AMP-activated Protein Kinase Signaling Cascade Research Articles

Physiological and skeletal muscle responses to high-intensity interval exercise in Thoroughbred horses.

The purpose of this study was to determine whether acute high-intensity interval exercise or sprint interval exercise induces greater physiological and skeletal muscle responses compared to moderate-intensity continuous exercise in horses. In a randomized crossover design, eight trained Thoroughbred horses performed three treadmill exercise protocols consisting of moderate-intensity continuous exercise (6 min at 70% VO2max; MICT), high-intensity interval exercise (6 × 30 s at 100% VO2max; HIIT), and sprint interval exercise (6 × 15 s at 120% VO2max; SIT). Arterial blood samples were collected to measure blood gas variables and plasma lactate concentration. Biopsy samples were obtained from the gluteus medius muscle before, immediately after, 4 h, and 24 h after exercise for biochemical analysis, western blotting and real-time RT-PCR. Effects of time and exercise protocol were analyzed using mixed models (p < 0.05). Heart rate and plasma lactate concentration at the end of exercise were higher in HIIT and SIT than those in MICT (heart rate, HIIT vs. MICT, p = 0.0005; SIT vs. MICT, p = 0.0015; lactate, HIIT vs. MICT, p = 0.0014; SIT vs. MICT, p = 0.0003). Arterial O2 saturation and arterial pH in HIIT and SIT were lower compared with MICT (SaO2, HIIT vs. MICT, p = 0.0035; SIT vs. MICT, p = 0.0265; pH, HIIT vs. MICT, p = 0.0011; SIT vs. MICT, p = 0.0023). Muscle glycogen content decreased significantly in HIIT (p = 0.0004) and SIT (p = 0.0016) immediately after exercise, but not in MICT (p = 0.19). Phosphorylation of AMP-activated protein kinase (AMPK) in HIIT showed a significant increase immediately after exercise (p = 0.014), but the increase was not significant in MICT (p = 0.13) and SIT (p = 0.39). At 4 h after exercise, peroxisome proliferator-activated receptor γ co-activator-1α mRNA increased in HIIT (p = 0.0027) and SIT (p = 0.0019) and vascular endothelial growth factor mRNA increased in SIT (p = 0.0002). Despite an equal run distance, HIIT and SIT cause more severe arterial hypoxemia and lactic acidosis compared with MICT. In addition, HIIT activates the AMPK signaling cascade, and HIIT and SIT elevate mitochondrial biogenesis and angiogenesis, whereas MICT did not induce any significant changes to these signaling pathways.

Understanding the Molecular Underpinnings of Neuronal Kinase CDK5 mediated Metabolic Vulnerabilities in Neuroendocrine Tumors

BackgroundAberrantly activated neuronal kinase CDK5 invoke alternations in mitochondrial dynamics and metabolic dysfunction associated with neurological disorders. Although pro‐tumorigenic role of CDK5 has been recently defined but there is insufficient data to understand the mechanistic role of CDK5 in cancer associated mitochondrial structure and function.HypothesisIn this study we intend to investigate putative signalling hotspots downstream of aberrantly activated CDK5 in neuroendocrine tumors of adrenal glands called Pheochromocytoma (PC).ApproachOur preliminary data for the first time showed deregulation of Ca2+ homeostasis and overexpression of CDK5 and its co‐activators‐p35/25 in malignant PC tumors inflicted with loss‐of‐function mutations in mitochondrial enzyme‐Succinate dehydrogenase B (SDHB). To elucidate role of CDK5 in tumorigenesis, we created a novel bi‐transgenic mouse model where doxycycline inducible overexpression of CDK5/p25 (p25OE) developed organ specific PC lesions.ResultsPhopshoproteomic analysis on p25OE PC tumors unraveled a tumor suppressive mechanism modulated by novel phosphorylation site on metabolic protein‐PRKAG2 (S‐65). PRKAG2 is a non‐catalytic regulatory subunit of AMP‐activated protein kinase (AMPK). S‐65 site on PRKAG2 were found downregulated in human PC tumors compared to normal adrenal medulla. Further biochemical characterization uncovered CDK5‐GSK3β‐AMPK crosstalk underlying human PC cellular proliferation. Picking CDK5 pocket with a novel CDK5 inhibitor ‘MRT3‐007’ induced anti‐proliferative effects with simultaneous increase in phosphorylation states of PRKAG2(S‐65) / AMPK(T‐172) sites followed by excessive mitochondrial fission.ConclusionThese intriguing initial findings set the ground to discover therapeutic vulnerabilities associated with CDK5/p25/PRKAG2/AMPK signaling cascade and Mitochondrial Dynamics in Neuroendocrine tumors.Schematic depiction of working model demonstrating a novel crosstalk between a metabolic sensor ‘PRKAG2/AMPK’ and ‘Mitochondrial Fission’ synapsed via neuronal kinase ‘CDK5/p25’ in Pheochromocytoma (PC tumors) inflicted with mutations in SDHB gene.Figure 1

