The effects of leg immobilization and retraining in combination with oral creatine intake, on muscle AMPK protein expression and phosphorylation status were investigated. A double-blind trial was performed in young healthy volunteers (n=22). A cast immobilized the right leg for 2 weeks, where after the knee-extensor muscles of that leg were retrained for 6 weeks Half of the subjects received creatine monohydrate throughout the study (CR; from 15g down to 2.5g daily), whilst the others ingested placebo (P; maltodextrin). Before and after immobilization and retraining needle biopsies were taken from the right and left vastus lateralis muscles. In the right leg of P and CR, immobilization did not affect AMPK ₁, ₂- and ₂-subunit expression or AMPK -subunit phosphorylation status. However, irrespective of the treatment received retraining increased the degree of -subunit phosphorylation by ~25% (p<0.05) and increased AMPK 1-subunit expression (p<0.05) in both groups. From the start to the end of the study, AMPK subunit protein expression and -subunit phosphorylation status were unchanged in the contralateral control leg. It is concluded that immobilization-induced muscle inactivity for 2 weeks does not alter AMPK ₁, ₂- and ₂ subunit expression or -AMPK phosphorylation status. Furthermore, the current observations indicate that AMPK probably is not implicated in the previously reported beneficial effects of oral creatine supplementation on muscle during immobilization and rehabilitative weight training.