Free oxygen radicals reduce bile flow in rats via an intracellular cyclic AMP-dependent mechanism

Background and Aim: Oxidative stress reduces hepatic bile formation. Cyclic adenosine monophosphate (cAMP) is important in bile production; however, the role of basal amounts of intracellular cAMP in bile formation is not known. The aim of this study was to determine whether oxygen radicals reduce bile flow by mechanisms involving intrahepatocyte cAMP levels. Methods: The effect of an oxygen radical (tert-butyl hydroperoxide; t-BHP) on hepatic bile flow was determined in Wistar rats in vivo and in isolated perfused liver. Intracellular cAMP was measured in isolated hepatocytes with and without t-BHP in culture medium, while adenylate cyclase activity was measured in purified plasma membranes. To examine whether intracellular cAMP restoration could reverse t-BHP-induced bile flow reduction, dibutyryl cAMP (DBcAMP), a cell-membrane permeating cAMP, was used to treat isolated liver perfused with t-BHP. Results: Bile flow was significantly reduced 10 min following t-BHP administration in vivo and in isolated perfused liver (control vs 0.1 mg/mL t-BHP in perfusate, 29.3 vs 23.1 mu g/kg per min, P < 0.05). Intracellular cAMP in isolated hepatocytes was reduced by adding t-BHP to the medium; this change was inhibited by DBcAMP. Adenylate cyclase activity in purified liver membrane fractions also was reduced by t-BHP. Administration of DBcAMP reversed bile flow reduction by t-BHP in isolated perfused liver. Conclusions: Bile flow reduction by oxygen radicals was at least partly explained by inactivation of adenylate cyclase causing decreases in intrahepatocytic cAMP. Exogenous DBcAMP administration restored intracellular cAMP preventing bile flow reduction after exposure to oxygen radicals.