Hepatic glycogen metabolism was studied in rats during the period of transition from the fed to fasted states. Glycogenic activity was measured in vivo based on the incorporation of [14C]glucose into liver glycogen. Its changes were almost parallel to the changes in glycogen synthase activity. Progressive accumulation of liver glycogen that occurred in the fed state was associated with a proportional increase in glycogenic activity. Within 4 h after the cessation of food intake, glycogenic activity showed a precipitous fall from the peak to its nadir without significant changes in glycogen content. Meanwhile, the glucose concentration in the portal vein decreased. Upon further development of fasting, glycogenic activity displayed a progressive regain, reciprocally as glycogen contents gradually decreased. The precipitous fall of glycogenic activity during the transition from the fed to fasted states was associated with a transient increase in plasma glucagon, and was partly overcome by the injection of anti-glucagon serum. It is concluded that the fall of portal venous concentration of glucose and secretion of glucagon act as a signal to initiate liver glycogen metabolism characteristic of the fasted or postabsorptive state.