Hernandez E V, Hu J G, Frambach D A, Gallemore R P
Jules Stein Eye Institute, University of California, Los Angeles School of Medicine.
Invest Ophthalmol Vis Sci. 1995 Jan;36(1):113-22.
To identify K+ conductances on the apical and basolateral membranes in cultured monolayers of fetal bovine and human retinal pigment epithelium (RPE).
Bovine and human RPE cells were grown on a permeable substrate for an average of 4 and 25 months, respectively, mounted in a modified Ussing chamber that allowed rapid solution changes at both membranes, and perfused with modified Ringer's solutions. Conventional microelectrode recording techniques were used to record intracellularly from RPE cells.
Electrical parameters under control conditions for bovine (n = 11) and human (n = 7) cultures respectively, were: transepithelial potential, 0.6 +/- 0.2 and 3.2 +/- 0.5 mV; Rt, 70 +/- 12 and 227 +/- 30 ohm.cm2; and Vap, -61 +/- 3 and -51 +/- 2 mV (mean +/- SEM). The relative K+ conductance (TK) was estimated from responses to 10-fold increases in [K+]o. For the apical and basal membranes, the values for TK were 0.65 and 0.37, respectively, in bovine and 0.33 and 0.45, respectively, in human RPE. Barium applied to either surface of the cultures produced membrane depolarizations and suppressed the responses significantly to K+. In bovine, a 10-fold decrease in basal [K+]o hyperpolarized Vba by 2 +/- 1 mV; a similar decrease in apical [K+]o hyperpolarized Vap by 14 +/- 1 mV. At both membranes, perfusion with Ba2+ unmasked a significant membrane depolarization induced by lowering [K+]o; this phenomenon, observed previously at the apical membrane in fresh explant bovine, frog, and toad RPE, appears to be an unmasking of Na+/K+ pump modulation by K+.
These results indicate the presence of Ba2+ sensitive K+ conductances at the apical and basal membranes of confluent monolayers of cultured fetal human and bovine RPE and the presence of Na+/K+ pump sites at both membranes of cultured bovine cells.
鉴定胎牛和人视网膜色素上皮（RPE）培养单层细胞顶膜和基底外侧膜上的钾离子电导。
牛和人RPE细胞分别在可渗透基质上生长平均4个月和25个月，安装在改良的尤斯灌流小室中，该小室允许对两侧膜快速更换溶液，并用改良的林格氏液灌注。采用传统微电极记录技术从RPE细胞进行细胞内记录。
牛（n = 11）和人（n = 7）培养物在对照条件下的电参数分别为：跨上皮电位，0.6±0.2和3.2±0.5 mV；Rt，70±12和227±30 ohm·cm2；以及Vap，-61±3和-51±2 mV（平均值±标准误）。相对钾离子电导（TK）通过对[K+]o增加10倍的反应进行估算。对于顶膜和基底膜，牛RPE中TK值分别为0.65和0.37，人RPE中分别为0.33和0.45。将钡应用于培养物的任何一侧表面都会导致膜去极化，并显著抑制对钾离子的反应。在牛中，基底[K+]o降低10倍使Vba超极化2±1 mV；顶膜[K+]o类似降低使Vap超极化14±1 mV。在两侧膜上，用Ba2+灌注揭示了因降低[K+]o诱导的显著膜去极化；这种现象先前在新鲜外植牛、蛙和蟾蜍RPE的顶膜中观察到，似乎是钾离子对钠钾泵调节的揭示。
这些结果表明在培养的胎牛和人RPE汇合单层细胞的顶膜和基底膜存在钡敏感的钾离子电导，并且在培养的牛细胞的两侧膜存在钠钾泵位点。
