Requirement of HCO3- for Cl- absorption in seawater-adapted eel intestine

The role of HCO3-/CO2 buffer in Cl- absorption was examined in the in vitro perfused eel intestine adapted to seawater. Cl- absorption, expressed as short-circuit current (Isc), was measured in either 20 mM HCO3-/1% CO2 Ringer or HEPES Ringer, pH 8.0. Unilateral (mucosal or serosal) substitution of HCO3-/CO2 with HEPES/O2 was without effect on Isc and transepithelial voltage (Vt), whereas bilateral removal of HCO3-/CO2 reduced Isc and Vt by 50%, indicating that the presence of HCO3-/CO2 buffer at one side of the epithelium is sufficient to keep Cl- absorption at the maximum rate. We examined in further detail the individual components of the HCO3-/CO2 system that stimulates Cl- absorption. We found that, in tissues bathed with HEPES Ringer, addition of 1% CO2 to the luminal or serosal solution (final pH = 7.6 in the chamber) had no effect on Isc and Vt, while both electrical parameters could be restored to control values by unilateral (luminal or serosal) substitution of HEPES Ringer with 20 mM HCO3-/1% CO2 Ringer or 20 mM HCO3- alone. Stimulation of Isc induced by unilateral (luminal or serosal) HCO3-/CO2 was inhibited by luminal or serosal 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS) (0.25 mM) or by serosal Na+ removal, whereas amiloride (1 mM), luminal or serosal, had no effect. Acetazolamide (0.1 mM, both sides) inhibited stimulation of Isc induced by luminal addition of HCO3-/CO2, whereas it was without effect when HCO3-/CO2 was added serosally or bilaterally.