1. To clarify the cause of uncoupling of Ca influx through Ca channels and the contractility of the myocardium in hibernating chipmunks, the electromechanical effects of two different internal Ca store inhibitors, caffeine and ryanodine, and a cardiotonic agent, isoprenaline, were investigated in papillary muscles of hibernating animals. 2. Ryanodine (10(-6) M), an inhibitor of internal Ca release, abolished the contraction with a marked inhibition of the action potential plateau (APp). In such preparations, an increase in Ca influx induced by isoprenaline (5 x 10(-8) M) failed to augment the contraction, indicating uncoupling of Ca influx and contraction. 3. In ryanodine pretreated preparations, 10 mM caffeine produced an early phase of APp, but did not affect the contraction abolished by ryanodine, while a higher concentration of caffeine (25 mM) markedly increased the contraction with an augmentation of the electrical response. 4. In the absence of ryanodine, caffeine (5 mM) almost abolished the contraction with a greater inhibition of APp. In such preparations, isoprenaline greatly increased the contraction with an augmentation of the early phase of APp. 5. These effects were not significantly affected by additional application of ryanodine, but were inhibited by nifedipine, a Ca channel blocker. 6. These observations suggest that in cardiac muscles of hibernating animals, lack of the positive inotropic effect of isoprenaline may be attributed to a rapid and effective sequestration of increased cytoplasmic Ca through Ca influx by internal stores, probably by enhancement of their ability to take up Ca.