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 (V(A) 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 Ba2+-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 T(K) almost-equal-to 0.9 and T(Cl) almost-equal-to 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 (R(A)) and basolateral (R(B)) cell membranes, as well as the paracellular or shunt resistance (R(S)). 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 (E(A)) and basolateral (E(B)) membranes were also calculated. They are: -69 +/- 5.0 and -24 +/- 5.0 mV, respectively. 4. Intracellular Cl- activity (a(cl)i) was measured using double-barrelled ion-selective microelectrodes. In the steady state a(Cl)i = 61 +/- 4.0 mM and the Nernst potential E(Cl) = - 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. Phase 2 is a delayed hyperpolarization of the basolateral membrane. Phase 3 is most probably caused by a decrease in [K+]i and a decrease in apical membrane K+ conductance. These voltage changes were accompanied by a monotonic decrease in an a(Cl)i of 26 +/- 3.0 mM (n = 4). 6. The second of the three phases that is produced following apical K+ reduction from 5 to 2 mM is characterized by an extra or delayed hyperpolarization at the basolateral membrane. During this phase a(Cl)i decreased by almost-equal-to 14 mM. The delayed hyperpolarization was blocked by apical bumetanide or basal DIDS, indicating that the chloride transport pathway is a primary determinant of this response. It is suggested that the drop in a(Cl)i, expressed at the basal membrane Cl- conductance during phase 2, generates the 'fast oscillation' component of the DC-recorded electroretinogram or electro-oculogram that is measured clinically across the human eye.
