The mechanism of recovery from an acid load in primary cultures of rabbit choroid plexus epithelium (CPE) was examined, with emphasis on Na(+)-dependent antiports. Cells were incubated in saline solutions buffered to pH 7.38 with either HEPES or HCO3- plus 95% O2/5% CO2. Intracellular pH (pHi) was determined from the steady-state distribution of [14C]benzoate. Recovery after acidification with NH4Cl was rapid (t1/2 = 5 min) and was dependent on external Na+ (EC50 = 12 mM). Hexamethyleneamiloride and ethylisopropylamiloride, potent inhibitors of the Na+/H+ antiport, blocked 80% of recovery when [Na+] was 5 mM with IC50 values of 100 nM. However, neither drug blocked recovery in normal [Na+]. 4,4'-Diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), an inhibitor of Cl-/HCO3- antiports, blocked recovery of pHi in a dose-related fashion in the presence of bicarbonate, but not in the presence of HEPES. No inhibition occurred with benzamil, an amiloride congener with high affinity for the Na+ channel, nor with dimethylbenzamil, an inhibitor of Na+/Ca2+ exchange. The carbonic anhydrase inhibitor acetazolamide also did not alter recovery from acidification. In CPE that had been pH-clamped with nigericin and KCl, the initial rate of 22Na+ uptake was very rapid (227 pmol/micrograms of DNA/min at pH 6.2), was dependent on external [Na+] with an EC50 value of 8 mM, and was inversely related to the pH of the medium. The maximal inhibition of 22Na+ uptake by hexamethyleneamiloride was 60% with an IC50 value of 76 nM.(ABSTRACT TRUNCATED AT 250 WORDS)