Inhibition of biliary bicarbonate secretion in ethinyl estradiol-induced cholestasis is not associated with impaired activity of the Cl-/HCO3- exchanger in the rat.

Abstract: Background/Aims: Bicarbonate is a major component of bile salt independent bile flow which is impaired in ethinyl estradiol (EE)-cholestasis. To examine this subject in EE-cholestasis, we studied: 1) basal and glucagon-stimulated biliary bicarbonate secretion both in vivo and in the isolated perfused rat liver (IPRL); 2) H+/HCO3- transport processes in isolated rat hepatocyte couplets. Methods: Rats received EE (5 mg . kg b.w.(-1)) for 5 days. Intracellular pH (pH(i)) was measured (BCECF-AM) using a single-cell microfluorimetric setup. Results: Bile flow was markedly (p<0.01) decreased in EE-treated rats. Bicarbonate concentration in bile was decreased (p<0.01) and bicarbonate secretion was 2.5-fold lower in EE-treated animals than in controls, both in bile-fistula rats [19.5+/-5.1 (n=23) vs 54.2+/-5.7 (n=20) nmol . min(-1). g liver(-1); p<0.01] and in the IPRL [11+/-2 (n=8) vs 24+/-3 (n=8) nmol . min(-1). g liver(-1); p<0.01]. In control IPRL, a bile/perfusate gradient for bicarbonate is maintained, while it is lost in EE-treated IPRL because of the lower bicarbonate concentration in bile. Glucagon stimulated bile flow and bicarbonate secretion to a similar extent in EE-treated and control IPRL (+25% vs +23%). Resting pH(i) of EE-treated hepatocyte couplets was higher in comparison with controls in KRB [7.25+/-0.07 (n=35) vs 7.20+/-0.05 (n=33); p<0.02] but similar in Hepes [7.08+/-0.07 (n=24) vs 7.05+/-0.06 (n=26)]. Basal activity of the Cl-/HCO3- exchanger was similar in EE-treated and control hepatocyte couplets [H+ flux = 2.87+/-1.12 (n=18) vs 3.01+/-1.23 mM/min (n=15)] and was stimulated to a similar extent by glucagon. Na+/HCO3- symport activity was increased in EE-treated hepatocyte couplets (p<0.05) while the Na+/H+ exchanger was unchanged. Conclusions: Bicarbonate biliary secretion is markedly impaired during EE-cholestasis in association with a marked decrease of bile salt independent bile flow. However, the Cl-/HCO3- exchanger and its hormonal regulation are normal, indicating that the lower bicarbonate excretion in EE-cholestasis is not due to a compromised activity of this anion exchanger. Since the bile/perfusate gradient for bicarbonate is dissipated in EE-treated IPRL, the impaired bicarbonate excretion could be caused by a reflux of biliary bicarbonate via leaky tight junctions.