DIFFERENTIAL-EFFECTS OF CALYCULIN-A AND OKADAIC ACID ON THE GLUCOSE-INDUCED REGULATION OF GLYCOGEN-SYNTHASE AND PHOSPHORYLASE ACTIVITIES INCULTURED-HEPATOCYTES

The effects of the phosphatase inhibitors calyculin A and okadaic acid were investigated to determine the roles of protein phosphatases type 1 and 2A in the regulation of the activities of glycogen synthase and phosphorylase by glucose in a primary culture of hepatocytes. Glycoge n synthesis, as measured by the incorporation of labelled glucose into glycogen, was inhibited in a dose-dependent manner by calyculin A (IC 50 = 2.2 nM) and okadaic acid (IC50 = 14 nM). Glucose-induced activati on of glycogen synthase was inhibited by calyculin A and okadaic acid with IC50 values of 3.7 nM and 90 nM, respectively. Phosphorylase was simultaneously activated by these inhibitors with calyculin A again be ing more active (P < 0.001) than okadaic acid. The differing potencies (P < 0.001) of these inhibitors on the activities of glycogen synthas e and phosphorylase were also observed with varying concentrations of glucose (5.6-60 mM) in the medium and at different incubation periods upto 120 min. It has been previously shown that both inhibitors inhibi t protein phosphatase-2A with equal potency and calyculin A is a more potent inhibitor of protein phosphatase-1 than okadaic acid. Heat- and proteinase-treated cytosolic fractions from hepatocytes incubated wit h calyculin A and okadaic acid showed similar differential inhibitory activities towards purified types 1 and 2-A protein phosphatases. Henc e, these data provide further evidence that protein phosphatase type-1 plays a major role in the control of glycogen synthesis by regulating the activities of glycogen synthase and phosphorylase.