Journal of Molecular and Cellular Cardiology

Editorial Board

2012-06-01

Differentiation and enrichment of cardiomyocytes from human pluripotent stem cells

Chunhui Xu — 2012-04-05

Abstract: Human cardiomyocytes derived from pluripotent stem cells hold great promise for cardiac cell therapy, disease modeling, drug discovery, and the study of developmental biology. Reaching these potentials fully requires the development of methods that enable efficient and robust generation of cardiomyocytes with expected characteristics. This review summarizes and discusses up-to-date methods that have been used to derive and enrich human cardiomyocytes from pluripotent stem cells, provides a brief overview of in vitro and in vivo characterization of these cardiomyocytes, and considers future advancement needed to further harness the power of these cells.Highlights: ► Described methods to derive and enrich cardiomyocytes from pluripotent stem cells. ► Discussed advantages and limitations of individual methods. ► Presented brief overview of in vitro and in vivo characterization of cardiomyocytes. ► Discussed future advancement needed to further harness the power of these cells.

Doxorubicin-induced cardiomyopathy: From molecular mechanisms to therapeutic strategies

2012-03-22

Abstract: The utility of anthracycline antineoplastic agents in the clinic is compromised by the risk of cardiotoxicity. It has been calculated that approximately 10% of patients treated with doxorubicin or its derivatives will develop cardiac complications up to 10years after the cessation of chemotherapy. Oxidative stress has been established as the primary cause of cardiotoxicity. However, interventions reducing oxidative stress have not been successful at reducing the incidence of cardiotoxicity in patients treated with doxorubicin. New insights into the cardiomyocyte response to oxidative stress demonstrate that underlying differences between in vitro and in vivo toxicities may modulate the response to superoxide radicals and related compounds. This has led to potentially new uses for pre-existing drugs and new avenues of exploration to find better pharmacotherapies and interventions for the prevention of cardiotoxicity. However, much work still must be done to validate the clinical utility of these new approaches and proposed mechanisms. In this review, the authors have reviewed the molecular mechanisms of the pathogenesis of acute and chronic doxorubicin-induced cardiotoxicity and propose potential pharmacological interventions and treatment options to prevent or reverse this specific type of heart failure.Highlights: ► The pathogenesis of doxorubicin-induced cardiotoxicity is complex and multifactorial. ► Oxidative stress is a major cause of doxorubicin-induced cardiotoxicity. ► Doxorubicin-induced heart failure can appear very late after the last administration. ► There is no specific curative or preventive treatment available.

Notch signaling and cardiac repair

Natalie Gude, Mark Sussman — 2012-03-23

Abstract: Notch signaling is critical for proper heart development and recently has been reported to participate in adult cardiac repair. Notch resides at the cell surface as a single pass transmembrane receptor, transits through the cytoplasm following activation, and acts as a transcription factor upon entering the nucleus. This dynamic and widespread cellular distribution allows for potential interactions with many signaling and binding partners. Notch displays temporal as well as spatial versatility, acting as a strong developmental signal, controlling cell fate determination and lineage commitment, and playing a pivotal role in embryonic and adult stem cell proliferation and differentiation. This review serves as an update of recent literature addressing Notch signaling in the heart, with attention to findings from noncardiac research that provide clues for further interpretation of how the Notch pathway influences cardiac biology. Specific areas of focus include Notch signaling in adult myocardium following pathologic injury, the role of Notch in cardiac progenitor cells with respect to differentiation and cardiac repair, crosstalk between Notch and other cardiac signaling pathways, and emerging aspects of noncanonical Notch signaling in heart.Highlights: ► Activation of Notch promotes myocardial repair. ► Notch signaling controls cardiac progenitor specification and differentiation. ► Notch crosstalks with survival pathways in heart. ► Noncanonical Notch signaling may participate in cardiac repair.

