Muscle metabolism, including the role of pyruvate dehydrogenase (PDH) in muscle lactate (Lac-) production, was examined during incremental exercise before and after 7 days of submaximal training on a cycle ergometer [2 h daily at 60% peak O2 uptake (VO2 max)]. Subjects were studied at rest and during continuous steady-state cycling at three stages (15 min each): 30, 65, and 75% of the pretraining VO2 max. Blood was sampled from brachial artery and femoral vein, and leg blood flow was measured by thermodilution. Biopsies of the vastus lateralis were obtained at rest and during steady-state exercise at the end of each stage. VO2 max, leg O2 uptake, and the maximum activities of citrate synthase and PDH were not altered by training; muscle glycogen concentration was higher. During rest and cycling at 30% VO2 max, muscle Lac- concentration ([Lac-]) and leg efflux were similar. At 65% VO2 max, muscle [Lac-] was lower (11.9 +/- 3.2 vs. 20.0 +/- 5.8 mmol/kg dry wt) and Lac- efflux was less [-0.22 +/- 0.24 (one leg) vs. 1.42 +/- 0.33 mmol/min] after training. Similarly, at 75% VO2 max, lower muscle [Lac-] (17.2 +/- 4.4 vs. 45.2 +/- 6.6 mmol/kg dry wt) accompanied less release (0.41 +/- 0.53 vs. 1.32 +/- 0.65 mmol/min) after training. PDH in its active form (PDHa) was not different between conditions. Calculated pyruvate production at 75% VO2 max fell by 33%, pyruvate reduction to lactate fell by 59%, and pyruvate oxidation fell by 24% compared with before training. Muscle contents of coenzyme A and phosphocreatine were higher during exercise after training. Lower muscle lactate production after training resulted from improved matching of glycolytic and PDHa fluxes, independently of changes in muscle O2 consumption, and was associated with greater phosphorylation potential.