The purpose of this study was to investigate the effects of sustained stretch and two common proprioceptive neuromuscular facilitation (PNF) stretch techniques on hamstring muscle activation and knee extension range of motion (ROM) in different athletic populations. Three stretch techniques: stretch-relax (SR), contract-relax (CR), and agonist contract-relax (ACR) were applied to 10 endurance athletes (EN), 10 high intensity athletes (HI), and 10 control subjects (C). The results revealed that ACR produced 89-110% greater hamstring EMG activity (P less than 0.05) and 9-13% more knee joint ROM than CR and SR, respectively. This same pattern was evident for the individual subject groups. Comparisons of mean data among the three subject groups revealed that the EN athletes generated 58-113% more hamstring EMG activity (P less than 0.05) than the HI and C groups, respectively, across all stretch conditions, whereas the EN group attained significantly less ROM than the HI and C groups for CR and ACR conditions. It was postulated that high intensity-short term activity training necessitates less hamstring resistance to knee extension than long term endurance training. The findings suggest that decreases in muscle activity may not be strongly related to increases in joint range of motion and that factors other than muscle relaxation are important in achieving increased ROM. Also, the differential effects of various stretch techniques between dissimilar athletic populations should be considered if stretch-induced injury is to be avoided.