Ca2+-activated adenylyl cyclase 1 introduces Ca2+-dependence to beta-adrenergic stimulation of HCN2 current

Yelena N. Kryukova, Lev Protas, Richard B. Robinson — 2012-03-30

Abstract: Previous observations show that β-adrenergic modulation of pacemaker current (If) in sinoatrial node (SAN) cells is impaired by disruption of normal Ca2+-homeostasis with ryanodine or BAPTA. Recently, the presence of Ca2+-activated adenylyl cyclase (AC) 1 was reported in SAN, and was proposed as a possible mechanism of Ca2+-dependence of β-adrenergic modulation. However, direct evidence that pacemaker (HCN) channels can be regulated by Ca2+-activated AC and that such regulation introduces Ca2+ dependence, is lacking. Here we co-expressed AC1 or AC6 with HCN2 in neonatal rat ventricular myocytes, which lack AC1. Although both isoforms have equivalent expression level and ability to interact with HCN2, only AC1 increases intracellular cAMP content, accelerates spontaneous beating rate and modifies HCN2 biophysics. Measured HCN2 current in the AC1 group activated ~10mV more positive than in GFP or AC6. The β-adrenergic agonist isoproterenol induced a further positive shift under control conditions, but failed to do so after pretreatment with the Ca2+ chelator BAPTA. In the AC6 group, isoproterenol shifted the HCN2 activation relation to a similar extent in the absence and presence of BAPTA. Thus, AC1 but not AC6 over-expression introduces Ca2+-sensitivity to the β-adrenergic response of HCN2. These results demonstrate physical and functional interaction between AC isoforms and the HCN2 pacemaker channel and support a key role of Ca2+ activated AC1 as a molecular mechanism in Ca2+-dependent modulation of β-adrenergic response of heart rate.Highlights: ► HCN2 was co-expressed with Adenylyl Cylase 1 (AC1) or 6 (AC6) in cardiac myocytes. ► AC1 but not AC6 increased [cAMP]i, accelerated beating rate and shifted HCN2 positive. ► In both AC1 and AC6 groups, isoproterenol shifted HCN2 activation relation positive. ► BAPTA prevented the isoproterenol effect in the AC1 but not AC6 group. ► AC1 plays a key role in sensitivity of β-adrenergic stimulation of HCN to [Ca]i.

Ankyrin-B reduction enhances Ca spark-mediated SR Ca release promoting cardiac myocyte arrhythmic activity

Emmanuel Camors, Peter J. Mohler, Donald M. Bers, Sanda Despa — 2012-03-05

Abstract: Ankyrin-B (AnkB) loss-of-function may cause ventricular arrhythmias and sudden cardiac death in humans. Cardiac myocytes from AnkB heterozygous mice (AnkB+/−) show reduced expression and altered localization of Na/Ca exchanger (NCX) and Na/K-ATPase (NKA), key players in regulating [Na]i and [Ca]i. Here we investigate how AnkB reduction affects cardiac [Na]i, [Ca]i and SR Ca release. We found reduced NCX and NKA transport function but unaltered [Na]i and diastolic [Ca]i in myocytes from AnkB+/− vs. wild-type (WT) mice. Ca transients, SR Ca content and fractional SR Ca release were larger in AnkB+/− myocytes. The frequency of spontaneous, diastolic Ca sparks (CaSpF) was significantly higher in intact myocytes from AnkB+/− vs. WT myocytes (with and without isoproterenol), even when normalized for SR Ca load. However, total ryanodine receptor (RyR)-mediated SR Ca leak (tetracaine-sensitive) was not different between groups. Thus, in AnkB+/− mice SR Ca leak is biased towards more Ca sparks (vs. smaller release events), suggesting more coordinated openings of RyRs in a cluster. This is due to local cytosolic RyR regulation, rather than intrinsic RyR differences, since CaSpF was similar in saponin-permeabilized myocytes from WT and AnkB+/− mice. The more coordinated RyRs openings resulted in an increased propensity of pro-arrhythmic Ca waves in AnkB+/− myocytes. In conclusion, AnkB reduction alters cardiac Na and Ca transport and enhances the coupled RyR openings, resulting in more frequent Ca sparks and waves although the total SR Ca leak is unaffected. This could enhance the propensity for triggered arrhythmias in AnkB+/− mice.Highlights: ► Reduced Na and Ca transport but unaltered [Na]i and diastolic [Ca]i in AnkB+/− myocytes. ► Ca transients, SR Ca load and fractional SR Ca release are larger in AnkB+/− myocytes. ► Higher Ca spark frequency and unaltered total SR Ca leak in intact AnkB+/− myocytes. ► The bias towards Ca spark-mediated leak is due to different cytosolic RyR regulation. ► More coordinated RyR openings increase the propensity for Ca waves in AnkB+/− mice.

