Reactive oxygen and nitrogen species are overproduced in the cardiovascular system in response to the exposure to doxorubicin, a cardiotoxic anticancer compound. Oxidant-induced cell injury involves the activation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) and pharmacological inhibition of PARP has recently been shown to improve myocardial contractility in doxorubicin-induced heart failure models. The current investigation, by utilizing an isolated perfused heart system capable of beat-to-beat intracellular calcium recording, addressed the following questions: (1) is intracellular calcium handling altered in hearts of rats after 6-week doxorubicin treatment, under baseline conditions, and in response to oxidative stress induced by hydrogen peroxide exposure in vitro; and (2) does pharmacological inhibition of PARP with the phenanthridinone-based PARP inhibitor PJ34 affect the changes in myocardial mechanical performance and calcium handling in doxorubicin-treated hearts under normal conditions and in response to oxidative stress. The results showed a marked elevation in intracellular calcium in the doxorubicin-treated hearts which was normalized by pharmacological inhibition of PARP. PARP inhibition also prevented the myocardial contractile disturbances and calcium overload that developed in response to hydrogen peroxide in the doxorubicin-treated hearts. We conclude that PARP activation contributes to the development of the disturbances in cellular calcium handling that develop in the myocardium in response to prolonged doxorubicin exposure.

In early stage, metabolic syndrome primarily disturbs SERCA2a function in the heart, but consequential haemodynamic dysfunction is prevented by upregulation of SERCA2a protein level and phosphorylation pathways regulating PLB. However, this compensated state is very vulnerable to a further decline in SERCA2a function.

The muscle Lim protein knock-out (MLP-KO) mouse model is extensively used for studying the pathophysiology of dilated cardiomyopathy. However, explanation is lacking for the observed long survival of the diseased mice which develop until adulthood despite the gene defect, which theoretically predestines them to early death due to heart failure. We hypothesized that adaptive changes of cardiac intracellular calcium (Ca(i)(2+)) handling might explain the phenomenon. In order to study the progression of changes in cardiac function and Ca(i)(2+) cycling, myocardial Ca(i)(2+)-transients recorded by Indo-1 surface fluorometry were assessed with concomitant measurement of hemodynamic performance in isolated Langendorff-perfused hearts of 3- and 9-month old MLP-KO animals. Hearts were challenged with beta-agonist isoproterenol and the sarcoplasmic reticular Ca(2+)-ATPase (SERCA2a) inhibitor cyclopiazonic acid (CPA). Cardiac mRNA content and levels of key Ca(2+) handling proteins were also measured. A decline in lusitropic function was observed in 3-month old, but not in 9-month old MLP-KO mice under unchallenged conditions. beta-adrenergic responses to isoproterenol were similar in all the studied groups. The CPA induced an increase in end-diastolic Ca(i)(2+)-level and a decrease in Ca(2+)-sequestration capacity in 3-month old MLP-KO mice compared to age-matched controls. This unfavorable condition was absent at 9 months of age. SERCA2a expression was lower in 3-month old MLP-KO than in the corresponding controls and in 9-month old MLP-KO hearts. Our results show time-related recovery of hemodynamic function and an age-dependent compensatory upregulation of Ca(i)(2+) handling in hearts of MLP-KO mice, which most likely involve the normalization of the expression of SERCA2a in the affected hearts.

