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1. We characterized the basolateral membrane Cl- and K+ conductances of the dark-adapted chick neural retina-retinal pigment epithelium (RPE)-choroid preparation. Conventional microelectrodes were used to measure apical (V(ap)) and basolateral (Vba) membrane voltage, and double-barreled Cl- and K+ selective microelectrodes were used to follow the time course and magnitude of ion concentration changes outside the basolateral (basal) membrane. 2. In response to a fivefold decrease in basal [Cl-]o, Vba rapidly depolarized by 6.4 +/- 0.7 (SE) mV, and the apparent resistance of the basolateral membrane (Rba) increased. The Cl- channel blocker 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) suppressed the Vba depolarization by 40% and blocked the Rba increase. Estimates of the relative Cl- conductance (transference number, TCl) from the DIDS-sensitive component of the Cl- diffusion potential gave an average value for TCl of 0.22 +/- 0.03. 3. Further evidence for a Cl- conductance was obtained by measuring changes in intracellular Cl- activity (aCli) induced by transtissue current. Depolarizing Vba elevated aiCl, whereas hyperpolarizing Vba had the opposite effect, consistent with conductive Cl- movement across the basal membrane. TCl estimated from these data averaged 0.23 +/- 0.02. 4. In response to a sixfold increase in basal [K+]o, Vba depolarized 6.1 +/- 0.8 mV. The amplitude of this K+ diffusion potential was inhibited 44 and 67% by 5 and 10 mM Ba2+, respectively. TK was estimated to be 0.61 +/- 0.05. 5. The rapid c-wave membrane hyperpolarizations in response to the light-evoked decrease in subretinal [K+]o were used to calculate the equivalent resistances of the apical membrane (R(ap)), basolateral membrane (Rba), and the paracellular shunt pathway (Rs). They were 152 +/- 10, 615 +/- 38, and 138 +/- 7 omega.cm2 (n = 11 tissues), respectively. From these data the equivalent electromotive force for the basal (Eba) and apical (Eap) membranes were estimated to be -45 +/- 2 and -77 +/- 1 mV, respectively. This estimate of Eba is in the range of that predicted from our estimates of TCl and TK, indicating that, in the dark-adapted chick retina, the resting conductance of the basal membrane can largely be accounted for by the Cl- and K+ conductances described here.