This study tested the hypothesis that paraxanthine, a caffeine metabolite, stimulates skeletal muscle potassium (K+) transport by an increase in Na+-K+ ATPase activity. The unidirectional transport of K+ into muscle (JinK) was studied using a perfused rat hind limb technique. Using 12 hind limbs, we examined the response to 20 min of paraxanthine perfusion (0.1 mM), followed by 20 min perfusion with 0.1 mM paraxanthine and 5 mM ouabain (n = 5) to irreversibly inhibit Na+-K+ ATPase activity. Paraxanthine stimulated JinK by 23 ± 5% within 20 min. Ouabain abolished the paraxanthine-induced stimulation of JinK, suggesting the increase in K+ uptake was due to activation of the Na+-K+ ATPase. To confirm the role of the Na+-K+ ATPase, 14 hind limbs were perfused for 20 min with 5 mM ouabain prior to 20 min perfusion with 0.1 mM paraxanthine and 5 mM ouabain (n = 6). Ouabain alone resulted in a 41 ± 7% decrease in JinK within 15 min. Inhibition of ouabain-sensitive JinK prevented the paraxanthine-induced increase in JinK. Hind limbs (n = 3) were also perfused with 0.1 mM paraxanthine for 60 min to examine the response to longer duration paraxanthine perfusion. The paraxanthine-induced increase in JinK continued for the entire 60 min. In another series, hind limbs were perfused with 0.01 (n = 9), 0.1 (n = 9), or 0.5 (n = 6) mM paraxanthine for 15 min. There was no concentration-dependent relationship between JinK and paraxanthine concentration, and 0.01, 0.1, and 0.5 mM paraxanthine increased JinK similarly (25 ± 5, 22 ± 4, and 27 ± 6%, respectively). The effect of paraxanthine on JinK could not be reversed by subsequent perfusion with paraxanthine-free perfusate. Caffeine (0.05-1.0 mM) had no effect on K+ transport. It is concluded that paraxanthine increases JinK in resting skeletal muscle by stimulating ouabain-sensitive Na+-K+ ATPase activity.Key words: caffeine, methylxanthine, ouabain, potassium transport, sodium pump, Na-K ATPase, VO2, glycolysis.