Excitation–contraction coupling in ventricular myocytes is enhanced by paracrine signaling from mesenchymal stem cells

2012-03-26

Abstract: In clinical trials mesenchymal stem cells (MSCs) are transplanted into cardiac ischemic regions to decrease infarct size and improve contractility. However, the mechanism and time course of MSC-mediated cardioprotection are incompletely understood. We tested the hypothesis that paracrine signaling by MSCs promotes changes in cardiac excitation–contraction (EC) coupling that protects myocytes from cell death and enhances contractility. Isolated mouse ventricular myocytes (VMs) were treated with control tyrode, MSC conditioned-tyrode (ConT) or co-cultured with MSCs. The Ca handling properties of VMs were monitored by laser scanning confocal microscopy and whole cell voltage clamp. ConT superfusion of VMs resulted in a time dependent increase of the Ca transient amplitude (ConT15min: ΔF/F0=3.52±0.38, n=14; Ctrl15min: ΔF/F0=2.41±0.35, n=14) and acceleration of the Ca transient decay (τ: ConT: 269±18ms n=14; vs. Ctrl: 315±57ms, n=14). Voltage clamp recordings confirmed a ConT induced increase in ICa,L (ConT: −5.9±0.5 pA/pF n=11; vs. Ctrl: −4.04±0.3 pA/pF, n=12). The change of τ resulted from increased SERCA activity. Changes in the Ca transient amplitude and τ were prevented by the PI3K inhibitors Wortmannin (100nmol/L) and LY294002 (10μmol/L) and the Akt inhibitor V (20μmol/L) indicating regulation through PI3K signal transduction and Akt activation which was confirmed by western blotting. A change in τ was also prevented in eNOS−/− myocytes or by inhibition of eNOS suggesting an NO mediated regulation of SERCA activity. Since paracrine signaling further resulted in increased survival of VMs we propose that the Akt induced change in Ca signaling is also a mechanism by which MSCs mediate an anti-apoptotic effect.Highlights: ► Bone marrow derived mesenchymal stem cells enhance cardiomyocyte E-CC. ► Changes in E-CC and survival are mediated by paracrine signaling. ► MSCs activate the PI3K/Akt pathway and eNOS dependent NO production. ► MSCs increase Ca transient amplitude by increasing ICa,L and SERCA activity. ► Increased SERCA activity depends on eNOS activity.

Reduction of heart failure by pharmacological inhibition or gene deletion of protein tyrosine phosphatase 1B

2012-03-19

Abstract: Protein tyrosine phosphatase 1B (PTP1B) regulates tyrosine kinase receptor-mediated responses, and especially negatively influences insulin sensitivity, thus PTP1B inhibitors (PTP1Bi) are currently evaluated in the context of diabetes. We recently revealed another important target for PTP1Bi, consisting in endothelial protection. The present study was designed to test whether reduction of PTP1B activity may be beneficial in chronic heart failure (CHF).We evaluated the impact of either a 2month pharmacological inhibition, or a gene deletion of PTP1B (PTP1B−/−) in CHF mice (2months post-myocardial infarction). PTP1Bi and PTP1B deficiency reduced adverse LV remodeling, and improved LV function, as shown by the increased LV fractional shortening and cardiac output (measured by echocardiography), the increased LV end systolic pressure, and the decreased LV end diastolic pressure, at identical infarct sizes. This was accompanied by reduced cardiac fibrosis, myocyte hypertrophy and cardiac expression of ANP. In vitro vascular studies performed in small mesenteric artery segments showed a restored endothelial function (i.e. improved NO-dependent, flow-mediated dilatation, increased eNOS phosphorylation) after either pharmacological inhibition or gene deletion. PTP1B−/− CHF also displayed an improved insulin sensitivity (assessed by euglycemic–hyperinsulinemic clamp studies), when compared to wild-type CHF associated with an increased insulin mediated mesenteric artery dilation.Thus, chronic pharmacological inhibition or gene deletion of PTP1B improves cardiac dysfunction and cardiac remodeling in the absence of changes in infarct size. Thus this enzyme may be a new therapeutic target in CHF. Diabetic patients with cardiac complications may potentially benefit from PTP1B inhibition via two different mechanisms, reduced diabetic complications, and reduced heart failure.Highlights: ► Protein tyrosine phosphatase 1B inhibition (PTP1Bi) was assessed in heart failure. ► PTP1Bi or gene deficiency reduced adverse LV remodeling, and improved LV function. ► PTP1Bi of gene deficiency reduced cardiac fibrosis and hypertrophy. ► PTP1Bi or gene deficiency restored endothelial function. ► PTP1B may be a new therapeutic target in CHF.

