The effect of several phenothiazines on the extent of cellular damage resulting from the calcium paradox was examined. Hearts treated with trifluoperazine, a potent calmodulin inhibitor, exhibited less cellular damage than untreated myocardium as reflected by light microscopy, high-energy phosphate content and the loss of protein and creatine phosphokinase into the perfusate. A dose response of this effect revealed a maximal response at about 1 microM trifluoperazine, a concentration which lies well within the range generally attributed to calmodulin inhibition. Several other lines of evidence were also obtained suggesting a possible role for calmodulin in calcium-overload induced necrosis. First, the phenothiazines had little influence on membrane changes believed responsible for altered calcium permeability. Second, trifluoperazine was without major effect unless included in the reperfusion buffer, indicating that the drug is only effective during the phase associated with calcium overload. Finally, less protection was afforded hearts exposed to phenothiazines such as chlorpromazine and promethazine, which are weaker inhibitors of calmodulin, than those treated with the potent inhibitor trifluoperazine. While other interpretations are possible, these studies are consistent with a role for calmodulin in calcium overload-induced heart failure.