Mitochondria-targeted drugs stimulate mitophagy and abrogate colon cancer cell proliferation

Mutations in the KRAS proto-oncogene are present in 50% of all colorectal cancers and are increasingly associated with chemotherapeutic resistance to frontline biologic drugs. Accumulating evidence indicates key roles for overactive KRAS mutations in the metabolic reprogramming from oxidative phosphorylation to aerobic glycolysis in cancer cells. Here, we sought to exploit the more negative membrane potential of cancer cell mitochondria as an untapped avenue for interfering with energy metabolism in KRAS variant-containing and KRAS WT colorectal cancer cells. Mitochondrial function, intracellular ATP levels, cellular uptake, energy sensor signaling, and functional effects on cancer cell proliferation were assayed. 3-Carboxyl proxyl nitroxide (Mito-CP) and Mito-Metformin, two mitochondria-targeted compounds, depleted intracellular ATP levels and persistently inhibited ATP-linked oxygen consumption in both KRAS WT and KRAS variant-containing colon cancer cells and had only limited effects on nontransformed intestinal epithelial cells. These anti-proliferative effects reflected the activation of AMP-activated protein kinase (AMPK) and the phosphorylation-mediated suppression of the mTOR target ribosomal protein S6 kinase B1 (RPS6KB1 or p70S6K). Moreover, Mito-CP and Mito-Metformin released Unc-51-like autophagy-activating kinase 1 (ULK1) from mTOR-mediated inhibition, affected mitochondrial morphology, and decreased mitochondrial membrane potential, all indicators of mitophagy. Pharmacological inhibition of the AMPK signaling cascade mitigated the anti-proliferative effects of Mito-CP and Mito-Metformin. This is the first demonstration that drugs selectively targeting mitochondria induce mitophagy in cancer cells. Targeting bioenergetic metabolism with mitochondria-targeted drugs to stimulate mitophagy provides an attractive approach for therapeutic intervention in KRAS WT and overactive mutant-expressing colon cancer.

Strawberry consumption improves aging-associated impairments, mitochondrial biogenesis and functionality through the AMP-activated protein kinase signaling cascade

Dietary polyphenols have been recently proposed as activators of the AMP-activated protein kinase (AMPK) signaling pathway and this fact might explain the relationship between the consumption of polyphenol-rich foods and the slowdown of the progression of aging. In the present work, the effects of strawberry consumption were evaluated on biomarkers of oxidative damage and on aging-associated reductions in mitochondrial function and biogenesis for 8weeks in old rats. Strawberry supplementation increased antioxidant enzyme activities, mitochondrial biomass and functionality, and decreased intracellular ROS levels and biomarkers of protein, lipid and DNA damage (P<0.05). Furthermore, a significant (P<0.05) increase in the expression of the AMPK cascade genes, involved in mitochondrial biogenesis and antioxidant defences, was also detected after strawberry intake. These in vivo results were then verified in vitro on HepG2 cells, confirming the involvement of AMPK in the beneficial effects exerted by strawberry against aging progression.

