Fasting of second-day fifth instar larval Manduca sexta leads to a rapid decrease in hemolymph glucose concentration from 3.39+/-0.29 to 0.33+/-0.06 mM in 1 h, along with a decrease in the fructose-2,6-bisphosphate content in the fat body (from 5.92+/-0.31 to 2.80+/-0.47 nmol fructose-2,6-bisphosphate/g fat body in 3 h) and activation of fat body glycogen phosphorylase (from 16% to 55-65% phosphorylase a). During re-feeding an increase in the glucose level in the hemolymph was observed (from 0.36+/-0.05 to 3.91+/-0.36 mM in 3 h), along with an increase in the fructose-2,6-bisphosphate level in the fat body (from 2.88+/-0.47 to 6.66+/-0.42 nmol fructose-2,6-bisphosphate/g fat body in 3 h) and inactivation of fat body glycogen phosphorylase (from 56% to 16% phosphorylase a). These data are consistent with the hypothesis that a decrease in hemolymph glucose both activates fat body glycogen phosphorylase and causes a decrease in fat body fructose-2,6-bisphosphate content. Both of these changes would favor conversion of stored glucose to trehalose in the fat body. When second-day larvae were decapitated, the changes in hemolymph glucose and fat body fructose-2,6-bisphosphate were very similar to those observed in fasting whole insects. These data are consistent with a direct role for glucose in controlling carbohydrate metabolism in Manduca sexta.