Triplicate biopsies from three different depths of the gluteus medius muscle were obtained in 36 endurance-raced horses, aged 8.42 +/- 2.85 yr. Twenty of the horses were considered excellent endurance performers according to the mean speed of their three fastest records in endurance events for the past 2 or 3 years, whereas 16 were moderate performers, with a mean racing speed < 12.5 km/h (in 120- to 180-km endurance rides), < 14 km/h (in 80- to 120-km endurance rides), or < 13.5 km/h (in 40- to 60-km endurance rides). Significant differences in muscle fiber type composition and fiber size were recorded; excellent performers had a higher percentage and a larger size of type I and type IIa fibers (high and low myosin adenosinetriphosphatase activity at pH 4.5, respectively) and a lower percentage of type IIb fibers (moderate myosin adenosinetriphosphatase activity at pH 4.5), including both type IIb oxidative (moderate to high NADH-tetrazolium reductase activity) and IIb nonoxidative (low NADH-tetrazolium reductase activity). The differences in distribution of myofiber types and in fiber sizes were more marked in the deeper parts compared with the superficial regions of muscle. Our results also imply a greater homogeneity among the fiber type sizes across the muscle in horses with a superior endurance performance than in horses that had been poorly or moderately endurance raced. Thus the results show that fiber type proportions and fiber size in equine skeletal muscle are directly related to the athletic ability of the horse for endurance events.