Abstract 71: Reperfusion-Induced Hypercontracture of Cardiomyocytes is Secondary to SR-Driven Cytosolic Ca2+ Oscillations and Mitochondrial Permeability Transition
Introduction: We showed previously that reperfusion causes hypercontracture and necrosis of cardiomyocytes due to oscillatory elevations of cytosolic Ca2+ originating from the sarcoplasmic reticulum (SR). It has also been shown that mitochondrial permeability transition (MPT) induces cell death in reperfused myocardium. We now investigated if SR-triggered Ca2+ oscillations and MPT are causally connected in signalling towards reperfusion-induced cardiomyocyte hyper-contracture.
Methods: To simulate ischemia and reperfusion, isolated cardiac myocytes from adult rat were superfused anoxically (pH 6.4) and then reperfused with a normoxic puffer (pH 7.4). Induction of MPT was detected in whole cells by the calcein loading procedure. Cytosolic Ca2+ was measured by use of fura-2.
Results: During simulated ischemia, no significant change in calcein fluorescence was observed. Reperfusion induced a marked decline in calcein fluorescence, which reflects induction of MPT. Concomitantly, reperfused cells developed hypercontracture. Application of the MPT inhibitor cyclosporine (500 nM) or inhibition of SR-Ca2+ release with ryanodine (3 μM) reduced the decline in calcein fluorescence and hypercontracture (calcein fluorescence as % of end-anoxic value after 5 min reperfusion: control: 42.2±2.4; cyclosporine: 90.5±1.8*; ryanodine: 86.8±1.5*; n=20; *p<0.05 vs. control; cell length as % of end-anoxic after 5 min reperfusion; control: 46.8±1.6; cyclosporine: 88.0±1.8*; ryanodine: 94.8±2.0*; n=20; *p<0.05).
Conclusions: Induction of MPT is involved in the development of reperfusion-induced hypercontracture of cardiac myocytes. It is triggered by oscillations of cytosolic Ca2+ originating from the SR during the early phase of reperfusion.