AMP-activated protein kinase and its multifaceted regulation of hepatic metabolism.

The current review summarizes recent advancements in our mechanistic and physiological understanding of the energy sensing AMP-activated protein kinase (AMPK) and its regulation of select aspects of hepatic metabolism. A highly conserved serine/threonine kinase, AMPK governs a multitude of cellular process to activate catabolic and inhibit anabolic pathways. Recent work has provided clarity as to the importance and contribution of the AMPK signaling cascade to various aspects of cellular metabolism, including lipid homeostasis, hepatic glucose production, mitochondrial metabolism, and autophagy. With more than 60 confirmed substrates, the physiological significance of AMPK signaling has been difficult to ascertain. The generation of targeted knock-in mutations on key AMPK substrates has begun to shed light on this complex system. Future studies are needed to further decipher the complexity, significance, and potential therapeutic targeting of hepatic AMPK signaling.

The protective effect of trimetazidine on myocardial ischemia/reperfusion injury through activating AMPK and ERK signaling pathway

IntroductionTrimetazidine (TMZ) is an anti-anginal drug that has been widely used in Europe and Asia. The TMZ can optimize energy metabolism via inhibition of long-chain 3-ketoacyl CoA thiolase (3-KAT) in the heart, with subsequent decrease in fatty acid oxidation and stimulation of glucose oxidation. However, the mechanism by which TMZ aids in cardioprotection against ischemic injury has not been characterized. AMP-activated protein kinase (AMPK) is an energy sensor that controls ATP supply from substrate metabolism and protects heart from energy stress. TMZ changes the cardiac AMP/ATP ratio by modulating fatty acid oxidation, thereby triggering AMPK signaling cascade that contributes to the protection of the heart from ischemia/reperfusion (I/R) injury. MethodsThe mouse model of in vivo regional ischemia and reperfusion by the ligation of the left anterior descending coronary artery (LAD) was used for determination of myocardial infarction. The infarct size was compared between C57BL/6J WT mice and AMPK kinase dead (KD) transgenic mice with or without TMZ treatment. The ex vivo working heart perfusion system was used to monitor the effect of TMZ on glucose oxidation and fatty acid oxidation in the heart. ResultsTMZ treatment significantly stimulates cardiac AMPK and extracellular signal-regulated kinase (ERK) signaling pathways (p<0.05 vs. vehicle group). The administration of TMZ reduces myocardial infarction size in WT C57BL/6J hearts, the reduction of myocardial infarction size by TMZ in AMPK KD hearts was significantly impaired versus WT hearts (p<0.05). Intriguingly, the administration of ERK inhibitor, PD98059, to AMPK KD mice abolished the cardioprotection of TMZ against I/R injury. The ex vivo working heart perfusion data demonstrated that TMZ treatment significantly activates AMPK signaling and modulating the substrate metabolism by shifting fatty acid oxidation to glucose oxidation during reperfusion, leading to reduction of oxidative stress in the I/R hearts. Therefore, both AMPK and ERK signaling pathways mediate the cardioprotection of TMZ against ischemic injury. The metabolic benefits of TMZ for angina patients could be due to the activation of energy sensor AMPK in the heart by TMZ administration.

