Pearce D, Verkman A S
Department of Medicine, University of California, San Francisco 94143.
Biophys J. 1989 Jun;55(6):1251-9. doi: 10.1016/S0006-3495(89)82920-0.
Two independent methods, induced osmosis and solvent drag, were used to determine the reflection coefficients for NaCl (sigma NaCl) in brush border and basolateral membrane vesicles isolated from rabbit proximal tubule. In the induced osmosis method, vesicles loaded with sucrose were subjected to varying inward NaCl gradients in a stopped-flow apparatus. sigma NaCl was determined from the osmolality of the NaCl solution required to cause no initial osmotic water flux as measured by light scattering (null point). By this method sigma NaCl was greater than 0.92 for both apical and basolateral membranes with best estimates of 1.0. sigma NaCl was determined by the solvent drag method using the Cl-sensitive fluorescent indicator, 6-methoxy-N-[3-sulfopropyl]quinolinium (SPQ), to detect the drag of Cl into vesicles by inward osmotic water movement caused by an outward osmotic gradient. sigma NaCl was determined by comparing experimental data with theoretical curves generated using the coupled flux equations of Kedem and Katchalsky. By this method we found that sigma NaCl was greater than 0.96 for apical and greater than 0.98 for basolateral membrane vesicles, with best estimates of 1.0 for both membranes. These results demonstrate that sigma NaCl for proximal tubule apical and basolateral membranes are near unity. Taken together with previous results, these data suggest that proximal tubule water channels are long narrow pores that exclude NaCl.
采用两种独立的方法，即诱导渗透法和溶剂拖曳法，来测定从兔近端小管分离出的刷状缘膜囊泡和基底外侧膜囊泡中氯化钠的反射系数（σNaCl）。在诱导渗透法中，将装载蔗糖的囊泡置于停流装置中，使其暴露于不同的内向氯化钠梯度下。通过光散射测量（零点），确定导致初始渗透水通量为零时所需的氯化钠溶液的渗透压，从而得出σNaCl。通过这种方法，顶端膜和基底外侧膜的σNaCl均大于0.92，最佳估计值为1.0。采用溶剂拖曳法，使用对氯离子敏感的荧光指示剂6-甲氧基-N-[3-磺丙基]喹啉鎓（SPQ），通过外向渗透梯度引起的内向渗透水运动来检测氯离子向囊泡内的拖曳，进而确定σNaCl。通过将实验数据与使用凯德姆和卡查尔斯基的耦合通量方程生成的理论曲线进行比较，来确定σNaCl。通过这种方法，我们发现顶端膜囊泡的σNaCl大于0.96，基底外侧膜囊泡的σNaCl大于0.98，两种膜的最佳估计值均为1.0。这些结果表明，近端小管顶端膜和基底外侧膜的σNaCl接近1。结合先前的结果，这些数据表明近端小管水通道是长而窄的孔道，可排除氯化钠。
