To investigate the effects of intermittent heavy exercise (HE) on sarcoplasmic reticulum (SR) maximal Ca2+-ATPase activity (Vmax) and Ca2+ uptake, a continuous two-stage standardized cycling test was performed before and after HE by untrained men [peak aerobic power (Vo -->Vo2 peak) = 42.9 +/- 2.7 ml. kg-1 x min-1]. The HE consisted of 16 bouts of cycling performed for 6 min each hour at 90% Vo2 peak. Tissue was obtained from the vastus lateralis by needle biopsy before and during each cycle test. Before HE, reductions (P < 0.05 micromol. g protein-1x min-1) of 16 and 31% were observed in Vmax and Ca2+ uptake, respectively, after 40 min of the standardized test. Resting Vmax and Ca2+ uptake were depressed (P < 0.05) by 19 and 30%, respectively, when measured 36-48 h after HE. During the standardized test, after HE, Vmax increased (P < 0.05) by 20%, whereas no change was observed in Ca2+ uptake. The HE protocol resulted in small increases (P < 0.05) and decreases (P < 0.05) in sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) 2a and SERCA1 expression, respectively, as determined by Western blotting techniques. These results indicate that SR Ca2+-sequestering function in response to a prolonged exercise test depends on prior activity status, such that rested muscles exhibit a decrease and prior exercised muscles, an increase in Ca2+-ATPase activity. Moreover, it appears that changes in SERCA content can occur in response to a sustained session of intermittent exercise.