Abstract 1136: Regulation of AMPK by androgen receptor signaling and its role in promoting prostate cancer through the use of autophagy

Abstract While we now know that the androgen receptor (AR) is important for progression to the deadly advanced stages of prostate cancer, it is poorly understood what AR-regulated processes drive this pathology. Autophagy, an enigmatic cellular recycling process, has received increased attention as of late due to its potential oncogenic role in late-stage cancers. One of the master regulators of autophagy is the 5′-AMP-activated protein kinase (AMPK). Previous work from several independent laboratories has suggested AR signaling promotes cancer progression through an AMPK-dependent signaling mechanism. Further, it was demonstrated this enzymatic cascade was specifically regulated by Ca2+/calmodulin-dependent protein kinase kinase 2 (CaMKK2). Given AMPK's known role as a regulator of autophagy, we hypothesized that AR-mediated CaMKK2-AMPK signaling could promote prostate cancer through increasing cellular rates of autophagy. To test this hypothesis, we initially assessed whether AR signaling could regulate autophagy and if so, what effect(s) this had on prostate cancer cell growth and tumor formation. We then used a variety of pharmacological and molecular approaches to determine whether CaMKK2-AMPK signaling was required and sufficient to regulate autophagy and subsequent prostate cancer cell growth in vitro and in vivo. Here, we have determined that 1) AR regulates cell metabolism and cell growth in part by increasing autophagy in prostate cancer cells, 2) functional autophagy was clinically detected in metastatic, castration-resistant cancers but not treatment-naïve, localized tumors and 3) autophagy is required for prostate cancer progression in preclinical animal models. Additionally, AR-mediated autophagy was mediated through the direct expression of CaMKK2 and subsequent phosphorylation/activation of AMPK. Correspondingly, levels of both CaMKK2 and phosphorylated/activated AMPK correlated with prostate cancer progression in genetic mouse models and patients. Mechanistically, AR-mediated autophagy appears to promote cell growth by augmenting intracellular lipid accumulation, a hallmark of prostate cancers. Taken together, our findings converge to demonstrate that AR signaling can co-opt the AMPK signaling cascade, a known homeostatic mechanism, to promote prostate cancer by increasing autophagy. The current study points to the potential utility of developing metabolic-targeted therapies directed towards the CaMKK2-AMPK signaling axis for the treatment of prostate cancer. Further, an inhibitor of this signaling cascade could serve as an alternative, more specific therapeutic compared to existing inhibitors of autophagy that, to date, have demonstrated limited efficacy in clinical trials due to their toxicity and poor pharmacokinetics. Citation Format: Yan Shi, Efrosini Tsouko, Alicia M. Blessing, Jayantha Tennakoon, Jenny J. Han, Michael M. Ittmann, Daniel E. Frigo. Regulation of AMPK by androgen receptor signaling and its role in promoting prostate cancer through the use of autophagy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1136. doi:10.1158/1538-7445.AM2015-1136

Abstract 12117: The Cardioprotection of Trimetazidine Against Ischemic Injury is Mediated by AMPK and ERK Signaling Pathway

Introduction: Trimetazidine (TMZ) is an anti-anginal drug that has been widely used in Europe and Asia. The TMZ can optimize energy metabolism via inhibition of long-chain 3-ketoacyl CoA thiolase (3-KAT) in the heart, with subsequent decrease in fatty acid oxidation and stimulation of glucose oxidation. However, the mechanism by which TMZ aids in cardioprotection against ischemic injury has not been characterized. Hypothesis: AMP-activated protein kinase (AMPK) is an energy sensor that control ATP supply from substrate metabolism and protect heart from energy stress. TMZ changes the cardiac AMP/ATP ratio via modulating fatty acid oxidation, thereby it may trigger AMPK signaling cascade that contribute to protection heart from ischemia/reperfusion (I/R) injury. Methods: The mouse in vivo regional ischemia and reperfusion by the ligation of the left anterior descending coronary artery (LAD) were used for determination of myocardial infarction. The infarct size was compared between C57BL/6J WT mice and AMPK kinase dead (KD) transgenic mice with or without TMZ treatment. The ex vivo working heart perfusion system was used to monitor the effect of TMZ on glucose oxidation and fatty acid oxidation in the heart. Results: TMZ treatment significantly stimulates cardiac AMPK and extracellular signal-regulated kinase (ERK) signaling pathways (p&lt;0.05 vs. vehicle group). The administration of TMZ reduces myocardial infarction size in WT C57BL/6J hearts, the reduction of myocardial infarction size by TMZ in AMPK KD hearts was significantly impaired versus WT hearts (p&lt;0.05). Intriguingly, the administration of ERK inhibitor, PD 98059, to AMPK KD mice abolished the cardioprotection of TMZ against I/R injury. The ex vivo working heart perfusion data demonstrated that TMZ treatment significantly activates AMPK signaling and modulating the substrate metabolism by shifting fatty acid oxidation to glucose oxidation during reperfusion, leading to reduction of oxidative stress in the I/R hearts. Conclusions: Both AMPK and ERK signaling pathways mediate the cardioprotection of TMZ against ischemic injury. The metabolic benefits of TMZ for angina patients could be due to the activation of energy sensor AMPK in the heart by TMZ administration.

