Cytosolic Ca²⁺ concentration of rat ventricular cells was measured under varying experimental conditions by using a fluorescent Ca²⁺ indicator, Fura-2. Resting [Ca²⁺]_i of rat myocyte was 150 ± 30 nM (n = 39), and this value was compatible with others. The Perfusion of cardioplegic solution significantly increased [Ca²⁺]_i, and this effect was further augmented by hypothermia (p<0.05). Application of nifedipine (5 x 10^-7 M) to the perfusate or pretreatment of caffeine (10 mM) had no apparent effect on this cardioplegia-induced [Ca²⁺]_i change. But Ni²⁺ (5 mM), an antagonist of Na⁺/Ca²⁺ exchange mechanism, prevented the [Ca²⁺]_i change during cardioplegia (p<0.05). Magnitude of cardioplegia-induced [Ca²⁺]_i increase was also dependent on the Ca²⁺ concentration of cardioplegic solution. These results suggest that Na⁺/Ca²⁺ exchange may play an important role in cardioplegia-induced [Ca²⁺]_i change. To rule out the possibility whether the protective effect of hypothermic cardioplegia is due to the preservation of high-energy phosphate store or decreasing the transmembrane ionic fluxes by phase transition, we exhausted a energy store of cardiac cell by application of 2,4 dinitrophenol to the bath and measured its effect on [Ca²⁺]_i change during cardioplegia. Hypothermic cardioplegia delayed the onset of [Ca²⁺]_i increase and decreased its amplitude compared to those of normothermic cardioplegia. From the above results, hypothermic cardioplegia may protect the cardiac cells from ischemic insult by preserving a high-energy phosphate store. Application of Ni²⁺ to the cardioplegic solution or reduction of external Ca²⁺ concentration also had some protective effect, since it prevented [Ca²⁺]_i increase during cardioplegia.