AMP-activated Protein Kinase In Heart Research Articles

AMP-activated Protein Kinase Phosphorylates Cardiac Troponin I at Ser-150 to Increase Myofilament Calcium Sensitivity and Blunt PKA-dependent Function

AMP-activated protein kinase (AMPK) is an energy-sensing enzyme central to the regulation of metabolic homeostasis. In the heart AMPK is activated during cardiac stress-induced ATP depletion and functions to stimulate metabolic pathways that restore the AMP/ATP balance. Recently it was demonstrated that AMPK phosphorylates cardiac troponin I (cTnI) at Ser-150 in vitro. We sought to determine if the metabolic regulatory kinase AMPK phosphorylates cTnI at Ser-150 in vivo to alter cardiac contractile function directly at the level of the myofilament. Rabbit cardiac myofibrils separated by two-dimensional isoelectric focusing subjected to a Western blot with a cTnI phosphorylation-specific antibody demonstrates that cTnI is endogenously phosphorylated at Ser-150 in the heart. Treatment of myofibrils with the AMPK holoenzyme increased cTnI Ser-150 phosphorylation within the constraints of the muscle lattice. Compared with controls, cardiac fiber bundles exchanged with troponin containing cTnI pseudo-phosphorylated at Ser-150 demonstrate increased sensitivity of calcium-dependent force development, blunting of both PKA-dependent calcium desensitization, and PKA-dependent increases in length dependent activation. Thus, in addition to the defined role of AMPK as a cardiac metabolic energy gauge, these data demonstrate AMPK Ser-150 phosphorylation of cTnI directly links the regulation of cardiac metabolic demand to myofilament contractile energetics. Furthermore, the blunting effect of cTnI Ser-150 phosphorylation cross-talk can uncouple the effects of myofilament PKA-dependent phosphorylation from β-adrenergic signaling as a novel thin filament contractile regulatory signaling mechanism.

Whole Body Deletion of AMP-activated Protein Kinase β2 Reduces Muscle AMPK Activity and Exercise Capacity

AMP-activated protein kinase (AMPK) β subunits (β1 and β2) provide scaffolds for binding α and γ subunits and contain a carbohydrate-binding module important for regulating enzyme activity. We generated C57Bl/6 mice with germline deletion of AMPK β2 (β2 KO) and examined AMPK expression and activity, exercise capacity, metabolic control during muscle contractions, aminoimidazole carboxamide ribonucleotide (AICAR) sensitivity, and susceptibility to obesity-induced insulin resistance. We find that β2 KO mice are viable and breed normally. β2 KO mice had a reduction in skeletal muscle AMPK α1 and α2 expression despite up-regulation of the β1 isoform. Heart AMPK α2 expression was also reduced but this did not affect resting AMPK α1 or α2 activities. AMPK α1 and α2 activities were not changed in liver, fat, or hypothalamus. AICAR-stimulated glucose uptake but not fatty acid oxidation was impaired in β2 KO mice. During treadmill running β2 KO mice had reduced maximal and endurance exercise capacity, which was associated with lower muscle and heart AMPK activity and reduced levels of muscle and liver glycogen. Reductions in exercise capacity of β2 KO mice were not due to lower muscle mitochondrial content or defects in contraction-stimulated glucose uptake or fatty acid oxidation. When challenged with a high-fat diet β2 KO mice gained more weight and were more susceptible to the development of hyperinsulinemia and glucose intolerance. In summary these data show that deletion of AMPK β2 reduces AMPK activity in skeletal muscle resulting in impaired exercise capacity and the worsening of diet-induced obesity and glucose intolerance.

