Protective effects of low and high doses of cyclosporin A against reoxygenation injury in isolated rat cardiomyocytes are associated with differential effects on mitochondrial calcium levels

Abstract In this study we aimed to determine the concentration range of cyclosporin A (CsA) which was effective in protecting against reoxygenation injury in isolated cardiomyocytes, and its effects on intramitochondrial free calcium levels ([Ca 2+ ] m ). We also determined whether a high [CsA] had any deleterious effect on normal myocyte function. Isolated adult rat ventricular myocytes were placed in a chamber on the stage of a fluorescence microscope for induction of hypoxia. [Ca 2+ ] m was determined from indo-1/am loaded cells where the cytosolic fluorescence signal had been quenched by superfusion with Mn 2+ . Cell length was measured using an edge-tracking device. Upon induction of hypoxia, control cells underwent rigor-contracture in 37± 1 min ( n =99) (T1); CsA had no effect on T1. The percentage of control cells which recovered upon reoxygenation depended on the time spent in rigor (T2). With a T2 of 21–30 min, only 36% of control cells recovered compared with 90% and 78% of cells treated with 0.2 μM and 1 μM CsA respectively. After 40 min in rigor, [Ca 2+ ] m was 280±60 nM in control-recovered cells (50% of cells) and 543±172 nM and 153±26 nM in cells treated with 0.2 and 1 μM CsA, respectively (all CsA treated cells recovered). In normoxic studies, CsA had no effect on cell contractility or [Ca 2+ ] m upon rapid pacing, even in presence of an elevated external [Ca 2+ ]. In conclusion, both low and high [CsA] protected against reoxygenation injury to cardiomyocytes despite having opposing effects on [Ca 2+ ] m , suggesting more than one mechanism of action. CsA had no effect on either cell contractility or [Ca 2+ ] m in normoxic cells.