Abstract P5-04-15: Investigating the role of the AMPK signaling cascade in breast cancer tumor dormancy

Abstract Breast cancer can recur in patients long after treatment for initial disease. The mechanisms underlying recurrence as well as the characteristics of these dormant tumor cells remain poorly understood. AMPK (AMP-activated protein kinase) is the central metabolic regulator of the cell, and has two catalytic isoforms, a1 and α2. The loss of AMPK, more specifically AMPKα2, is observed in primary breast cancer and its expression could function to limit tumor expansion in vivo. We hypothesize that the expression of AMPKα2 will confer a survival advantage in dormant breast cancer cells by upregulating the p38MAPK signaling axis. In order to investigate breast cancer dormancy mechanisms in vitro, we generated mammospheres using the estrogen-dependent MCF-7 human breast cancer cell line, which has low expression of AMPKα2. Control MCF-7 cells expressing GFP and MCF-7 cells overexpressing the AMPKα1 or AMPKα2 isoforms were cultured as mammospheres and exposed to various metabolic stressors including estrogen, glucose, and serum deprivation. Spheroids were evaluated for size, viability, gene expression at the RNA level by qRT-PCR, and intracellular signaling by immunoblot as well as immunohistochemistry. AMPKα2 expressing mammospheres were larger and more viable than either the GFP or AMPKα1 expressing spheres. In addition, these cells were resistant to serum deprivation, and upregulated members of the p38MAPK signaling axis, a key player in tumor dormancy. To evaluate estrogen deprivation-induced dormancy in vivo, MCF-7 cell lines expressing either GFP or AMPKα2 embedded in a matrigel matrix were implanted in athymic nude mice containing supplemental slow-release estradiol pellets. After one week, estradiol pellets were removed to induce a dormant period for roughly thirty days. Evaluation of matrigel plugs at this time indicated that AMPKα2 expressing cells demonstrated significant survival whereas GFP expressing cells were significantly depleted. Estadiol pellets were then re-implanted for five weeks to determine if the AMPKα2 expressing cells still responded to estradiol supplementation.This resulted in the AMPKα2 expressing tumors rapidly expanding to a maximal size as compared to the GFP expressing tumors. Consistent with our findings in vitro using mammosphere cultures, the AMPKα2 expressing cells had higher levels of phospho-p38MAPK than the GFP expressing cells, as seen by immunofluorescence. We conclude that the expression of AMPKα2 promotes both survival and dormancy in estrogen-sensitive breast tumor cells, thus implying a significant role in breast cancer recurrence. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-04-15.

Recent Insights into the Role of Hypothalamic AMPK Signaling Cascade upon Metabolic Control

In 2004, two seminal papers focused on the role of AMP-activated protein kinase (AMPK) in the hypothalamus opened new avenues of research in the field of the central regulation of energy homeostasis. Over the following 8 years, hundreds of studies have firmly established hypothalamic AMPK as a key sensor and integrator of hormonal and nutritional signals with neurochemical and neurophysiological responses to regulate whole-body energy balance. In this review article we aim to discuss the most recent findings in this particular area of research, highlighting the function of hypothalamic AMPK in appetite, thermogenesis, and peripheral glucose metabolism. The diversity of mechanisms by which hypothalamic AMPK regulates energy homeostasis illustrates the importance of this evolutionary-conserved energy signaling cascade in the control of this complex and fundamental biological process.

