To examine the role of AMP-activated protein kinase (AMPK) in muscle glucose transport, we generated muscle-specific transgenic mice (TG) carrying cDNAs of inactive ␣2 (␣2i TG) and ␣1 (␣1i TG) catalytic subunits. Extensor digitorum longus (EDL) muscles from wild type and TG mice were isolated and subjected to a series of in vitro incubation experiments. In ␣2i TG mice basal ␣2 activity was barely detectable, whereas basal ␣1 activity was only partially reduced. Known AMPK stimuli including 5-aminoimidazole-4-carboxamide-1-␤-4-ribofuranoside (AICAR), rotenone (a Complex I inhibitor), dinitrophenol (a mitochondrial uncoupler), muscle contraction, and sorbitol (producing hyperosmolar shock) did not increase AMPK ␣2 activity in ␣2i TG mice, whereas ␣1 activation was attenuated by only 30 -50%. Glucose transport was measured in vitro using isolated EDL muscles from ␣2i TG mice. AICAR-and rotenone-stimulated glucose transport was fully inhibited in ␣2i TG mice; however, the lack of AMPK ␣2 activity had no effect on contraction-or sorbitol-induced glucose transport. Similar to these observations in vitro, contraction-stimulated glucose transport, assessed in vivo by 2-deoxy-D-[ 3 H]glucose incorporation into EDL, tibialis anterior, and gastrocnemius muscles, was normal in ␣2i TG mice. Thus, AMPK ␣2 activation is essential for some, but not all, insulin-independent glucose transport. Muscle contraction-and hyperosmolarity-induced glucose transport may be regulated by a redundant mechanism in which AMPK ␣2 is one of multiple signaling pathways.Recent reports suggest that AMP-activated protein kinase (AMPK), 2 a member of a metabolite-sensing protein kinase family, controls blood glucose homeostasis by regulating glucose transport in skeletal muscle and glucose production in the liver (1, 2). In skeletal muscle activation of AMPK by pharmacological stimulation and transient expression of an AMPK-active mutant increases glucose transport (3-6). AMPK also seems to play a role in enhancing muscle (7) and whole body (8) insulin sensitivity and responsiveness for glucose transport. Because skeletal muscle accounts for ϳ80% of disposal of an oral glucose load (9, 10) and because type 2 diabetes is associated with reduced muscle glucose disposal (11), AMPK may be critical in the control of metabolic homeostasis and perhaps the development of type 2 diabetes (12, 13). Not surprisingly, AMPK is now considered a drug target for the treatment of type 2 diabetes (14).AMPK is a serine/threonine kinase consisting of a catalytic ␣ subunit and regulatory ␤ and ␥ subunits (15-17). Different isoforms have been reported for each subunit (␣1 and ␣2, ␤1 and ␤2, ␥1, ␥2, and ␥3) with tissue-specific distribution. In skeletal muscle, ␣2 (18, 19), ␤2 (20, 21), and ␥1 (18) or ␥3 (22) are the major isoforms expressed and form the majority of AMPK heterotrimer complexes. AMPK is activated in response to decreases in intracellular ATP and concomitant increases in AMP, increasing the AMP:ATP ratio (15-17).It has long been believed that there are two m...

Leptin activates the long form of the leptin receptor (LRb) to control feeding and neuroendocrine function and thus regulate adiposity. While adiposity influences insulin sensitivity, leptin also regulates glucose homeostasis independently of energy balance. Disruption of the LRb/STAT3 signal in s/s mice results in hyperphagia, neuroendocrine dysfunction, and obesity similar to LRb null db/db mice. Insulin resistance and glucose intolerance are improved in s/s compared to db/db animals, however, suggesting that LRb/STAT3-independent signals may contribute to the regulation of glucose homeostasis by leptin. Indeed, caloric restriction normalized glycemic control in s/s animals, but db/db mice of similar weight and adiposity remained hyperglycemic. These differences in glucose homeostasis were not attributable to differences in insulin production between s/s and db/db animals but rather to decreased insulin resistance in s/s mice. Thus, in addition to LRb/STAT3-mediated adiposity signals, non-LRb/STAT3 leptin signals mediate an important adiposity-independent role in promoting glycemic control.

