Molecular mode of inhibition of glycogenolysis in rat liver by the dihydropyridine derivative, BAY R3401: inhibition and inactivation of glycogen phosphorylase by an activated metabolite

The racemic prodrug BAY R3401 suppresses hepatic glycogenolysis. BAY W1807, the active metabolite of BAY R3401, inhibits muscle glycogen phosphorylase a and b. We investigated whether BAY R3401 reduces hepatic glycogenolysis by allosteric inhibition or by phosphatase-catalyzed inactivation of phosphorylase. In gel-filtered liver extracts, racemic BAY U6751 (containing active BAY W1807) was tested for inhibition of phosphorylase in the glycogenolytic (in which only phosphorylase a is active) and glycogen-synthetic (for the evaluation of a:b ratios) directions. Phosphorylase inactivation by endogenous phosphatase was also studied. In liver extracts, BAY U6751 (0.9-36 micromol/l) inhibited glycogen synthesis by phosphorylase b (notwithstanding the inclusion of AMP), but not by phosphorylase a. Inhibition of phosphorylase-a-catalyzed glycogenolysis was partially relieved by AMP (500 micromol/l). BAY U6751 facilitated phosphorylase-a dephosphorylation. Isolated hepatocytes and perfused livers were tested for BAY R3401-induced changes in phosphorylase-a:b ratios and glycogenolytic output. Though ineffective in extracts, BAY R3401 (0.25 micromol/l-0.5 mmol/l) promoted phosphorylase-a dephosphorylation in hepatocytes. In perfused livers exposed to dibutyryl cAMP (100 micromol/l) for maximal activation of phosphorylase, BAY R3401 (125 micromol/l) inactivated phosphorylase by 63% but glucose output dropped by 83%. Inhibition of glycogenolysis suppressed glucose-6-phosphate (G6P) levels. Activation of glycogen synthase after phosphorylase inactivation depended on the maintenance of G6P levels by supplementing glucose (50 mmol/l). We conclude that the metabolites of BAY R3401 suppress hepatic glycogenolysis by allosteric inhibition and by the dephosphorylation of phosphorylase a.