Over 20 years ago, successful 3-day-perfusion preservation of canine kidneys was obtained. Since then, consistent 5-day preservation has not been reported. In this study, we investigated how the perfusate calcium concentration affected both mitochondrial function and posttransplant viability in dog kidneys preserved for 5 days. Dog kidneys were preserved by machine perfusion (5 degrees C) using a hydroxyethyl starch-gluconate solution that contained either 0.0, 0.5, 1.5, or 5.0 mM calcium. Mitochondria isolated from preserved kidneys has a loss of respiratory control when either 0.0, 1.5, or 5.0 mM calcium were present. However, the use of a perfusate with 0.5 mM calcium preserved the mitochondrial function at levels equivalent to controls for 5 days. Transplantation of kidneys preserved for 5 days with 0.0 or 1.5 mM calcium yielded poor survival (0% and 17%, respectively). The use of a 0.5-mM calcium perfusate increased posttransplant survival to 63% (5 of 8 transplanted). Donor pretreatment of kidneys with chlorpromazine (2.5 mg/kg i.v.) did not improve the function of mitochondria isolated from preserved kidneys but did increase survival in the 1.5-mM calcium group to 67% (4 of 6 transplanted) and in the 0.5 mM calcium group to 100% (7 of 7 transplanted). This is the first report to document consistently successful 5-day preservation of canine kidneys and clearly shows the importance of the perfusate calcium concentration in long-term kidney preservation. The specific mechanism by which calcium or chlorpromazine exert their effect is not known, but it is apparent that excessively high or low concentrations of calcium are damaging to the preserved organ, and an optimal calcium concentration combined with metabolic inhibition of calcium-dependent pathways can significantly improve the function of organs preserved for extended time periods.