Stripped rabbit ileal mucosa was studied in vitro in Ussing chambers under short-circuit conditions using the pH-stat technique to determine basal rates of luminal alkalinization; the contribution of the shunt pathway to the alkalinization process; the effects of Na, Cl, or HCO3 removal from the bathing solutions on luminal alkalinization; and the effects of epinephrine, ouabain, 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS), acetazolamide, prostaglandin E1 (PGE1), A23187, sugars, or amino acids on the alkalinization process. Results from these studies reveal that, under basal conditions, the rate of luminal alkalinization accounts for 81% of the basal short-circuit current (Isc), although there was no correlation between the rate of alkalinization and Isc. The contribution of the shunt to the alkalinization process accounts for less than 10% of the mucosal-to-serosal HCO3 flux. Removal of Cl from the bathing solutions increased the rate of luminal alkalinization and decreased Isc. Sodium removal from the bathing solutions reduced both Isc and the rate of luminal alkalinization. Addition of DIDS to the luminal or serosal bathing solution reduced luminal alkalinization less than 30%. Acetazolamide, PGE1, and A23187 were all without effect on luminal alkalinization. Addition of 3-O-methyl-D-glucose or L-alanine to the luminal bathing solution did not alter luminal alkalinization but increased Isc, D-Glucose added to the luminal bathing solution reduced luminal alkalinization. This effect appears to result from metabolic acid production since 1) it is not seen with L-alanine or 3-O-methyl-D-glucose; 2) in the absence of HCO3 in the bathing solutions, D-glucose increased luminal acidification; and 3) luminal addition of fructose also reduced the rate of luminal alkalinization. Addition of epinephrine to the serosal bathing solution stimulates a Na-dependent serosal alkalinization process. These results suggest that luminal alkalinization results from Na-dependent, transcellular HCO3 transport and that a Na-dependent, HCO3 absorptive process is stimulated by adrenergic agents.