Evidence against luminal one-for-one Cl(-)-HCO-3 exchange in urodele small intestine

The ratio of Cl- absorbed to HCO3- secreted by the in vitro small intestine of Amphiuma was measured using 36Cl and titration. The aim was to estimate the stoichiometry and thereby elucidate the underlying transport mechanisms. For every mole of HCO3- secreted 1.8 mol of Cl- underwent net absorption. Indirect measures of net Cl- absorption and HCO3- secretion were validated. Several known and putative Cl- transport inhibitors were examined for their ability to inhibit the anion transport events. Disulfonic stilbenes [4-acetamido-4'-isothiocyanostilbene-2',2'-disulfonic acid and 4,4'-diisothiocyano-2,2'-disulfonate stilbene (DIDS)] and the diuretics piretanide and furosemide inhibited the Cl- absorptive flux (JClm----s) and simultaneously the HCO3- secretory flux (JHCO3-). The diuretics acetazolamide and bumetanide also reduced JHCO3- and JClm----s, although the latter effect was not statistically significant. The ratio of inhibition, delta JClm----s/delta JHCO3-, varied from 1.2 to 1.8 for the different inhibitors. The presence of Cl(-)-HCO3- exchange at the serosal membrane was deduced from the reduction of JClm----s and JHCO3- by serosally added stilbenes, the reduction of Cl- absorption when serosal Cl- was replaced (reduced Cl(-)-Cl- exchange), inhibition of the secretory-to-mucosal Cl- flux by serosal stilbenes, and enhancement of JHCO3- when serosal medium HCO3- was elevated. Addition of DIDS to the mucosal medium did not alter the secretory flux. The observations are not consistent with one-for-one exchange of Cl- for HCO3- at the mucosal membrane. Other possible mechanisms including rheogenic Cl- uptake in parallel with electrodiffusional efflux of HCO3- are discussed. The observed coupling ratio is compatible with a one-for-one exchange of Cl- for HCO3- at the serosal membrane.