The present study was performed to test the hypothesis that the interstitial glucose concentration in human skeletal muscle is decreased for a prolonged period following a single bout of exercise, while blood flow has returned to resting levels. Muscle interstitial concentrations of glucose, lactate, pyruvate and urea were monitored in six healthy individuals during 8 h following a 2-h one-leg exercise session by microdialysis at low perfusion flow rate. Simultaneously the blood flow was measured by the microdialysis ethanol technique. The blood glucose and the control leg interstitial glucose concentrations were stable during the experiment averaging 5.7 +/- 0.1 and 4.1 +/- 0.3 mm, respectively. In contrast, the interstitial glucose concentration in the exercise leg was markedly decreased, averaging 1.9 +/- 0.5 mm, during the first 5.5 h following exercise (P < 0.01), after which it returned towards normal values. Muscle blood flow at the site of the microdialysis catheter, measured as the ethanol outflow-to-inflow ratio, did not change significantly over time in the control or exercise leg and did not differ significantly between the two legs. Interstitial concentrations of lactate, pyruvate and urea were not significantly different between the control and exercise leg. The study shows that the interstitial glucose concentration in skeletal muscle is markedly decreased for several hours following a single exercise session. The decreased interstitial glucose concentration may serve to limit the rate of post-exercise muscle glucose uptake to a rate compatible with normal blood glucose levels and may also be speculated to have a positive long-term health implication by augmenting muscle insulin sensitivity.