Acetazolamide-induced changes of the apical (Vap) and basal (Vba) membrane potentials of the retinal pigment epithelial cell were studied in an in-vitro retinal pigment epithelium (RPE)-choroid of the frog. Both Vap and Vba were hyperpolarized by acetazolamide placed on either the apical or basal side of the RPE. In all cases, acetazolamide on the apical side hyperpolarized Vba more than Vap and decreased the transepithelial potential (TEP) across the RPE. In most cases, acetazolamide on the basal side hyperpolarized both Vap and Vba to almost equal degrees and hardly changed the TEP. We conclude that the Diamox response (a decrease of the ocular standing potential induced by an intravenous sodium acetazolamide) may be triggered by effects of acetazolamide on the apical side of the RPE and generated mainly by a hyperpolarization of the basal membrane of the RPE cell.