Transport properties of 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxyinosine (DDI) were characterized in the isolated rat choroid plexus. AZT and DDI competitively inhibited the active transport of [3H]benzylpenicillin, a prototypic organic anion, with Ki values of 85.4 +/- 13.1 and 155 +/- 22 microM, respectively. Accumulation of [3H]DDI was against an electrochemical potential via a saturable process (K(m) = 29.7 +/- 4.9 microM, Vmax = 13.5 +/- 2.4 pmol min-1/microL tissue) that was inhibited by metabolic inhibitors (carbonylcyanide p-trifluoromethoxyphenylhydrazone, 10 microM, and rotenone, 30 microM) and sulphydryl reagents (p-chloromercuribenzoic acid, 100 microM, and p-chloromercuribenzenesulphonic acid, 100 microM), but did not require an inwardly directed Na+ gradient. Accumulation of [3H]DDI was inhibited by benzylpenicillin and AZT in a dose-dependent manner, with IC50 values of 91.6 +/- 28.9 and 294 +/- 84 microM, respectively. In contrast, no significant accumulation of [3H]AZT was observed. These results suggest that DDI is transported, at least in part, by the transport system for organic anions located on the rat choroid plexus, whereas AZT is recognized, but not transported by this system.