Six subjects performed isometric contraction (66% maximal force) to fatigue with the knee extensor muscles. Biopsies were taken from the quadriceps femoris muscle at rest, at fatigue and 1 min after termination of contraction. In three of the subjects recovery from contraction occurred in the presence of an intact circulation (non-occluded, NON) to the thigh, whereas in the other three the circulation during recovery was occluded (OCC). Glycogen synthase fractional activity (GSF) decreased in all subjects from (mean +/- SE) 0.53 +/- 0.06 at rest to 0.37 +/- 0.04 at fatigue (P < 0.001). In the OCC group GSF returned to the pre-exercise value within 1 min after termination of contraction (0.59 +/- 0.07 at rest vs. 0.57 +/- 0.04 at 1 min post-exercise), whereas in the NON group GSF increased to a higher extent (0.48 +/- 0.09 at rest vs. 0.70 +/- 0.06 at 1 min post-exercise). The increase in GSF during the 1-min recovery was almost three-fold higher in the NON group (0.15 +/- 0.02 vs. 0.38 +/- 0.03). Cyclic AMP-dependent protein kinase (cAMP-PK) (assayed at 0/100 microM and 0.2/100 microM cAMP) did not change at fatigue or during recovery in either group. Glycogen synthase phosphatase (GSP) increased at fatigue by approximately 30% (P < 0.05 vs. rest). It is concluded that isometric contraction mediated inactivation of GS (i.e. phosphorylation of GS) is due to activation of a protein kinase(s) but not cAMP-PK. The rapid activation of GS in the NON group demonstrates that a humoral factor(s), possibly insulin and/or oxygen, is responsible for this phenomenon.