Background: Racehorses commonly train and race in one direction, which may result in gait asymmetries. This study quantified gait symmetry in two cohorts of Thoroughbreds differing in their predominant exercising direction; we hypothesized that there would be significant differences in the direction of asymmetry between cohorts. Methods: 307 Thoroughbreds (156 from Singapore Turf Club (STC)—anticlockwise; 151 from Hong Kong Jockey Club (HKJC)—clockwise) were assessed during a straight-line, in-hand trot on firm ground with inertial sensors on their head and pelvis quantifying differences between the minima, maxima, upward movement amplitudes (MinDiff, MaxDiff, UpDiff), and hip hike (HHD). The presence of asymmetry (≥5 mm) was assessed for each variable. Chi-Squared tests identified differences in the number of horses with left/right-sided movement asymmetry between cohorts and mixed model analyses evaluated differences in the movement symmetry values. Results: HKJC had significantly more left forelimb asymmetrical horses (Head: MinDiff p < 0.0001, MaxDiff p < 0.03, UpDiff p < 0.01) than STC. Pelvis MinDiff (p = 0.010) and UpDiff (p = 0.021), and head MinDiff (p = 0.006) and UpDiff (p = 0.017) values were significantly different between cohorts; HKJC mean values indicated left fore- and hindlimb asymmetry, and STC mean values indicated right fore- and hindlimb asymmetry. Conclusion: the asymmetry differences between cohorts suggest that horses may adapt their gait to their racing direction, with kinematics reflecting reduced ‘outside’ fore- and hindlimb loading.