Hays S R, Alpern R J
Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235.
Am J Physiol. 1990 Oct;259(4 Pt 2):F628-35. doi: 10.1152/ajprenal.1990.259.4.F628.
To examine mechanisms of H+ extrusion in the inner stripe of outer medullary collecting duct (OMCDIS), cell pH (pHi) was measured microfluorometrically in in vitro perfused tubules by use of 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. In total absence of luminal and peritubular Na+, pHi recovery from an acid load (NH3/NH+4 pulse) occurred at an initial rate of 0.13 +/- 0.02 pH units/min, whereas in the presence of 135 mM peritubular Na+, pHi recovered at 1.40 +/- 0.28 pH units/min. Na(+)-dependent pHi recovery was completely inhibited by 1.0 mM peritubular amiloride. Luminal Na+ (135 mM) addition had no effect on pHi recovery. Na(+)-independent pHi recovery from acid load was manifest by a triphasic response: 1) initial slow alkalinization; 2) slow cell acidification; and 3) a final phase that exhibited gradually increasing rates of alkalinization, returning pHi above the initial control level (pre-NH3/NH+4 pulse). Luminal N-ethylmaleimide (NEM, 500 microM), an H(+)-ATPase inhibitor, significantly inhibited initial rate of pHi recovery and total pHi recovery; whereas 500 microM peritubular NEM had no effect on initial rate of pHi recovery. Luminal SCH 28080 (100 microM), an H(+)-K(+)-ATPase inhibitor, had no effect on initial rate of pHi recovery or total pHi recovery. Thus rabbit OMCDIS possesses both an apical membrane NEM-sensitive, SCH 28080-insensitive, Na(+)-independent H+ extrusion mechanism (likely a simple H(+)-translocating ATPase) and a basolateral membrane amiloride-sensitive Na(+)-H+ antiporter.
为研究外髓集合管内带（OMCDIS）中H⁺排出的机制，利用2',7'-双（羧乙基）-5（6）-羧基荧光素，通过微荧光法在体外灌注小管中测量细胞pH（pHi）。在完全没有管腔和肾小管周围Na⁺的情况下，酸负荷（NH₃/NH₄⁺脉冲）后pHi恢复的初始速率为0.13±0.02pH单位/分钟，而在存在135mM肾小管周围Na⁺的情况下，pHi以1.40±0.28pH单位/分钟的速率恢复。1.0mM肾小管周围氨氯吡咪完全抑制了依赖Na⁺的pHi恢复。添加管腔Na⁺（135mM）对pHi恢复没有影响。酸负荷后不依赖Na⁺的pHi恢复表现为三相反应：1）初始缓慢碱化；2）缓慢的细胞酸化；3）最后阶段表现出逐渐增加的碱化速率，使pHi恢复到初始对照水平（NH₃/NH₄⁺脉冲前）以上。管腔N-乙基马来酰亚胺（NEM，500μM），一种H⁺-ATP酶抑制剂，显著抑制pHi恢复的初始速率和总pHi恢复；而500μM肾小管周围NEM对pHi恢复的初始速率没有影响。管腔SCH 28080（100μM），一种H⁺-K⁺-ATP酶抑制剂，对pHi恢复的初始速率或总pHi恢复没有影响。因此，兔OMCDIS具有一种顶端膜对NEM敏感、对SCH 28080不敏感、不依赖Na⁺的H⁺排出机制（可能是一种简单的H⁺转运ATP酶）和一种基底外侧膜对氨氯吡咪敏感的Na⁺-H⁺反向转运体。
