Muscle contractions induce an increase in glucose transport. The acute effect of muscle contractions on glucose transport is independent of insulin and reverses rapidly after cessation of exercise. As the acute increase in glucose transport reverses, a marked increase in the sensitivity of muscle to insulin occurs. The mechanism for this phenomenon is unknown. We hypothesize that an increase in insulin sensitivity is a general phenomenon that occurs during reversal of an increase in cell surface GLUT4 induced by any stimulus, not just exercise. To test this hypothesis, epitrochlearis, rat soleus, and flexor digitorum brevis muscles were incubated for 30 min with a maximally effective insulin concentration (1.0 mU/ml). Muscles were allowed to recover for 3 h in the absence of insulin. Muscles were then exposed to 60 microU/ml insulin for 30 min followed by measurement of glucose transport. Preincubation with 1.0 mU/ml insulin resulted in an approximately 2-fold greater increase in glucose transport 3.5 h later in response to 60 microU/ml insulin than that which occurred in control muscles treated with 60 microU/ml insulin. Pretreatment of muscles with combined maximal insulin and exercise stimuli greatly amplified the increase in insulin sensitivity. The increases in glucose transport were paralleled by increases in cell surface GLUT4. We conclude that stimulation of glucose transport by any agent is followed by an increase in sensitivity of glucose transport to activation that is mediated by translocation of more GLUT4 to the cell surface.