Prostaglandin E2 has been reported both to stimulate glycogen-phosphorylase activity (glycogenolytic effect) and to inhibit the glucagon-stimulated glycogen-phosphorylase activity (antiglycogenolytic effect) in rat hepatocytes. It was the purpose of this study to resolve this apparent contradiction and to characterize the signalling pathways and receptor subtypes involved in the opposing prostaglandin E2 actions. Prostaglandin E2 (10 microM) increased glucose output, glycogen-phosphorylase activity and inositol trisphosphate formation in hepatocyte cell culture and/or suspension. In the same systems, prostaglandin E2 decreased the glucagon-stimulated (1 nM) glycogen-phosphorylase activity and cAMP formation. The signalling pathway leading to the glycogenolytic effect of PGE2 was interrupted by incubation of the hepatocytes with 4 beta-phorbol 12-myristate 13-acetate (100 nM) for 10 min, while the antiglycogenolytic effect of prostaglandin E2 was not attenuated. The signalling pathway leading to the antiglycogenolytic effect of prostaglandin E2 was interrupted by an incubation of cultured hepatocytes with pertussis toxin (100 ng/ml) for 18 h, whereas the glycogenolytic effect of prostaglandin E2 was enhanced. The EP1/EP3 prostaglandin-E2-receptor-specific prostaglandin E2 analogue Sulproston had a stronger glycogenolytic potency than the EP3 prostaglandin-E2-receptor-specific prostaglandin E2 analogue Misoprostol. The antiglycogenolytic potency of both agonists was equal. It is concluded that the glycogenolytic and the antiglycogenolytic effects of prostaglandin E2 are mediated via different signalling pathways in hepatocytes possibly involving EP1 and EP3 prostaglandin E2 receptors, respectively.