We examined the influence of 8 wk of endurance training on basal levels of fat oxidation and its association with changes in norepinephrine (NE) kinetics, resting metabolic rate (RMR), and body composition in 18 healthy elderly persons (66.1 +/- 1.4 yr; 10 men, 8 women). Fatty acid appearance rate and total body fat oxidation were determined from [14C]palmitate infusion and indirect calorimetry, NE kinetics were determined from infusions of [3H]NE, RMR was determined from the ventilated hood technique, and body composition was determined from underwater weighing. Endurance training increased peak oxygen consumption by 11% (1.9 +/- 0.1 to 2.1 +/- 0.1 l/min; P < 0.01) and increased RMR by 7% (1.20 +/- 0.02 to 1.28 +/- 0.02 kcal/min; P < 0.01). Endurance training increased NE appearance rate by 35% (0.51 +/- 0.04 to 0.69 +/- 0.04 micrograms/min; P < 0.01), whereas no change in NE clearance was noted. Endurance training increased fat oxidation by 22% (201.0 +/- 11.2 vs. 244.0 +/- 15.2 mumol/min; P < 0.01) but did not alter fatty acid appearance rate. Approximately two-thirds of the variation (r2 = 0.65) for the increase in fat oxidation was explained by increased NE appearance rate (r2 = 0.51; P < 0.01) and changes in fat-free weight (r2 = 0.14; P < 0.01). We conclude that 1) endurance training shifts in vivo basal substrate utilization toward greater fat oxidation in elderly individuals and 2) enhanced fat oxidation is associated with increased activity of the sympathetic nervous system and alterations in fat-free mass.