High plasma fatty acid availability and a positive energy balance in sedentary individuals reduce insulin sensitivity. This study's purpose was to determine whether high plasma fatty acid availability and systemic caloric excess after exercise also impair insulin sensitivity. On two separate occasions, seven nonobese women performed 90 min of exercise at ∼65% peak oxygen uptake. In one trial, a lipid + heparin emulsion (Lipid) was infused overnight to increase plasma fatty acid availability. In the other trial, saline was infused as control. The next morning, a muscle biopsy was taken to measure muscle glycogen and intramuscular triglyceride (IMTG) concentrations. Three hours after the overnight infusion was stopped, insulin sensitivity was assessed with an intravenous glucose tolerance test, using minimal model analysis (S_i). During the overnight infusions, plasma fatty acid concentration was approximately fourfold higher [means (SD): 0.84 (0.36) vs. 0.22 (0.09) mmol/l; P = 0.003], and the next morning IMTG concentration was ∼30% greater [49.2 (6.6) vs. 38.3 (7.7) mmol/kg dry wt; P = 0.036] in Lipid compared with saline. However, muscle glycogen concentration was not different between trials (P = 0.82). Lipid caused a 24-h surplus of ∼1100 kcal above energy balance (P = 0.00001), whereas energy balance was maintained in saline. Despite these differences in fatty acid and energy availability, S_i the morning after exercise was not different between trials (P = 0.72). Thus insulin sensitivity the morning after a single exercise session was not reduced despite overnight exposure to a fourfold increase in plasma fatty acid concentration, elevated IMTG concentration, and systemic delivery of ∼1,100-kcal excess.