Nutrient Stress Activates Inflammation and Reduces Glucose Metabolism by Suppressing AMP-Activated Protein Kinase in the Heart

OBJECTIVEHeart failure is a major cause of mortality in diabetes and may be causally associated with altered metabolism. Recent reports indicate a role of inflammation in peripheral insulin resistance, but the impact of inflammation on cardiac metabolism is unknown. We investigated the effects of diet-induced obesity on cardiac inflammation and glucose metabolism in mice.RESEARCH DESIGN AND METHODSMale C57BL/6 mice were fed a high-fat diet (HFD) for 6 weeks, and heart samples were taken to measure insulin sensitivity, glucose metabolism, and inflammation. Heart samples were also examined following acute interleukin (IL)-6 or lipid infusion in C57BL/6 mice and in IL-6 knockout mice following an HFD.RESULTSDiet-induced obesity reduced cardiac glucose metabolism, GLUT, and AMP-activated protein kinase (AMPK) levels, and this was associated with increased levels of macrophages, toll-like receptor 4, suppressor of cytokine signaling 3 (SOCS3), and cytokines in heart. Acute physiological elevation of IL-6 suppressed glucose metabolism and caused insulin resistance by increasing SOCS3 and via SOCS3-mediated inhibition of insulin receptor substrate (IRS)-1 and possibly AMPK in heart. Diet-induced inflammation and defects in glucose metabolism were attenuated in IL-6 knockout mice, implicating the role of IL-6 in obesity-associated cardiac inflammation. Acute lipid infusion caused inflammation and raised local levels of macrophages, C-C motif chemokine receptor 2, SOCS3, and cytokines in heart. Lipid-induced cardiac inflammation suppressed AMPK, suggesting the role of lipid as a nutrient stress triggering inflammation.CONCLUSIONSOur findings that nutrient stress activates cardiac inflammation and that IL-6 suppresses myocardial glucose metabolism via inhibition of AMPK and IRS-1 underscore the important role of inflammation in the pathogenesis of diabetic heart.

Phosphorylation of translation factors in response to anoxia in turtles, Trachemys scripta elegans: role of the AMP-activated protein kinase and target of rapamycin signalling pathways

Long-term survival of oxygen deprivation by animals with well-developed anoxia tolerance depends on multiple biochemical adaptations including strong metabolic rate depression. We investigated whether the AMP-activated protein kinase (AMPK) could play a regulatory role in the suppression of protein synthesis that occurs when turtles experience anoxic conditions. AMPK activity and the phosphorylation state of ribosomal translation factors were measured in liver, heart, red muscle and white muscle of red-eared slider turtles (Trachemys scripta elegans) subjected to 20 h of anoxic submergence. AMPK activity increased twofold in white muscle of anoxic turtles compared with aerobic controls but remained unchanged in liver and red muscle, whereas in heart AMPK activity decreased by 40%. Immunoblotting with phospho-specific antibodies revealed that eukaryotic elongation factor-2 phosphorylation at the inactivating Thr56 site increased six- and eightfold in red and white muscles from anoxic animals, respectively, but was unchanged in liver and heart. The phosphorylation state of the activating Thr389 site of p70 ribosomal protein S6 kinase was reduced under anoxia in red muscle and heart but was unaffected in liver and white muscle. Exposure to anoxia decreased 40S ribosomal protein S6 phosphorylation in heart and promoted eukaryotic initiation factor 4E-binding protein-1 (4E-BP1) dephosphorylation in red muscle, but surprisingly increased 4E-BP1 phosphorylation in white muscle. The changes in phosphorylation state of translation factors suggest that organ-specific patterns of signalling and response are involved in achieving the anoxia-induced suppression of protein synthesis in turtles.

AMP‐activated protein kinase: a core signalling pathway in the heart

Over the past decade, AMP-activated protein kinase (AMPK) has emerged as an important intracellular signalling pathway in the heart. Activated AMPK stimulates the production of ATP by regulating key steps in both glucose and fatty acid metabolism. It has an inhibitory effect on cardiac protein synthesis. AMPK also interacts with additional intracellular signalling pathways in a coordinated network that modulates essential cellular processes in the heart. Evidence is accumulating that AMPK may protect the heart from ischaemic injury and limit the development of cardiac myocyte hypertrophy to various stimuli. Heart AMPK is activated by hormones, cytokines and oral hypoglycaemic drugs that are used in the treatment of type 2 diabetes. The tumour suppressor LKB1 is the major regulator of AMPK activity, but additional upstream kinases and protein phosphatases also contribute. Mutations in the regulatory gamma2 subunit of AMPK lead to an inherited syndrome of hypertrophic cardiomyopathy and ventricular pre-excitation, which appears to be due to intracellular glycogen accumulation. Future research promises to elucidate the molecular mechanisms responsible for AMPK activation, novel downstream AMPK targets, and the therapeutic potential of targeting AMPK for the prevention and treatment of myocardial ischaemia or cardiac hypertrophy.