The main aim of this study was to assess the kinetics of intracellular free calcium (Ca(2+)i) handling by isolated rat hearts rendered ischemic for 30 min followed by 30 min of reperfusion analyzing the upstroke and downslope of the Ca(2+)i transient. Changes in mechanical performance and degradation of membrane phospholipids--estimated by tissue arachidonic acid content--were correlated with Ca(2+)i levels of the heart. The fluorescence ratio technique was applied to estimate Ca(2+)i. The disappearance of mechanical activity of the heart preceded that of the Ca(2+)i transient in the first 2 min of ischemia. The slope of upstroke of the Ca(2+)i transient, reflecting Ca2+ release, decreased by 60%, while the duration of the downslope of the transient, reflecting Ca2+ sequestration, expressed a significant prolongation (105 +/- 17 vs. 149 +/- 39 msec) during the first 3 min of ischemia. At about 20 min of ischemia end-diastolic pressure expressed a 3.5-fold increase (contracture) when the fluorescence ratio showed a 2-fold elevation. Reperfusion was accompanied with a further precipitous increase in end-diastolic pressure, while resting Ca(2+)i remained at end-ischemic levels. Increases in the arachidonic acid (AA) content of the ischemic and postischemic hearts were proportional to Ca(2+)i levels. In summary, the present findings indicate that both calcium release and removal are hampered during the early phase of ischemia. Moreover, a critical level of Ca(2+)i and a critical duration of ischemia may exist to provoke contracture of the heart. Upon reperfusion the hearts show membrane phospholipid degradation and signs of stunning exemplified by elevated AA levels, partial recovery of Ca(2+)i handling and sustained depression of mechanical performance.

Heat shock (HS) pretreatment of the heart is effective in mitigating the deleterious effects of ischaemia/reperfusion. The main objective of this study was to determine whether the beneficial effect of HS is associated with the preservation of intracellular Ca2+ handling in the ischaemic/reperfused, isolated rat heart. Twenty-four hours after raising body core temperature to 42 degrees C for 15 min, rat hearts were perfused according to Langendorff and subjected to 30 min ischaemia followed by 20 min reperfusion. Cyclic changes of cytoplasmic calcium ion [Ca2+i] levels were measured by surface fluorometry using Indo-1 AM. Reperfused HS hearts showed improved recovery of contractile function compared with control hearts: end-diastolic pressure: 45+/-11 vs. 64+/-22 mmHg; developed pressure: 72+/-12 vs. 41+/-20 mmHg; maximum rate of pressure increase (+dP/dtmax): 1,513+/-305 vs. 938+/-500 mmHg/s; maximum rate of pressure decrease (-dP/dtmax): -1,354+/-304 vs. -806+/-403 mmHg/s. HS hearts displayed a significantly lower end-diastolic cytosolic [Ca2+] ([Ca2+]i) after reinstallation of flow. The dynamic parameters of the Ca2+i transients, i.e. the maximum rate of increase/decrease (+/-dCa2+i/dtmax) and amplitude, did not differ between reperfused control and HS hearts. The novel finding of this study is that improved performance of the HS-preconditioned heart after an ischaemic insult is associated with a reduced end-diastolic Ca2+i load, and most likely, preserved Ca2+ sensitivity of the myocardial contractile machinery.

No abstract

The endothelium‐mediated vasomotor function of skeletal muscle arterioles has been shown to be impaired in HHcy. Thus we hypothesized that HHcy modulates the reactive hyperemic response of coronary circulation following occlusion. Rats received 1g/kg of body weight methionine daily, for 5 weeks in the drinking water to induce mild HHcy (~ 30 μmol vs. ~ 7 μmol). In Langendorff‐perfused hearts reactive hyperemia (RH) after 5 min of global ischemia was studied prior to and after the simultaneous administration of the nitric oxide synthase inhibitor L‐NAME (10−4M) and the cyclooxygenase inhibitor indomethacin (INDO 10−5M) into the perfusate. We have found that the early peak values of reactive hyperemia were not significantly different in HHcy and control (C) groups (at 2 min of RH: HHcy: 139±14 vs. C: 147±19% of basal flow). The later phase of reactive hyperemia however, was significantly diminished in HHcy hearts (at 8 min of RH: HHcy: 116±7 vs. C: 130±14% of basal flow). Presence of L‐NAME+INDO eliminated the difference between the reactive hyperemia of control and HHcy hearts. On the basis of these findings we propose that cardiac reactive hyperemia is impaired in HHcy, primarily due to reduced release of endothelium‐derived dilator factors. (Grants: Hungarian Sci. Res. Funds/OTKA T48376, 61694, 68502, T67984; and Am. Heart Assoc. NE Aff. 0555897T, USA).