Role of bicarbonate in biliary excretion of diisothiocyanostilbene disulfonate

Abstract

Hepatic transport of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) was studied in isolated perfused rat livers and in isolated rat hepatocytes to determine if DIDS-induced decrease in biliary HCO3- excretion is due to a DIDS-HCO3- exchange and/or due to inhibition of Cl(-)-HCO3- exchange. In isolated perfused rat livers, DIDS reversibly decreased biliary HCO3- concentration and excretion. The changes in biliary HCO3- concentration were inversely related to biliary DIDS concentration. DIDS was concentrated in bile, indicating active hepatic transport. Replacement of perfusate HCO3- with equimolar dimethyloxazolidinedione (DMO) or tricine decreased biliary excretion, but not hepatic uptake, of DIDS. Biliary excretion of DIDS was also associated with a decrease in bile pH, and this decrease in pH was greater in the presence of HCO3-. HCO3-, but not DMO or tricine, stimulated DIDS efflux from preloaded hepatocytes. DIDS efflux was also temperature dependent and increased with increasing extracellular pH. Collectively, these results are consistent with the presence of a DIDS-HCO3- (OH-) exchange mechanism at the canalicular membrane. HCO3(-)-dependent Cl- uptake in hepatocytes was competitively inhibited by DIDS (Ki = 0.24 mM), confirming the presence of DIDS-inhibitable Cl(-)-HCO3- exchange. However, the ability of DIDS to decrease biliary HCO3- excretion persisted when perfusate Cl- was replaced by isethionate. Moreover, biliary HCO3- concentration returned to base line despite the presence of 2-6 mM DIDS in bile. Thus it seems unlikely that the inhibition of Cl(-)-HCO3- exchange by DIDS is a major mechanism of inhibition of HCO3- excretion. We, therefore, conclude that a DIDS-HCO3- (OH-) exchange at the canalicular membrane is the most likely explanation for the observed decrease in biliary HCO3- excretion.

Intracellular pH regulation in isolated rat bile duct epithelial cells.

“…Hepatocyte anion transporters are known to be relatively insensitive to DIDS (21,39,40). Moreover, in the isolated perfused rat liver preparation where the integrated function ofhepatocytes and ductal system can be evaluated, the liver excretes DIDS into the bile at biliary concentrations up to 5 mM resulting in a decrease in biliary HCO-concentration that is reversible after DIDS withdrawal (45). Thus, in the intact organ, high DIDS concentrations are tolerated without irreversible inhibition.…”

Hepatocyte Heterogeneity in Bile Formation and Hepatobiliary Transport of Drugs

“…Mechanisms of Canalicular Transport There is evidence that transport of or ganic anions at this level occurs through ex change with one or more HCO3 or SO4" ions, [79,161], The bicarbonate ion may enter bile through antiport with chloride and con sequently its biliary accumulation is pro duced by the prevailing chloride gradient across the membrane that, in its turn is pro duced by a conductive (membrane-poten tial-driven) pathway [3,79], Potential carrier proteins for facilitated diffusion of organic anions at the canalicular level have recently been isolated and poly clonal antibodies have been raised against one of the candidates [79], Interestingly, photoaffinity labeling with non-bile acid anionic and cationic probes [77] in canalicular membranes all show spe cific labeling in the 100-kD region [40, 79,155], Probably several, closely related, car rier proteins are involved [79], Bile acid transport can operate separately from biliary excretion of anionic compounds such as BSP and also from cationic agents [3,89,155], Infusion of large amounts of bile acids stimulates the biliary output of some but not all organic anions and cations [75,76], In mutant animals with clearly deficient biliary excretion of most organic anions, bile acid output and organic cation transport is Acinar Heterogeneity in Canalicular Transport Both for taurocholic acid ( fig. 15) and taurodeoxycholic acid, it was shown that ex cretion is slower in retrograde perfusions where zone-3 cells are mainly involved in biliary excretion than in antegrade perfu sions where zone-1 cells are mainly involved [22,29,134], Two explanations can be giv en: either zone-3 cells have less canalicular carriers or zone-3 cells have more bile acid binding protein, resulting in lower free con centrations and thus slower biliary excre tion.…”

“…Mechanisms of Bile Formation Bile flow is maintained by the active se cretion of bile acids [3,31,89,155], GSH [142,143,148] and bicarbonate [79,161], Separate ATP-dependent carrier systems for these agents have recently been described [79,89,142,143,155], In addition, the neg ative membrane potential may help in extru sion of these anionic compounds [79,111], Excretion of GSH induces a part of the so-junctional complexes. Various inorganic ion exchangers such as a CIVHCO3 transporter [79,161] have been demonstrated in the can aliculus as well as transport carriers for bile acids [89,155], non-bile acid organic anions [154,156], organic cations [75,77,155] and uncharged compounds.…”

“…Various inorganic ion exchangers such as a CIVHCO3 transporter [79,161] have been demonstrated in the can aliculus as well as transport carriers for bile acids [89,155], non-bile acid organic anions [154,156], organic cations [75,77,155] and uncharged compounds.…”

The anion transport inhibitor DIDS increases rat hepatocyte K+ conductance via uptake through the bilirubin pathway.

“…In isolated perfused rat liver, DIDS is taken up and secreted into bile (Anwer, Nolan & Hardison, 1988). In guinea-pig liver, SITS is transported into bile at rates comparable to those of BSP secretion (Rutishauser, 1983).…”