Intestinal HCO₃⁻secretion and NaCl absorption are essential for counteracting dehydration in marine teleost fish. We investigated how these two processes are coordinated in toadfish. HCO₃⁻stimulated a luminal positive short-circuit current ( I_sc) in intestine mounted in Ussing chamber, bathed with the same saline solution on the external and internal sides of the epithelium. The I_scincreased proportionally to the [HCO₃⁻] in the bath up to 80 mM NaHCO₃, and it did not occur when NaHCO₃was replaced with Na⁺-gluconate or with NaHCO₃in Cl⁻-free saline. HCO₃⁻(20 mM) induced a ∼2.5-fold stimulation of I_sc, and this [HCO₃⁻] was used in all subsequent experiments. The HCO₃⁻-stimulated I_scwas prevented or abolished by apical application of 10 μM bumetanide (a specific inhibitor of NKCC) and by 30 μM 4-catechol estrogen [CE; an inhibitor of soluble adenylyl cyclase (sAC)]. The inhibitory effects of bumetanide and CE were not additive. The HCO₃⁻-stimulated I_scwas prevented by apical bafilomycin (1 μM) and etoxolamide (1 mM), indicating involvement of V-H⁺-ATPase and carbonic anhydrases, respectively. Immunohistochemistry and Western blot analysis confirmed the presence of an NKCC2-like protein in the apical membrane and subapical area of epithelial intestinal cells, of Na⁺/K⁺-ATPase in basolateral membranes, and of an sAC-like protein in the cytoplasm. We propose that sAC regulates NKCC activity in response to luminal HCO₃⁻, and that V-H⁺-ATPase and intracellular carbonic anhydrase are essential for transducing luminal HCO₃⁻into the cell by CO₂/HCO₃⁻hydration/dehydration. This mechanism putatively coordinates HCO₃⁻secretion with NaCl and water absorption in toadfish intestine.

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