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

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 ([Ca2+](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, [Ca2+](m) was determined from indo-1/am loaded cells where the cytosolic fluorescence signal had been quenched by superfusion with Mn2+. 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 mu M and 1 mu M CsA respectively. After 40 min in rigor, [Ca2+](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 mu M CsA, respectively (all CsA treated cells recovered). in normoxic studies, CsA had no effect on cell contractility or [Ca2+](m) upon rapid pacing, even in presence of an elevated external [Ca2+]. In conclusion, both low and high [CsA] protected against reoxygenation injury to cardiomyocytes despite having opposing effects on [Ca2+](m), suggesting more than one mechanism of action. CsA had no effect on either cell contractility or [Ca2+](m) in normoxic cells. (C) 2000 Harcourt Publishers Ltd.