We investigated the importance of the two catalytic ␣-isoforms of the 5-AMP-activated protein kinase (AMPK) in 5-aminoimidazole-4-carboxamide-1-␤-4-ribofuranoside (AICAR) and contraction-induced glucose uptake in skeletal muscle. Incubated soleus and EDL muscle from whole-body ␣ 2 -or ␣ 1 -AMPK knockout (KO) and wild type (WT) mice were incubated with 2.0 mM AICAR or electrically stimulated to contraction. Both AICAR and contraction increased 2DG uptake in WT muscles. KO of ␣ 2 , but not ␣ 1 , abolished AICAR-induced glucose uptake, whereas neither KO affected contraction-induced glucose uptake. AICAR and contraction increased ␣ 2 -and ␣ 1 -AMPK activity in wild type (WT) muscles. During AICAR stimulation, the remaining AMPK activity in KO muscles increased to the same level as in WT. During contraction, the remaining AMPK activity in ␣ 2 -KO muscles was elevated by 100% probably explained by a 2-3-fold increase in ␣ 1 -protein. In ␣ 1 -KO muscles, ␣ 2 -AMPK activity increased to similar levels as in WT. Both interventions increased total AMPK activity, as expressed by AMPK-P and ACC␤-P, in WT muscles. During AICAR stimulation, this was dramatically reduced in ␣ 2 -KO but not in ␣1-KO, whereas during contraction, both measurements were essentially similar to WT in both KO-muscles. The results show that ␣ 2 -AMPK is the main donor of basal and AICAR-stimulated AMPK activity and is responsible for AICAR-induced glucose uptake. In contrast, during contraction, the two ␣-isoforms seem to substitute for each other in terms of activity, which may explain the normal glucose uptake despite the lack of either ␣ 2 -or ␣ 1 -AMPK. Alternatively, neither ␣-isoform of AMPK is involved in contractioninduced muscle glucose uptake.The 5Ј-AMP-activated protein kinase (AMPK) 1 is a multisubstrate serine/threonine protein kinase that is ubiquitously expressed and functions as an intracellular fuel sensor activated by depletion of high energy phosphor compounds (1, 2). Activation of AMPK initiates a complex series of signaling events, causing an increase in uptake and oxidation of substrates for ATP synthesis concurrent with decreasing ATP consuming biosynthetic processes such as protein (3, 4), lipid (1), and glycogen synthesis (5, 6).Both human and rodent studies have shown that AMPK in skeletal muscle is activated during exercise in vivo (7-10) and during contraction in vitro (11-15) probably by several coinciding mechanisms. These involve decreased ATP/AMP and PCr/Cr ratios (16, 17), decreased pH (16), and reduction of muscle glycogen content (6, 15) and substrate delivery (18 -20). Therefore, it is tempting to ascribe a role for AMPK in muscle metabolism in response to exercise, and in particular investigators have hypothesized a role for AMPK in contraction-stimulated glucose uptake (11,13,21).AMPK may also be activated by treatment with the adenosine analogue 5-aminoimidazole-4-carboxamide-1-␤-4-ribofuranoside (AICAR) in rat, mouse, and human skeletal muscle in vitro (6,11,21,22) and in vivo in conscious rats (23). AICAR is...

Growth differentiation factor 15 (GDF15; also known as MIC-1) is a divergent member of the TGF-β superfamily and is associated with body-weight regulation in humans and rodents. However, the cognate receptor of GDF15 is unknown. Here we show that GDF15 binds specifically to GDNF family receptor α-like (GFRAL) with high affinity, and that GFRAL requires association with the coreceptor RET to elicit intracellular signaling in response to GDF15 stimulation. We also found that GDF15-mediated reductions in food intake and body weight of mice with obesity were abolished in GFRAL-knockout mice. We further found that GFRAL expression was limited to hindbrain neurons and not present in peripheral tissues, which suggests that GDF15-GFRAL-mediated regulation of food intake is by a central mechanism. Lastly, given that GDF15 did not increase energy expenditure in treated mice with obesity, the anti-obesity actions of the cytokine are likely driven primarily by a reduction in food intake.

