INTRODUCTION:: The hypothesis that brief intermittent exercise induced increases in human skeletal muscle metabolic mRNA is dependent on relative workload was investigated. METHODS:: Trained (n=10) and untrained (n=8) subjects performed exhaustive intermittent cycling exercise (4x4 min @ 85% of VO2 peak, interspersed by 3 min). Trained subjects also performed the intermittent exercise at the same absolute workload as untrained, corresponding to 70% of VO2 peak (n=6). RESULTS:: Exercise at 85% of VO2 peak elevated (P<0.001) venous plasma lactate to 10.1+/-0.4 mM and 10.8+/-0.5 mM in trained and untrained, respectively. PGC-1alpha mRNA expression was increased (P<0.001) ~4-5 fold for several hours after exercise in both groups. After exercise at 70% of VO2 peak, venous plasma lactate was less (P<0.001) elevated (3.1+/-0.7 mM) and PGC-1alpha mRNA content was less (P<0.05) increased (~3 fold) than after exercise at 85% of VO2 peak. Likewise, PDK4 and HKII mRNA expression were only increased (P<0.05) subsequent to exercise performed at 85% of VO2 peak in trained subjects. HIF2alpha mRNA only increased (P<0.05) 3 h into recovery in trained, with no difference between the 70% and 85% of VO2 peak trial. No change in HIF1alpha, PFK, CS, LDH-A or LDH-B mRNA expression was detected after any of the exercise trials. CONCLUSION:: The relative intensity of brief intermittent exercise is of major importance for the exercise induced increase of several mRNA's, including PGC-1alpha.