Bicarbonate secretion occurs in almost all segments of the gastrointestinal tract. This study examined HCO(3)(-) secretion in the ileum, since it is less understood than HCO(3)(-) secretion in other intestinal segments. Mouse ileal mucosa was mounted in vitro in Ussing chambers, and the mucosal alkalinization rate (J(OH)) was determined by pH stat titration, while the mucosal side was bathed with a buffer-free solution (100% O(2)) and the serosal side with a HCO(3)(-)/CO(2)-buffered solution. The transmural potential difference (PD) was recorded. The mucosal alkalinization rate (J(OH)) was higher in the presence of mucosal Cl(-) than in its absence. Forskolin, an activator of adenylate cyclase, enhanced J(OH) and PD in both the presence and absence of mucosal Cl(-). Mucosal SO(4)(2-) also caused an increase in J(OH), although the magnitude was smaller than that induced by Cl(-). Mucosal Cl(-)-dependent J(OH) was partially inhibited by acetazolamide, 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB), tenidap and probably also by niflumic acid, but not by glibenclamide, DIDS or bumetanide. The forskolin-induced J(OH) value and PD were both inhibited by NPPB and probably also by tenidap. It is concluded that HCO(3)(-)- secretion in the ileum follows a mucosal Cl(-)-dependent pathway and a cAMP-activated pathway, each being distinct from each other. The Cl(-)-dependent pathway is probably mediated by the slc26a6 anion exchanger, and possibly also by the slc26a3 anion exchanger. The cAMP-activated HCO(3)(-) secretion is probably mediated by the cystic fibrosis transmembrane conductance regulator.