The effect of acetylglyceryl ether phosphorylcholine (AGEPC) on glycogenolysis and phosphatidylinositol 4,5-bisphosphate has been studied in rat hepatocytes. Previously, this laboratory demonstrated that AGEPC stimulated glucose output from the perfused rat liver and promoted the breakdown of phosphoinositides in rat hepatocytes (Shukla, S. D., Buxton, D. B., Olson, M.S., and Hanahan, D.J. (1983) J. Biol. Chem. 258, 10212-10214). In the present study, addition of AGEPC (10(-13) to 10(-9) M) to rat hepatocytes failed to stimulate glucose output, whereas epinephrine (10(-5)M) and glucagon (10(-7)M) stimulated glucose output by 100% or more in these same cells. The effects of AGEPC, epinephrine, vasopressin, and glucagon on glycogen phosphorylase activity and the breakdown of phosphatidylinositol 4,5-bisphosphate were compared in hepatocytes. AGEPC (10(-9)M) promoted the breakdown of phosphatidylinositol 4,5-bisphosphate in a fashion similar to epinephrine (10(-5)M) and vasopressin (10(-7)M). In contrast to the two calcium-mobilizing hormones, epinephrine and vasopressin, AGEPC did not cause an activation of glycogen phosphorylase. Glucagon activation of glycogen phosphorylase was not accompanied by a significant effect on phosphatidylinositol 4,5-bisphosphate hydrolysis. Thus, AGEPC is a chemical mediator which induces the degradation of phosphatidylinositol 4,5-bisphosphate without activating glycogenolysis in hepatocytes.