The purpose of this study was to determine the optimal treadmill slope for trotters to produce the same heart rate and blood lactate responses as on the track during a standardized exercise test. Nine 2-year-old French trotters performed exercise tests on a training track and on a treadmill set at an incline of 0, 2 or 4%. For all horses, track testing was performed on day 1 and then on the treadmill according to a Latin-square design. The track test utilized three steps each of 3 min at speeds of 470, 530, 590 m/min and the same speeds were used on the treadmill. Derived physiological variables such as the speed at a HR of 200 bpm (V200) and the speed inducing blood lactate concentrations of 4 mmol/L (V4) were calculated. There were significant differences for V200and V4(P<0.05) between the track and the treadmill data when the treadmill was set at inclines of 0 and 4%, but no significant differences with the treadmill set at a 2% incline. The optimal treadmill incline to reproduce similar responses to those on the track was determined by regression analysis, and was found to be 2.4% for the two most often utilized derived physiological variables, V4and V200.