Further study on the role of HSP70 on Ca super(2+) homeostasis in rat ventricular myocytes subjected to simulated ischemia

We hypothesized that activation of heat shock protein 70 (HSP70) by preconditioning, which is known to confer delayed cardioprotection, attenuates the impaired handling of Ca super(2+) at multiple sites. To test the hypothesis, we determined how the ryanodine receptor (RyR), sarco(endo)plasmic reticulum Ca super(2+)-ATPase (SERCA), and Na super(+)/Ca super(2+) exchanger (NCX) handled Ca super(2+) in rat ventricular myocytes preconditioned with a Kappa -opioid receptor agonist, U50488H (UP), followed by blockade of HSP70 with a selective antisense oligonucleotide and subsequently subjected to simulated ischemia. We determined the following: 1) the Ca super(2+) transients induced by electrical stimulation and caffeine, which provide the overall picture of Ca super(2+) homeostasis; 2) expression of RyR, SERCA, and NCX; and 3) Ca super(2+) fluxes via NCX by the use of super(45)Ca super(2+) in the rat ventricular myocyte. We found that UP increased the activity of RyR, SERCA, and NCX and the expression of RyR and SERCA. These effects led to increases in the release of Ca super(2+) from the sarcoplasmic reticulum via RyR and in the removal of Ca super(2+) from the cytoplasm by reuptake of Ca super(2+) to the SR via SERCA and by extrusion of Ca super(2+) out of the cell via NCX. UP also reduced mitochondrial Ca super(2+) accumulation. All of the effects of UP were either abolished or significantly attenuated by blockade of HSP70 synthesis with a selective antisense oligonucleotide. The results are evidence that activation of HSP70 by preconditioning improves the ischemia-impaired Ca super(2+) homeostasis at multiple sites in the heart, which may be responsible, at least partly, for attenuated Ca super(2+) overload, improved recovery in contractile function, and cardioprotection.