Leptin is an adipocyte-derived hormone that communicates the status of body energy stores to the brain to regulate feeding and energy balance. The inability of elevated leptin levels to adequately suppress feeding in obesity suggests attenuation of leptin action under these conditions; the activation of feedback circuits due to high leptin levels could contribute to this leptin resistance. Using cultured cells exogenously expressing the long form of the leptin receptor (LRb) or an erythropoietin receptor/LRb chimera, we show that chronic stimulation results in the attenuation of LRb signaling and the establishment of a state in which the receptor is refractory to reactivation. Mutation of LRb Tyr1138 (the site that recruits signal transducer and activator of transcription 3) alleviated this feedback inhibition, suggesting that signal transducer and activator of transcription 3 mediates the induction of a feedback inhibitor, such as suppressor of cytokine signaling 3 (SOCS3), during chronic LRb stimulation. Indeed, manipulation of the expression or activity of the LRb-binding tyrosine phosphatase, SH2-domain containing phosphatase-2, by overexpression of wild-type and dominant negative isoforms or RNA interference-mediated knockdown did not alter the attenuation of LRb signals. In contrast, SOCS3 overexpression repressed LRb signaling, whereas RNA interference-mediated knockdown of SOCS3 resulted in increased LRb signaling that was not attenuated during chronic ligand stimulation. These data suggest that Tyr1138 of LRb and SOCS3 represent major effector pathways for the feedback inhibition of LRb signaling. Furthermore, we show that mice expressing an LRb isoform mutant for Tyr1138 display increased activity of the leptin-dependent growth and immune axes, suggesting that Tyr1138-mediated feedback inhibition may regulate leptin sensitivity in vivo.

Secretion of leptin from adipose tissue communicates body energy status to the neuroendocrine system by activating the long form of the leptin receptor (LRb). Lack of leptin or LRb (as in db/db mice) results in obesity that stems from the combined effects of hyperphagia and decreased energy expenditure. We have previously generated mice in which LRb is replaced with a mutant LRb (LRb S1138 ) that specifically disrupts LRb3 STAT3 (signal transducer and activator of transcription-3) signaling; mice homozygous for this mutant (s/s) display increased feeding and are obese. We have now examined energy expenditure in s/s and db/db mice. Consistent with the increased lean body mass of s/s animals, locomotor activity and acute cold tolerance (partly a measure of shivering thermogenesis) in s/s mice were modestly but significantly improved compared with db/db mice, although they were decreased compared with wild-type mice. Total and resting metabolic rates were similarly depressed in s/s and db/db mice, however. Indeed, s/s and db/db mice display similar reductions in thyroid function and brown adipose tissue expression of uncoupling protein-1, which is regulated by sympathetic nervous system (SNS) tone. Thus, the LRb3 STAT3 signal is central to both the control of energy expenditure by leptin and the neuroendocrine regulation of the SNS and the thyroid axis. Diabetes 53:3067-3073, 2004 T he incidence of type 2 diabetes in industrialized nations has increased dramatically over the past two decades and continues to increase; much of this increase is attributable to the skyrocketing incidence of obesity in these populations (1,2). The recent identification of numerous regulators of appetite and energy expenditure has facilitated the molecular study of appetite and energy expenditure and potential mechanisms underlying obesity (3-6).One such regulator is leptin, the product of the obese (ob) gene (4,7). Leptin is a hormone that is secreted by adipose tissue to signal the status of body energy stores to the central nervous system (8). As a signal of energy sufficiency, adequate leptin levels suppress feeding and permit energy-costly neuroendocrine functions (4,9 -12). Conversely, negative energy balance decreases leptin levels, increasing the drive to feed and triggering neuroendocrine responses that limit energy expenditure (9). In addition to limiting energy-costly nonsurvival functions such as growth and reproduction, low leptin levels decrease minute-to-minute energy utilization by decreasing the metabolic rate of various tissues. Thus, the absence of leptin or the "long" signaling form of the leptin receptor (LRb) in ob/ob and db/db mice, respectively, results in a phenotype of obesity secondary to hyperphagia and decreased metabolic rate (due at least in part to hypothyroidism and decreased sympathetic nervous system [SNS] tone) plus altered nutrient partitioning, decreased growth, and infertility (4,5).Total momentary energy expenditure equals the sum of energy expended in physical work (e.g., volitional movement and ...

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Mice deficient in c-jun-NH(2)-terminal kinase 1 (JNK1) exhibit decreased fasting blood glucose and insulin levels, and protection against obesity-induced insulin resistance, suggesting increased glucose disposal into skeletal muscle. Thus, we assessed whether JNK1 deficiency enhances muscle glucose metabolism. Ex vivo insulin or contraction-induced muscle [(3)H]2-deoxyglucose uptake was not altered in JNK1 knockout mice, demonstrating that JNK1 does not regulate blood glucose levels via direct alterations in muscle. In vivo muscle [(3)H]2-deoxyglucose uptake in response to a glucose injection was also not enhanced by JNK1 deficiency, demonstrating that a circulating factor was not required to observe altered muscle glucose uptake in the knockout mice. JNK1 deficiency did not affect muscle glycogen levels or the protein expression of key molecules involved in glucose metabolism. This study is the first to directly demonstrate that enhanced skeletal muscle glucose metabolism does not underlie the beneficial effects of JNK1 deficiency in lean mice.