The AMPK signaling cascade in metabolic regulation: view from the chair

The AMP-activated protein kinase (AMPK) system is a keyplayer in the regulation of cellular energy levels. By switch-ing on catabolic pathways that generate ATP, this molecularswitch triggers substrate oxidation processes during physicalexercise and mediates the metabolic actions of severalhormones and adipokines known to control whole-bodyenergy balance. In this symposium, we learned about thediscovery of AMPK, from the details of its complex molecularstructure to its molecular mechanisms of activation bymetabolic stresses, exercise and humoral factors. As AMPKis also activated by two classes of antidiabetic drugs (forexample, biguanides and thiazolidinediones) it represents anattractive target for the treatment of obesity and itsassociated metabolic abnormalities. The potential upstreamand downstream signals that may be involved in relayingAMPK’s metabolic actions were also discussed by severalspeakers of the symposium as summarized below.The mammalian AMPK system was discovered two decadesago by the first speaker of this session, Grahame Hardie, whoinitially reported that AMPK phosphorylates and inactivateskey enzymes of lipid biosynthesis, that is acetyl-CoAcarboxylase and 3-hydroxy-3-methylglutaryl-CoA reductase.

Adiponectin modulates carnitine palmitoyltransferase-1 through AMPK signaling cascade in rat cardiomyocytes

Adiponectin, an adipocyte-derived polypeptide hormone, plays an important role in regulating fatty acid oxidation. β-oxidation of fatty acids supplies most of the cardiac energy and carnitine palmitoyltransferase (CPT)-1 serves as a key regulator during this process. To characterize the potential effects of adiponectin on CPT-1, we incubated rat neonatal cardiomyocytes with globular adiponectin (gAd). Results showed that gAd promoted the activity and mRNA expression of CPT-1. The underlying signal pathway involved in this modulatory effect was further investigated. Inhibition of AMP-activated protein kinase (AMPK) with adenine 9-β- d-arabinofuranoside (AraA) completely abrogated gAd-mediated AMPK and acetyl coenzyme A carboxylase (ACC) phosphorylation and suppressed the promotion of CPT-1 activity. gAd also induced the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and peroxisome proliferator-activated receptor (PPAR)-α, which was inhibited by AraA. SB202190, a p38MAPK inhibitor, blocked gAd-stimulated PPAR-α phosphorylation. When AMPK and/or p38MAPK was inhibited, gAd-enhanced mRNA expression of CPT-1 was partially reduced. In conclusion, our study suggests that the activation of AMPK signaling cascade participates in the promotion effect of gAd on CPT-1.

Adiponectin actions in the cardiovascular system

Obesity is strongly associated with the pathogenesis of type 2 diabetes, hypertension, and cardiovascular disease. Levels of the hormone adiponectin are downregulated in obese individuals, and several experimental studies show that adiponectin protects against the development of various obesity-related metabolic and cardiovascular diseases. Adiponectin exhibits favorable effects on atherogenesis, endothelial function, and vascular remodeling by modulation of signaling cascades in cells of the vasculature. More recent findings have shown that adiponectin directly affects signaling in cardiac cells and is beneficial in the setting of pathological cardiac remodeling and acute cardiac injury. Several of these effects of adiponectin have been attributed to the activation of the 5' AMP-activated protein kinase signaling cascade and other signaling proteins. This review will discuss the epidemiological and experimental studies that have elucidated the role of adiponectin in a variety of cardiovascular diseases.

