Five trained standardbred horses were studied trotting at two speeds (300 and 500 m × min-1) for 4 h and 1 h, respectively, and one horse at maximal speed. Their mean muscle glycogen content was reduced from 103 to 43 mmol × kg-1 in the 1-h experiment and from 119 to 48 mmol × kg-1 in the 4-h experiment. At maximal trotting speed, muscle glycogen declined from 88 to 46 mmol × kg-1 in 3 aggregate min of exercise. Glycogen depletion patterns in slow twitch (ST), fast twitch, high oxidative (FTH) and fast twitch (FT) fibres were estimated on the basis of the periodic acid-Schiff reaction. The ST and FTH fibres were depleted at all exercise intensities. These fibres then probably sustained almost all the exercise at slow trotting speeds as long as their glycogen stores were not depleted. After prolonged trotting and at high speeds, there appeared to be a progressive increase in FT fibre recruitment. Liver glycogen was reduced from 151 to 95 mmol × kg-1 in the 1-h run and from 116 to 19 mmol × kg-1 in the 4-h run, respectively. The fatigue found in horses trotting for long periods of time or at high speeds could be related to a selective depletion of glycogen in the ST and FTH fibres.