Uterine cells are recruited to the infarcted heart and improve cardiac outcomes in female rats

2012-03-16

Abstract: We evaluated the hypothesis that uterine cells home to the heart after injury and improve cardiac outcomes. Premenopausal women have fewer cardiovascular complications than age-matched men, but the mechanisms responsible for this protection have not been conclusively identified. Hysterectomy was performed in young female rats (leaving the ovaries intact), and 7days later the left coronary artery was ligated to produce a myocardial infarction (MI). Cardiac function at 28days post-MI was measured using echocardiography. Fractional shortening was best in non-hysterectomized (non-Hx) females and lower in both Hx females and males. Uteri were then removed from GFP rats and heterotopically transplanted into non-GFP recipients to investigate homing of uterine cells to the infarcted myocardium. Seven days later, the uterine transplant recipients underwent coronary ligation. GFP+ cells were found in the recipient hearts 7days after MI and persisted for 6months. Confocal analysis showed that homed uterine cells were located around blood vessels, suggesting their involvement in neovascularization. We then evaluated uterine cell transplantation by intravenously injecting GFP+ uterine cells into Hx females immediately after MI. These GFP+ cells were found to home to the injured myocardium, stimulate angiogenesis, improve cardiac function, and increase survival. This study demonstrates that uterine cells can home to the injured myocardium, enhance tissue repair, and prevent cardiac dysfunction. Uterine cells may play a role in the prevention of cardiovascular complications in females.Highlights: ► Hysterectomy decreases cardiac recovery in female rats after MI. ► The uterus contains regenerative cells that home to the infarcted myocardium. ► These mobilized uterine cells persisted in the injured myocardial tissue. ► Transplanted uterine cells stimulated angiogenesis and prevented infarct scar thinning. ► Transplanted uterine cells prevented cardiac dysfunction after MI.

Neuronal nitric oxide synthase is up-regulated by angiotensin II and attenuates NADPH oxidase activity and facilitates relaxation in murine left ventricular myocytes

2012-04-05

Abstract: Angiotensin II (Ang II) is critical in myocardial pathogenesis, mostly via stimulating NADPH oxidase. Neuronal nitric oxide synthase (nNOS) has recently been shown to play important roles in modulating myocardial oxidative stress and contractility. Here, we examine whether nNOS is regulated by Ang II and affects NADPH oxidase production of intracellular reactive oxygen species (ROSi) and contractile function in left ventricular (LV) myocytes. Our results showed that Ang II induced biphasic effects on ROSi and LV myocyte relaxation (TR50) without affecting the amplitude of sarcomere shortening and L-type Ca2+ current density: TR50 was prolonged at 30min but was shortened after 3h (or after Ang II treatment in vivo). Correspondingly, ROSi was increased, followed by a reduction to control level. Quantitative RT-PCR and immunoblotting experiments showed that Ang II (3h) increased the mRNA and protein expression of nNOS and increased NO production (nitrite assay) in LV myocyte homogenates, suggesting that nNOS activity may be enhanced and involved in mediating the effects of Ang II. Indeed, n(omega)-nitro-l-arginine methyl ester (l-NAME) or a selective nNOS inhibitor, S-methyl-l-thiocitrulline (SMTC) increased NADPH oxidase production of superoxide/ROSi and abolished faster myocyte relaxation induced by Ang II. The positive lusitropic effect of Ang II was not mediated by PKA-, CaMKII-dependent signaling or peroxynitrite. Conversely, inhibition of cGMP/PKG pathway abolished the Ang II-induced faster relaxation by reducing phospholamban (PLN) Ser16 phosphorylation. Taken together, these results clearly demonstrate that myocardial nNOS is up-regulated by Ang II and functions as an early adaptive mechanism to attenuate NADPH oxidase activity and facilitate myocardial relaxation.Highlights: ► Ang II activated NADPH oxidase (NOX2) and prolonged LV myocyte relaxation at 30min. ► nNOS is upregulated by Ang II (3h), inhibited NOX2 and shortened myocyte relaxation. ► Myocyte contraction and L-type Ca2+ current density were not affected by Ang II (3h). ► cGMP/PKG-dependent PLN-p mediated faster relaxation induced by nNOS and Ang II. ► PKA, CaMKII-pathway or peroxynitrite did not prevent the lusitropic effect of nNOS.

