Glycogen depletion was used as an experimental tool to examine the relationship between excess post-exercise oxygen consumption and lactate metabolism in 6-g rainbow trout. A 5-day starvation period reduced whole-body glycogen stores by 50% and slightly lowered resting lactate levels; resting oxygen consumption, glucose, ATP, and creatine phosphate levels were not affected. After a 5-min bout of exhaustive exercise, significantly less glycogen was utilized by the glycogen-depleted fish, 40% less lactate was accumulated, and glucose levels did not rise in comparison with the control group. Creatine phosphate recovered more quickly in the glycogen-depleted fish, whereas ATP was unaffected. Recovery from excess post-exercise oxygen consumption was not significantly different despite the large absolute differences in lactate removed and glycogen resynthesized. This experimental test demonstrates that the classical oxygen debt hypothesis does not completely explain the excess post-exercise oxygen consumption in the trout.