The effects of adrenergic agonists and antagonists on hepatic glycogenolysis were investigated using isolated perfused rat liver. Comparative studies on the glucose output by various adrenergic agonists and the inhibitory action of various antagonists on epinephrine-induced glycogenolysis indicated that this glycogenolysis was mediated by alpha 1-adrenergic receptors. Epinephrine-induced glucose output from the liver was detected repetitively when epinephrine was repeatedly added to the perfused liver, while the glucose output decreased gradually when epinephrine was infused repetitively to the liver perfused with Ca2+-free buffer. Infusion of epinephrine to the perfused liver caused a release of Ca2+ associated with the glucose output, and a close correlation was found between the amounts of glucose and Ca2+ released from the liver with the infusion of epinephrine at various concentrations. The amounts of glucose production induced by epinephrine in the absence of extracellular Ca2+ were smaller than in the presence of extracellular Ca2+ when the liver received a large or sustained stimulation of epinephrine. These results suggest that the epinephrine mobilizes Ca2+ from the intracellular store to activate glycogenolysis, while entry of extracellular Ca2+ into the cell is required in order to obtain a large or sustained hormonal stimulation of glycogenolysis and to supply the intracellular Ca2+ store.