. In acid-loaded cells, the antiport can be detected as an Na+-induced, amiloride-sensitive alkalinization. At pHi greater than or equal to 7.0, amiloride-sensitive net H+ fluxes are not detectable. To investigate whether at this pHi the transporter is operative in a different mode, e.g., Na+/Na+ exchange, 22Na+ uptake was measured as a function of pHi. The results indicate that the antiport is relatively inactive at pHi greater than or equal to 7.0. Comparison of the rates of H+ efflux (or equivalent OH- uptake) and Na+ uptake indicate that Na+/Na+ countertransport through this system is negligible at all values of pHi and that the Na+:H+ stoichiometry is 1:1. Measurements of pHi in Na+-loaded cells suspended in Na+-free medium revealed an amiloride-sensitive cytoplasmic acidification, which is indicative of exchange of internal Na+ for external H+. The symmetry of the system was analyzed by measuring the effect of extracellular pH (pHo) on Na+ efflux. Unlike cytoplasmic acidification, lowering pHo failed to activate the antiport. The results indicate that the amiloride-sensitive Na+/H+ exchanger is reversible but asymmetric. The system is virtually inactive at pHi greater than or equal to 7.0 but can be activated by protonation of a modifier site on the cytoplasmic surface. Activation can also occur by depletion of cellular Na+. It is proposed that Na+ may also interact with the modifier site, stabilizing the unprotonated (inactive) form.
利用²²Na⁺通量和细胞质pH值（pHi）测定法，研究大鼠胸腺淋巴细胞中Na⁺/H⁺交换系统激活的可逆性、对称性及机制。在酸负荷细胞中，该反向转运体可被检测为Na⁺诱导的、氨氯地平敏感的碱化。当pHi大于或等于7.0时，无法检测到氨氯地平敏感的净H⁺通量。为研究在此pHi时转运体是否以不同模式运作，如Na⁺/Na⁺交换，测量了作为pHi函数的²²Na⁺摄取量。结果表明，在pHi大于或等于7.0时，反向转运体相对不活跃。H⁺外流速率（或等效OH⁻摄取速率）与Na⁺摄取速率的比较表明，在所有pHi值下，通过该系统的Na⁺/Na⁺逆向转运可忽略不计，且Na⁺:H⁺化学计量比为1:1。对悬浮于无Na⁺培养基中的Na⁺负荷细胞进行pHi测量，发现有氨氯地平敏感的细胞质酸化，这表明内部Na⁺与外部H⁺发生了交换。通过测量细胞外pH值（pHo）对Na⁺外流的影响来分析该系统的对称性。与细胞质酸化不同，降低pHo未能激活反向转运体。结果表明，氨氯地平敏感的Na⁺/H⁺交换体是可逆的但不对称。该系统在pHi大于或等于7.0时几乎不活跃，但可通过细胞质表面修饰位点的质子化激活。细胞内Na⁺耗竭也可引发激活。有人提出，Na⁺可能也与修饰位点相互作用，稳定未质子化（无活性）形式。
