The effect of phenothiazines upon maintenance of membrane integrity in the cultured myocardial cell

The cultured myocardial cell provides a defined model for examining factors which are responsible for maintaining cellular viability and sarcolemmal integrity. Our data indicates that the spontaneous loss of myocyte membrane integrity is a calcium-dependent process and thus provides a method for examining the mechanism through which calcium exerts this effect. Antimyosin antibody staining and propidium iodide uptake were used to quantitate membrane integrity. The integrity of the cell membrane was inversely related to the calcium concentration in the culture medium. This loss of membrane integrity was calmodulin-dependent as demonstrated by the following: (1) phenothiazines (trifluoperazine >chlorpromazine>promethazine) and structurally dissimilar calmodium-inhibitors prevented the formation of sarcolemmal defects at concentrations similar to those known to inhibit calmodulin; (2) phenothiazines and calcium demonstrated a competitive interaction with respect to this effect on membrane integrity. Electron microscopy confirmed the integrity of the sarcolemma of the cells exposed to high phenothiazine concentrations although metabolic alterations occurred in these cells as evidenced by an increased membrane permeability to the low molecular weight probe propidium iodide, degenerative changes in the fine structure of the mitochondria, the accumulation of autophagic vacuoles in the cytoplasm and the loss of contractile ability. These findings indicate that calmodulin inhibitory compounds are capable of preserving the membrane integrity of cardiac myocytes, interfering with a calcium-dependent process that is associated with the spontaneous attrition of these cells in culture. Significant intracellular alterations appear at high doses of these agents even while the sarcolemma is free of gross defects.