This study examined the effect of heat stress on physiological responses and exercise performance in elite road cyclists. Eleven members of the Australian National Road Cycling Squad completed two 30 min cycling time-trials in an environmental chamber set at either 32°C, (HT) or 23°C (NT) with a relative humidity of 60% in each circumstance. The trials were separated by two days, with six subjects performing HT first. Power output was 6.5% lower (P<0.05) during HT compared with NT. Mean skin temperature and sweat rate were higher (P<0.05) in HT compared with NT. In contrast, rectal temperature was remarkably similar throughout each trial. During the first 10 min of exercise in HT when power output was not different between trials, blood lactate was higher (P<0.05), and blood pH lower (P<0.05). In contrast, during the last 10 min of exercise when power output was reduced (P<0.05), blood lactate was lower (P<0.05), and pH higher (P<0.05), in HT. These data indicate that heat stress is associated with a reduced power output during self-paced exercise in highly trained men. This decrease in performance appears to be associated with factors associated with body temperature rather than metabolic capacity.