Long-chain acyl-CoA esters inhibit phosphorylation of AMP-activated protein kinase at threonine-172 by LKB1/STRAD/MO25

Activation of the AMP-activated protein kinase (AMPK) results in acute changes in cellular metabolism and transcriptional events that make the cell more robust when encountering an energy challenge. AMPK is thought to be inhibited by glycogen, the major storage form of intracellular carbohydrate. We hypothesized that long-chain acyl-CoA esters (LCACEs) might also inhibit AMPK signaling. Cytosolic LCACEs are available for immediate transport and oxidation within the mitochondria and accordingly may be representative of the lipid energy charge of the cell. We found that LCACEs inhibited phosphorylation of AMPK by the recombinant AMPK kinase (AMPKK) LKB1/STRAD/MO25 in a concentration-dependent manner. Palmitoyl-CoA (PCoA) did not affect the activity of phosphothreonine-172 AMPK. PCoA potently inhibited AMPKK purified from liver. Conversely, PCoA stimulated the kinase activity of LKB1/STRAD/MO25 toward the peptide substrate LKB1tide. Octanoyl-CoA, palmitate, and palmitoylcarnitine did not inhibit AMPKK activity. Removal of AMP from the reaction mixture resulted in reduced AMPKK activity in the presence of PCoA. In conclusion, these results demonstrate that the AMPKK activity of LKB1/STRAD/MO25 is substrate specific and distinct from the kinase activity of LKB1/STRAD/MO25 toward the peptide substrate LKB1tide. They also demonstrate that LCACEs inhibit the AMPKK activity of LKB1/STRAD/MO25 in a specific manner with a dependence on both a long fatty chain and a CoA moiety. These results suggest that the AMPK signaling cascade may directly sense and respond to the lipid energy charge of the cell.

NO PAIN, NO GAIN?

Wouldn't it be nice to enjoy the benefits of exercise without the effort?For example, endurance physical training results in elevated muscle mitochondrial content for increased aerobic respiration and the transition of fast-twitch fibers into slower oxidative fibers. One step towards the goal of gain without pain is the elucidation of the signaling cascades that increase both metabolic and contractile capacities in muscle. Recently, it has been appreciated that one important signaling cascade during exercise involved AMP-activated protein kinase (AMPK). Indeed, the activation of AMPK can mediate some of the metabolic effects associated with exercise. However, the role of this pathway in modifying muscle contractile properties remains unclear. The paper by Charles T. Putman and collaborators examines this question and further investigates the effects of activating the AMPK pathway on glycolytic and oxidative metabolic profiles.In order to test these hypotheses, the authors injected rats for a month with a compound, AICAR, that is known to activate AMPK. The team looked at the activity of several mitochondrial enzymes as markers of mitochondrial biogenesis, the expression of uncoupling protein-3 (UCP3), a mitochondrial protein that is associated with the regulation of burning fat and, finally,the proportion of oxidative fibers in muscle.The first set of results in fast-twitch muscle revealed that the activity of mitochondrial enzymes was elevated in the groups of rats treated with AICAR. This result suggests that AMPK plays a role in mitochondrial biogenesis due to exercise.Also, the increased mitochondrial biogenesis led to an increase in UCP3 in the treated rats. However, the increase in UCP3 protein level was higher than for other mitochondrial proteins, which suggests that UCP3 might be specifically targeted in mitochondria by the AMPK signaling pathway. The team suspects that as UCP3 expression has been associated with the metabolism of lipids, the preferential upregulation of this protein could be related to the increased oxidation of fatty acids that occurs during exercise.For the second part of their study, the authors examined the expression of various myosin heavy chain isoforms, which are markers of the different fiber types in fast-twitch muscle. The proportion of oxidative fibers was not increased in AICAR-treated animals. Therefore, activation of AMPK cannot mimic the contractile adaptations that occur in muscle during endurance training.Overall, this study shows that activation of the AMPK signaling cascade leads to some of the beneficial metabolic effects of exercise, without affecting muscle contractile properties. This type of pioneering study is fundamental to our understanding of how exercise affects organismal physiology. Finally, the search for evolutionary conserved signaling pathways in other organisms should be of great interest for comparative physiologists in the years to come.