Association of Stat3 with HSF1 plays a critical role in G-CSF-induced cardio-protection against ischemia/reperfusion injury

2012-03-07

Abstract: Granulocyte colony-stimulating factor (G-CSF) has been shown to be cardio-protective against ischemia through activating Jak2/Stat3 pathway, however, the mechanism is unclear. Heat shock transcription factor 1 (HSF1), a definite endogenous protective protein in cardiomyocytes, may interact with Stat family under stress conditions. We hypothesized that G-CSF could induce cardio-protection against ischemia/reperfusion (I/R) through association of HSF1 with Stat3. To test the hypothesis, we built cardiac I/R injury model with HSF1 knockout (KO) mice and wild type (WT) mice by occlusion of the left anterior descending (LAD) coronary artery for 30min and subsequent release of the occlusion for 24h. These mice were administered with G-CSF (100μg/kg/day) or vehicle subcutaneously for 3days before surgery. As expected, G-CSF induced significant cardio-protections against I/R injury, characterized by higher ejection fraction (EF%), lower left ventricular end diastolic pressure (LVEDP), increased dp/dt value and decreased infarct area as compared with the vehicle treatment in WT mice. In HSF1-KO mice, however, these cardio-protections induced by G-CSF were greatly attenuated. Inhibition of oxidative stress-induced cardiomyocyte apoptosis by G-CSF also disappeared due to the deficiency of HSF1 in vitro and in vivo. Furthermore, G-CSF increased the phosphorylation and the association of Stat3 with HSF1, which enhanced transcriptional activity of HSF1. Inhibition of either Stat3 or HSF1 by pharmacological agents suppressed G-CSF-induced association of the two proteins and anti-apoptotic effect on cardiomyocytes. Our data suggest that G-CSF stimulates phosphorylation and association of Stat3 with HSF1 and therefore enhances transcriptional activity of HSF1, leading to the cardio-protection against I/R injury.Highlights: ► HSF1 plays a key role in G-CSF-induced cardiac protection against I/R. ► G-CSF induced interaction of Stat3 with HSF1 on cardiomyocytes following I/R. ► Inhibitor of Stat3 or HSF1 partly abolished the anti-apoptotic effects of G-CSF.

Physiological consequences of transient outward K+ current activation during heart failure in the canine left ventricle

2012-03-14

Abstract: Background: Remodeling of ion channel expression is well established in heart failure (HF). We determined the extent to which Ito is reduced in tachypacing-induced HF and assessed the ability of an Ito activator (NS5806) to recover this current. Method and results: Whole-cell patch clamp was used to record Ito in epicardial (Epi) ventricular myocytes. Epi- and endocardial action potentials were recorded from left ventricular wedge preparations. Right ventricular tachypacing-induced heart failure reduced Ito density in Epi myocytes (Control=22.1±1.9pA/pF vs 16.1±1.4 after 2weeks and 10.7±1.4pA/pF after 5weeks, +50mV). Current decay as well as recovery of Ito from inactivation progressively slowed with the development of heart failure. Reduction of Ito density was paralleled by a reduction in phase 1 magnitude, epicardial action potential notch and J wave amplitude recorded from coronary-perfused left ventricular wedge preparations. NS5806 increased Ito (at +50mV) from 16.1±1.4 to 23.9±2.1pA/pF (p<0.05) at 2weeks and from 10.7±1.4 to 14.4±1.9pA/pF (p<0.05) in 5weeks tachypaced dogs. NS5806 increased both fast and slow phases of Ito recovery in 2 and 5-week HF cells and restored the action potential notch and J wave in wedge preparations from HF dogs. Conclusions: The Ito agonist NS5806 increases the rate of recovery and density of Ito, thus reversing the HF-induced reduction in these parameters. In wedge preparations from HF dogs, NS5806 restored the spike-and-dome morphology of the Epi action potential providing proof of principal that some aspects of electrical remodelling during HF can be pharmacologically reversed.Highlights: ► Heart failure (HF) reduced phase 1 repolarization in single and multicellular tissue. ► Ito density was reduced in HF Epi cells compared to normal Epi cells. ► The Ito activator NS5806 increased Ito density toward normal in HF cells. ► NS5806 restored phase 1 repolarization in wedge preparations from failing hearts. ► Restoration of Ito may be a novel approach to the treatment of HF.