Alpha1 and beta adrenergic stimulation on regulation of amp-activated protein kinase in heart

Alpha1 and beta adrenergic stimulation on regulation of amp-activated protein kinase in heart

Activation of AMPK α- and γ-isoform complexes in the intact ischemic rat heart

AMP-activated protein kinase (AMPK) plays a key role in modulating cellular metabolic processes. AMPK, a serine-threonine kinase, is a heterotrimeric complex of catalytic alpha-subunits and regulatory beta- and gamma-subunits with multiple isoforms. Mutations in the cardiac gamma(2)-isoform have been associated with hypertrophic cardiomyopathy and pre-excitation syndromes. However, physiological regulation of AMPK complexes containing different subunit isoforms is not well defined and is important for an understanding of the function of this signaling pathway in the intact heart. We evaluated the kinase activity associated with heart AMPK complexes containing specific alpha- and gamma-subunit isoforms of AMPK in an in vivo rat model of regional ischemia. Left coronary artery occlusion activated the immunoprecipitated alpha(1)-isoform (6-fold, P < 0.01) and alpha(2)-isoform (9-fold, P < 0.01) in the ischemic left ventricle compared with sham controls. The degree of alpha-subunit activation depended on the extent of ischemia and paralleled echocardiographic contractile dysfunction. The regulatory gamma(1)- and gamma(2)-isoforms were expressed in the heart. The gamma(1)- and gamma(2)-isoforms coimmunoprecipitated with alpha(1)- and alpha(2)-isoforms in proportion to alpha-subunit content. gamma(1)-Isoform immunocomplexes accounted for 70% of AMPK activity and AMPK phosphorylation (Thr(172)) in hearts. Ischemia similarly increased AMPK activity associated with the gamma(1)- and gamma(2)-isoform complexes threefold (P < 0.01 for each). Thus AMPK catalytic alpha(1)- and alpha(2)-isoforms are activated by regional ischemia in vivo in the heart, irrespective of the regulatory gamma(1)- or gamma(2)-isoforms to which they are complexed. Despite the pathophysiological importance of gamma(2)-isoform mutations, gamma(1)-isoform complexes account for most of the AMPK activity in the ischemic heart.

Covalent activation of heart AMP‐activated protein kinase in response to physiological concentrations of long‐chain fatty acids

Rat hearts were perfused for 1 h with 5 mm glucose with or without palmitate or oleate at concentrations characteristic of the fasting state. The inclusion of fatty acids resulted in increased activities of the alpha-1 or the alpha-2 isoforms of AMP-activated protein kinase (AMPK), increased phosphorylation of acetyl-CoA carboxylase and a decrease in the tissue content of malonyl-CoA. Activation of AMPK was not accompanied by any changes in the tissue contents of ATP, ADP, AMP, phosphocreatine or creatine. Palmitate increased phosphorylation of Thr172 within AMPK alpha-subunits and the activation by palmitate of both AMPK isoforms was abolished by protein phosphatase 2C leading to the conclusion that exposure to fatty acid caused activation of an AMPK kinase or inhibition of an AMPK phosphatase. In vivo, 24 h of starvation also increased heart AMPK activity and Thr172 phosphorylation of AMPK alpha-subunits. Perfusion with insulin decreased both alpha-1 and alpha-2 AMPK activities and increased malonyl-CoA content. Palmitate prevented both of these effects. Perfusion with epinephrine decreased malonyl-CoA content without an effect on AMPK activity but prevented the activation of AMPK by palmitate. The concept is discussed that activation of AMPK by an unknown fatty acid-driven signalling process provides a mechanism for a 'feed-forward' activation of fatty acid oxidation.