The effects of minimal increments in plasma insulin concentrations on hepatic glucose production and glucose uptake, skeletal muscle net glycogen synthesis and glycogenolysis, glycogen synthase and phosphorylase activity, glucose-6-phosphate and uridinediphosphoglucose (UDPG) concentrations were examined in 24-h and in 6-h fasted conscious rats. Insulin was infused for 120 min at rates of 1.5, 3, 6, 12, 24, and 108 pmol/kg per min in 24-h fasted rats and at rates of 3, 6, 9, 12, 36, and 108 pmol/kg per min in 6-h fasted rats while endogenous insulin release was inhibited by SRIF infusion and plasma glucose was maintained at the basal level. All rats received an infusion of [3-3H]glucose. The portion of the muscle glucose-6-phosphate (G6P) pool derived from net glycogenolysis was estimated from the ratio of specific activities of muscle UDPG and plasma glucose. Minimal increments in the circulating insulin levels, which did not stimulate glucose uptake, caused: (a) the increase in skeletal muscle glycogen synthase activity and the decrease in the rate of muscle glycogenolysis and in the G6P concentration; (b) the inhibition of hepatic glucose production. Net muscle glycogen synthesis was not stimulated despite submaximal activation of glycogen synthase, and its onset correlated with the rise in muscle G6P levels. Thus, insulin's inhibition of muscle glycogenolysis is the most sensitive insulin action on skeletal muscle and its dose-response characteristics resemble those for the inhibition of hepatic glucose production. These findings indicate that skeletal muscle glycogen synthase may play a major role in carbohydrate homeostasis even under postabsorptive (basal insulin) conditions and support the notion that insulin may exert some of its effects on the liver through an indirect or peripheral mechanism.