Ca²⁺ paradox damage has been suggested to be determined by Na⁺ entry during Ca²⁺ depletion and exchange of Na⁺ for Ca²⁺ during Ca²⁺ repletion. With the use of ²³Na nuclear magnetic resonance, we previously observed a Ca²⁺ paradox without a prior Na⁺ increase. We have now demonstrated a Na⁺ increase during Ca²⁺ and Mg²⁺ depletion without the occurrence of the Ca²⁺ paradox during Ca²⁺ repletion. Isolated rat hearts were perfused for 20 min with a Ca²⁺-free or a Ca²⁺- and Mg²⁺-free (Ca²⁺/Mg²⁺-free) solution under hypothermic conditions (20 and 25°C). Intracellular Na⁺ concentration ([Na⁺]_i) increased from 11.9 ± 1.2 to 26.9 ± 5.8 mM (P < 0.001) during Ca²⁺/Mg²⁺-free perfusion at 20°C, whereas no significant change in [Na⁺]_i occurred during 20 min of Ca²⁺-free perfusion at 20°C. In addition, we confirmed that [Na⁺]_i did not change significantly during 20 min of normothermic Ca²⁺-free perfusion. Creatine kinase release during normothermic Ca²⁺ repletion in the 20°C groups was ~10% and in the 25°C groups 75% of the release in the normothermia group. Recovery of rate-pressure product was ~50% in the 20°C groups versus 0% in the normothermia group. In conclusion, hypothermic Ca²⁺/Mg²⁺-free perfusion results in a significant increase of [Na⁺]_i, which does not contribute to the extent of the Ca²⁺ paradox on normothermic Ca²⁺ repletion.