AMP-activated protein kinase (AMPK) β1 or β2 subunits are required for assembling of AMPK heterotrimers and are important for regulating enzyme activity and cellular localization. In skeletal muscle, α2β2γ3-containing heterotrimers predominate. However, compensatory up-regulation and redundancy of AMPK subunits in wholebody AMPK α2, β2, and γ3 null mice has made it difficult to determine the physiological importance of AMPK in regulating muscle metabolism, because these models have normal mitochondrial content, contraction-stimulated glucose uptake, and insulin sensitivity. In the current study, we generated mice lacking both AMPK β1 and β2 isoforms in skeletal muscle (β1β2M-KO). β1β2M-KO mice are physically inactive and have a drastically impaired capacity for treadmill running that is associated with reductions in skeletal muscle mitochondrial content but not a fiber-type switch. Interestingly, young β1β2M-KO mice fed a control chow diet are not obese or insulin resistant but do have impaired contraction-stimulated glucose uptake. These data demonstrate an obligatory role for skeletal muscle AMPK in maintaining mitochondrial capacity and contraction-stimulated glucose uptake, findings that were not apparent in mice with single mutations or deletions in muscle α, β, or γ subunits.is an evolutionarily conserved stress-sensing kinase that controls energy metabolism and appetite by responding to nutrients and hormones (1). The regulation of AMPK activity depends on AMP and ADP regulated phosphorylation of the α catalytic subunit at T172 by the upstream kinases LKB1 and Ca 2+ /CaM-dependent protein kinase kinase (CaMKKβ; refs. 2 and 3). AMPK exists as a heterotrimer, consisting of an α catalytic subunit (α1, α2), a scaffolding β subunit (β1, β2) and a nucleotide-binding γ subunit (γ1, γ2, γ3) (1). The C-terminal of the β subunit contains a highly conserved α and γ subunit-binding sequence (SBS) that is required for the formation of a stable, active AMPK αβγ complex (4). We recently reported on the physiological effects of germline deletion of β1 (5) and β2 (6) isoforms in mice. We showed that β1 null mice have reduced AMPK α-subunit expression and activity in liver, adipose tissue and the hypothalamus (5). In contrast, AMPK β2 null mice have reduced AMPK activity in skeletal muscle, are aminoimidazole carboxamide ribonucleotide (AICAR) insensitive and have reduced exercise tolerance despite a greater than 50% increase in muscle β1 protein expression (6). The phenotype of β2 null mice was similar to that of mice lacking α2 (7) or γ3 (8) subunits or muscle-specific overexpression of an α2 kinase dead (KD) mutation (9, 10).During exercise, AMPK is activated in an intensity-dependent manner (for review, see ref. 11). Mice with reduced AMPK in muscle are exercise intolerant, an effect shown not to be due to cardiac impairments in AMPK (12-14). However, the cause for this reduction in exercise capacity remains largely unknown, because mitochondrial content and glucose uptake are not altered (6,7,10,12,(15)(16)(17) or onl...

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We tested the hypothesis that 5'AMP-activated protein kinase (AMPK) plays an important role in regulating the acute, exercise-induced activation of metabolic genes in skeletal muscle, which were dissected from whole-body alpha2- and alpha1-AMPK knockout (KO) and wild-type (WT) mice at rest, after treadmill running (90 min), and in recovery. Running increased alpha1-AMPK kinase activity, phosphorylation (P) of AMPK, and acetyl-CoA carboxylase (ACC)beta in alpha2-WT and alpha2-KO muscles and increased alpha2-AMPK kinase activity in alpha2-WT. In alpha2-KO muscles, AMPK-P and ACCbeta-P were markedly lower compared with alpha2-WT. However, in alpha1-WT and alpha1-KO muscles, AMPK-P and ACCbeta-P levels were identical at rest and increased similarly during exercise in the two genotypes. The alpha2-KO decreased peroxisome-proliferator-activated receptor gamma coactivator (PGC)-1alpha, uncoupling protein-3 (UCP3), and hexokinase II (HKII) transcription at rest but did not affect exercise-induced transcription. Exercise increased the mRNA content of PGC-1alpha, Forkhead box class O (FOXO)1, HKII, and pyruvate dehydrogenase kinase 4 (PDK4) similarly in alpha2-WT and alpha2-KO mice, whereas glucose transporter GLUT 4, carnitine palmitoyltransferase 1 (CPTI), lipoprotein lipase, and UCP3 mRNA were unchanged by exercise in both genotypes. CPTI mRNA was lower in alpha2-KO muscles than in alpha2-WT muscles at all time-points. In alpha1-WT and alpha1-KO muscles, running increased the mRNA content of PGC-1alpha and FOXO1 similarly. The alpha2-KO was associated with lower muscle adenosine 5'-triphosphate content, and the inosine monophosphate content increased substantially at the end of exercise only in alpha2-KO muscles. In addition, subcutaneous injection of 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR) increased the mRNA content of PGC-1alpha, HKII, FOXO1, PDK4, and UCP3, and alpha2-KO abolished the AICAR-induced increases in PGC-1alpha and HKII mRNA. In conclusion, KO of the alpha2- but not the alpha1-AMPK isoform markedly diminished AMPK activation during running. Nevertheless, exercise-induced activation of the investigated genes in mouse skeletal muscle was not impaired in alpha1- or alpha2-AMPK KO muscles. Although it cannot be ruled out that activation of the remaining alpha-isoform is sufficient to increase gene activation during exercise, the present data do not support an essential role of AMPK in regulating exercise-induced gene activation in skeletal muscle.