Effects of short-term, high-intensity and long-term, moderate-intensity exercise on biochemically assessed sarcoplasmic reticulum (SR) ATPase protein were analyzed in muscle homogenates of the rat after treadmill runs to exhaustion (avg, time to exhaustion 2 min 48 sec and 1 h 29 min, respectively) . The exercise-induced changes in SR Ca²⁺ -ATPase activity were muscle type-specific. After short-term exercise, a decrease in the activity occurred in the soleus muscle and the superficial region of the vastus lateralis muscle whereas long-term exercise depressed the rate of ATP hydrolysis in the soleus muscle and the deep region of the vastus lateralis muscle. The concentration of fluorescein isothiocyanate, a competitor at the ATP-binding site, for 50% inhibition of SR Ca²⁺ -ATPase activity fluctuated only in the soleus muscle subjected to short-term exercise ; it was increased by 31%. This change occurring in the soleus muscle would elevate SR Ca²⁺ -ATPase activity at a given concentration of ATP. The results presented here suggest that acute short-term exercise to exhaustion may exert a remarkably inhibitory factor on SR Ca²⁺ -ATPase protein of slow-twitch muscle, which can overcome the positive effect probably arising from the phosphorylation of the phospholamban.