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Our previous study has demonstrated that ovariectomy (Ovx) significantly increased the left ventricular developed pressure (LVDP) and the maximal rate of developed pressure over time ([+ or -]dP/d[t.sub.max]) in the isolated perfused rat heart and the effects were reversed by female sex hormone replacement. In the present investigation, we studied the effects of Ovx for 6 wk on [Ca.sup.2+] homeostasis that determines the contractile function. Particular emphasis was given to [Ca.sup.2+] handling by ryanodine receptor (RyR) and [Na.sup.+]-[Ca.sup.2+] exchange (NCX). [sup.45][Ca.sup.2+] fluxes via the RyR, NCX, and [Ca.sup.2+]-ATPase (SERCA) were compared with their expression in myocytes from Ovx rats with and without estrogen replacement. Furthermore, we correlated the handling of [Ca.sup.2+] by these [Ca.sup.2+] handling proteins with the overall [Ca.sup.2+] homeostasis by determining the [Ca.sup.2+] transients induced by electrical stimulation and caffeine, which reveals the dynamic changes of cytosolic [Ca.sup.2+] concentration ([[[Ca.sup.2+]].sub.i]) in the heart. In addition, we determined the expression and contribution of protein kinase A (PKA) to the regulation of the aforementioned [Ca.sup.2+] handling proteins in Ovx rats. It was found that after Ovx there were 1) increased [Ca.sup.2+] fluxes via RyR and NCX, which were reversed not only by estrogen replacement, but more importantly by blockade of PKA; 2) an increased expression of PKA; and 3) no increase in expression of NCX and SERCA. We suggest that hyperactivities of RyR and NCX are a result of upregulation of PKA. The increased release of [Ca.sup.2+] through RyR and removal of [Ca.sup.2+] by NCX are believed to be responsible for the greater contractility and faster relaxation after Ovx. ovariectomy doi: 10.1152/ajpcell.00368.2006.

We hypothesized that activation of heat shock protein 70 (HSP70) by preconditioning, which is known to confer delayed cardioprotection, attenuates the impaired handling of C[a.sup.2+] at multiple sites. To test the hypothesis, we determined how the ryanodine receptor (RYR), sarco(endo)plasmic reticulum C[a.sup.2+]-ATPase (SERCA), and N[a.sup.+]/C[a.sup.2+] exchanger (NCX) handled C[a.sup.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 C[a.sup.2+] transients induced by electrical stimulation and caffeine, which provide the overall picture of C[a.sup.2+] homeostasis; 2) expression of RyR, SERCA, and NCX; and 3) C[a.sup.2+] fluxes via NCX by the use of [45.sup.C[a.sup.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 C[a.sup.2+] from the sarcoplasmic reticulum via RyR and in the removal of C[a.sup.2+] from the cytoplasm by reuptake of C[a.sup.2+] to the SR via SERCA and by extrusion of C[a.sup.2+] out of the cell via NCX. UP also reduced mitochondrial C[a.sup.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 C[a.sup.2+] homeostasis at multiple sites in the heart, which may be responsible, at least partly, for attenuated C[a.sup.2+] overload, improved recovery in contractile function, and cardioprotection. intracellular C[a.sup.2+], [kappa]-opioid receptor; N[a.sup.+]/C[a.sup.2+] exchanger; ryanodine receptor; sarco(endo)plasmic reticulum C[a.sup.2+]-ATPase