Increased myofilament Ca2+ sensitivity and diastolic dysfunction as early consequences of Mybpc3 mutation in heterozygous knock-in mice

2012-03-26

Abstract: Hypertrophic cardiomyopathy (HCM) is frequently caused by mutations in MYBPC3 encoding cardiac myosin-binding protein C (cMyBP-C). The mechanisms leading from gene mutations to the HCM phenotype remain incompletely understood, partially because current mouse models of HCM do not faithfully reflect the human situation and early hypertrophy confounds the interpretation of functional alterations. The goal of this study was to evaluate whether myofilament Ca2+ sensitization and diastolic dysfunction are associated or precede the development of left ventricular hypertrophy (LVH) in HCM. We evaluated the function of skinned and intact cardiac myocytes, as well as the intact heart in a recently developed Mybpc3-targeted knock-in mouse model carrying a point mutation frequently associated with HCM. Compared to wild-type, 10-week old homozygous knock-in mice exhibited i) higher myofilament Ca2+ sensitivity in skinned ventricular trabeculae, ii) lower diastolic sarcomere length, and faster Ca2+ transient decay in intact myocytes, and iii) LVH, reduced fractional shortening, lower E/A and E′/A′, and higher E/E′ ratios by echocardiography and Doppler analysis, suggesting systolic and diastolic dysfunction. In contrast, heterozygous knock-in mice, which mimic the human HCM situation, did not exhibit LVH or systolic dysfunction, but exhibited higher myofilament Ca2+ sensitivity, faster Ca2+ transient decay, and diastolic dysfunction. These data demonstrate that myofilament Ca2+ sensitization and diastolic dysfunction are early phenotypic consequences of Mybpc3 mutations independent of LVH. The accelerated Ca2+ transients point to compensatory mechanisms directed towards normalization of relaxation. We propose that HCM is a model for diastolic heart failure and this mouse model could be valuable in studying mechanisms and treatment modalities.Highlights: ► Absence of left ventricular hypertrophy in heterozygous Mybpc3-targeted knock-in mice. ► Myofilament Ca2+ sensitization in heterozygous Mybpc3-targeted knock-in mice. ► Diastolic dysfunction independent of left ventricular hypertrophy. ► Hypertrophic cardiomyopathy as a model of diastolic heart failure.

Adenosine-mediated inhibition of 5′-AMP-activated protein kinase and p38 mitogen-activated protein kinase during reperfusion enhances recovery of left ventricular mechanical function

Mohamed A. Omar, Sanam Verma, Alexander S. Clanachan — 2012-04-02

Abstract: Attenuation of excessive rates of myocardial glycolysis limits proton production and Ca2+ overload during reperfusion and improves recovery of post-ischemic left ventricular (LV) function. In order to elucidate mechanisms underlying glycolytic inhibition by adenosine (ADO), this study tested the hypothesis that the beneficial effects of ADO are due to Ser/Thr protein phosphatase (PP)-mediated inhibition of 5′-AMP-activated protein kinase (AMPK) and phosphofructokinase-2 (PFK-2). In isolated perfused working rat hearts subjected to global ischemia (GI) and reperfusion, ADO (500μmol/l), added 5min prior to the onset of GI and present throughout reperfusion, inhibits glycolysis and proton production during reperfusion and improves post-ischemic LV work. These metabolic effects of ADO are also evident during aerobic perfusion. Assays of glycolytic intermediates show that ADO-induced glycolytic inhibition occurs at the step catalyzed by PFK-1, an effect mediated by reduced activation of PFK-2 by AMPK. The PP1 and PP2A inhibitors, cantharidin (5μmol/l) or okadaic acid (0.1μmol/l), added 10min prior to ADO prevent ADO-induced inhibition of glycolysis and AMPK, as well as ADO-induced cardioprotection. ADO also inhibits p38 MAPK phosphorylation during reperfusion in a cantharidin-sensitive manner, and pharmacological inhibition of p38 MAPK (by SB202190, 10μmol/l) during reperfusion also reduces glycolysis and is cardioprotective. These results indicate that attenuation of glycolysis during reperfusion and cardioprotection can be achieved by inhibition of the stress kinases, AMPK and p38 MAPK.Graphical abstract: Highlights: ► Adenosine inhibits glycolysis via actions on AMPK, PFK-2 and PFK-1. ► Adenosine effects are mainly evident during reperfusion. ► Adenosine effects are mediated by activation of protein phosphatases. ► p38 MAPK inhibitor reduces glycolysis by a mechanism distinct than adenosine.