The relative contributions of the different Ca transport systems involved in cardiac relaxation were evaluated at 25 and 35 degrees C in isolated rabbit, ferret, and cat ventricular myocytes during twitches, caffeine-induced contractures in normal Tyrode solution, and caffeine-induced contractures in Na- and Ca-free solution. The time course of intracellular [Ca] decline these contractions in rabbit ventricular myocytes allowed estimates of the relative contributions of the sarcoplasmic reticulum (SR) Ca pump, Na/Ca exchange, sarcolemmal Ca pump, and the mitochondrial calcium uniporter (with the latter two considered together as "slow mechanisms"). The percent contributions of the SR Ca pump, the Na/Ca exchange, and the slow mechanisms were 70, 27 and 3% at 25 degrees C and 74, 23, and 3% at 35 degrees C. Warming from 25 to 35 degrees C decreases twitch contractions in rabbit and ferret myocytes and caffeine-induced contractures in normal Tyrode solution and Na- and Ca-free solution in all species. In contrast, in cat myocytes warming increased twitches, possibly because of a stronger effect of temperature on Ca influx. We conclude that increased temperature accelerates all of the Ca transport systems involved in relaxation. Despite large changes in each Ca transport system with warming, the relative contributions during relaxation remain similar at physiological temperature.