Positron emission tomography permits noninvasive measurement of regional glucose uptake in vivo in humans. We employed this technique to determine the effect of FFA on glucose uptake in leg, arm, and heart muscles. Six normal men were studied twice under euglycemic hyperinsulinemic (serum insulin approximately 500 pmol/liter) conditions, once during elevation of serum FFA by infusions of heparin and Intralipid (serum FFA 2.0+/-0.4 mmol/liter), and once during infusion of saline (serum FFA 0.1+/-0.01 mmol/liter). Regional glucose uptake rates were measured using positron emission tomography-derived F-18-fluoro-2-deoxy-D-glucose kinetics and the three-compartment model described by Sokoloff (Sokoloff, L., M. Reivich, C. Kennedy, M. C. Des Rosiers, C. S. Patlak, K. D. Pettigrew, 0. Sakurada, and M. Shinohara. 1977. J. Neurochem. 28: 897-916). Elevation of plasma FFA decreased whole body glucose uptake by 31+/-2% (1,960+/-130 vs. 2,860+/-250-mu-mol/min, P < 0.01, FFA vs. saline study). This decrease was due to inhibition of glucose uptake in the heart by 30+/-8% (150+/-33 vs. 200+/-28-mu-mol/min, P < 0.02), and in skeletal muscles; both when measured in femoral (1,594+-261 vs. 2,272+/-328-mu-mol/min, 25+/-13%) and arm muscles (1,617+/-411 to 2,305+/-517-mu-mol/min, P < 0.02, 31+/-6%). Whole body glucose uptake correlated with glucose uptake in femoral (r = 0.75, P < 0.005), and arm muscles (r = 0.69, P < 0.05) but not with glucose uptake in the heart (r = 0.04, NS). These data demonstrate that the glucose-FFA cycle operates in vivo in both heart and skeletal muscles in humans.