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Apical and basal membrane ion transport mechanisms in bovine retinal pigment epithelium.

1991; Wiley; Volume: 435; Issue: 1 Linguagem: Inglês

10.1113/jphysiol.1991.sp018518

1469-7793

Daniel P. Joseph, Sheldon S. Miller,

Neuroscience and Neural Engineering

1. Intracellular voltage recordings using conventional and double‐barrelled chloride‐selective microelectrodes have been used to identify several transport mechanisms at the apical and basolateral membranes of the isolated bovine retinal pigment epithelium (RPE)‐choroid preparation. Intracellular recordings were obtained from two cell populations, melanotic (pigmented) and amelanotic (non‐pigmented). The electrical properties of these two populations are practically identical. For melanotic cells the average apical resting membrane potential (VA) is ‐61 +/‐ 2 mV (mean +/‐ S.E.M., n = 49 cells, thirty‐three eyes). For these cells the ratio of apical to basolateral membrane resistance (a) was 0.22 +/‐ 0.02. The mean transepithelial voltage and resistance were 6 +/‐ 1 mV and 138 +/‐ 7 omega cm2, respectively. 2. The apical membrane, which faces the distal retina, contains a Ba(2+)‐inhibitable K+ conductance and a ouabain‐inhibitable, electrogenic Na(+)‐K+ pump. In addition it contains a bumetanide‐sensitive mechanism, the putative Na(+)‐K(+)‐Cl‐ cotransporter. The basolateral membrane contains a DIDS (4,4'‐diisothiocyanostilbene‐2,2'‐disulphonic acid)‐inhibitable chloride channel. The relative conductances of the apical and basolateral membranes to K+ and Cl‐ are TK approximately 0.9 and TCl approximately 0.7, respectively. 3. The ouabain‐induced fast phase of apical membrane depolarization (0‐30 s) was used to calculate the equivalent resistances of the apical (RA) and basolateral (RB) cell membranes, as well as the paracellular or shunt resistance (RS). They are: 3190 +/‐ 400, 17920 +/‐ 2730 and 2550 +/‐ 200 omega (mean +/‐ S.E.M., n = 9 tissues), respectively. From these data the equivalent electromotive forces (EMF) at the apical (EA) and basolateral (EB) membranes were also calculated. They are: ‐69 +/‐ 5.0 and ‐24 +/‐ 5.0 mV, respectively. 4. Intracellular Cl‐ activity (aiCl) was measured using double‐barreled ion‐selective microelectrodes. In the steady state aiCl = 61 +/‐ 4.0 mM and the Nernst potential ECl = ‐13.5 +/‐ 1.5 mV (mean +/‐ S.E.M., n = 4). 5. In the intact eye or in retina, RPE‐choroid preparations it has been shown that the transition between light and dark alters the K+ concentration in the extracellular (or subretinal) space between the photoreceptors and the apical membrane of the RPE. These light‐induced changes in subretinal [K+]o were qualitatively simulated in vitro by altering apical K+ between 5 and 2 mM. This produced a sequence of voltage changes at the apical and basolateral membranes that had three operationally distinct phases. Phase 1 is generated by the combination of an apical membrane K+ diffusion potential and inhibition of the electrogenic Na(+)‐K+ pump.(ABSTRACT TRUNCATED AT 400 WORDS)