Although developmental differences in left ventricular function after cardioplegic arrest and rewarming have been postulated, whether differences exist at the level of the myocyte remains unexplored. This project tested the hypothesis that there is a differential effect of hypothermic hyperkalemic cardioplegic arrest with subsequent rewarming on contractile function of immature compared with adult ventricular myocytes. Myocytes were isolated from the left ventricular free wall of five immature and five adult rabbits and incubated for 2 hours in hyperkalemic modified Ringer's solution at 4° C (cardioplegia) or for 2 hours in cell culture medium at 37° C (normothermia). Myocytes were resuspended ('fewarmed') in 37° C cell culture medium after the incubation protocol. Normothermic baseline contractile performance was lower in immature, compared with adult, myocytes. Specifically, myocyte shortening velocity was 62 ± 4 μm/sec in immature and 112 ± 6 μm/sec in adult myocytes (p < 0.01). After cardioplegia and rewarming, immature myocyte contractile function was unchanged, whereas adult myocyte contractile function was significantly diminished. For example, myocyte shortening velocity was 65 ± 4 μm/sec in immature and 58 ± 3 μm/sec in adult myocytes (p < 0.01 versus normothermic). Myocyte surface area, which reflects myocyte volume, was increased after cardioplegia and rewarming in adults (3582 ± 55 versus 3316 ± 46 μm2, p < 0.01), but remained unchanged in immature myocytes (2212 ± 27 versus 2285 ± 28 μm2, p = not significant). These unique findings demonstrate a preservation of myocyte contractile function and volume regulation in immature myocytes after cardioplegic arrest and rewarming. Thus this study directly demonstrates that developmental differences exist in myocyte responses to hypothermic hyperkalemic cardioplegic arrest with subsequent rewarming.