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The Mouse Metabolic Phenotyping Center (MMPC)Live Program was established in 2023 by the National Institute for Diabetes, Digestive and Kidney Diseases (NIDDK) at the National Institutes of Health (NIH) to advance biomedical research by providing the scientific community with standardized, high quality phenotyping services for mouse models of diabetes and obesity. Emerging as the next iteration of the MMPC Program which served the biomedical research community for 20 years (2001-2021), MMPCLive is designed as an outwardly-facing consortium of service cores that collaborate to provide reduced-cost consultation and metabolic, physiologic, and behavioral phenotyping tests on live mice for U.S. biomedical researchers. Four MMPCLive Centers located at universities around the country perform complex and often unique procedures in vivo on a fee for service basis, typically on mice shipped from the client or directly from a repository or vendor. Current areas of expertise include energy balance and body composition, insulin action and secretion, whole body carbohydrate and lipid metabolism, cardiovascular and renal function, food intake and behavior, microbiome and xenometabolism, and metabolic pathway kinetics. Additionally, an opportunity arose to reduce barriers to access and expand the diversity of the biomedical research workforce by establishing the VIBRANT Program. Directed at researchers historically underrepresented in the biomedical sciences, VIBRANT-eligible investigators have access to testing services, travel and career development awards, expert advice and experimental design consultation, and short internships to learn test technologies. Data derived from experiments run by the Centers belongs to the researchers submitting mice for testing which can be made publicly available and accessible from the MMPCLive database following publication. In addition to services, MMPCLive staff provide expertise and advice to researchers, develop and refine test protocols, engage in outreach activities, publish scientific and technical papers, and conduct educational workshops and training sessions to aid researchers in unraveling the heterogeneity of diabetes and obesity.
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Understanding the role of dentate gyrus (DG) mossy cells (MCs) in learning and memory has rapidly evolved due to increasingly precise methods for targeting MCs and for in vivo recording and activity manipulation in rodents. These studies have shown MCs are highly active in vivo, strongly remap to contextual manipulation, and that their inhibition or hyperactivation impairs pattern separation and location or context discrimination. Less well understood is how MC activity is modulated by neurohormonal mechanisms, which might differentially control the participation of MCs in cognitive functions during discrete states, such as hunger or satiety. In this study, we demonstrate that glucagon-like peptide-1 (GLP-1), a neuropeptide produced in the gut and the brain that regulates food consumption and hippocampal-dependent mnemonic function, might regulate MC function through expression of its receptor, GLP-1R. RNA-seq demonstrated that most, though not all, Glp1r in hippocampal principal neurons is expressed in MCs, and in situ hybridization revealed strong expression of Glp1r in hilar neurons. Glp1r-ires-Cre mice crossed with Ai14D reporter mice followed by co-labeling for the MC marker GluR2/3 revealed that almost all MCs in the ventral DG expressed Glp1r and that almost all Glp1r-expressing hilar neurons were MCs. However, only ~60% of dorsal DG MCs expressed Glp1r, and Glp1r was also expressed in small hilar neurons that were not MCs. Consistent with this expression pattern, peripheral administration of the GLP-1R agonist exendin-4 (5 µg/kg) increased cFos expression in ventral but not dorsal DG hilar neurons. Finally, whole-cell patch-clamp recordings from ventral MCs showed that bath application of exendin-4 (200 nM) depolarized MCs and increased action potential firing. Taken together, this study adds to known MC activity modulators a neurohormonal mechanism that may preferentially affect ventral DG physiology and may potentially be targetable by several GLP-1R pharmacotherapies already in clinical use.
Assuntos
Receptor do Peptídeo Semelhante ao Glucagon 1 , Fibras Musgosas Hipocampais , Animais , Camundongos , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Exenatida/farmacologia , Exenatida/metabolismo , Fibras Musgosas Hipocampais/fisiologia , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Hipocampo/metabolismo , Giro Denteado/metabolismoRESUMO
To identify new gene regulatory pathways controlling skeletal muscle energy metabolism, comparative studies were conducted on muscle-specific transgenic mouse lines expressing the nuclear receptors peroxisome proliferator-activated receptor α (PPARα; muscle creatine kinase [MCK]-PPARα) or PPARß/δ (MCK-PPARß/δ). MCK-PPARß/δ mice are known to have enhanced exercise performance, whereas MCK-PPARα mice perform at low levels. Transcriptional profiling revealed that the lactate dehydrogenase b (Ldhb)/Ldha gene expression ratio is increased in MCK-PPARß/δ muscle, an isoenzyme shift that diverts pyruvate into the mitochondrion for the final steps of glucose oxidation. PPARß/δ gain- and loss-of-function studies in skeletal myotubes demonstrated that PPARß/δ, but not PPARα, interacts with the exercise-inducible kinase AMP-activated protein kinase (AMPK) to synergistically activate Ldhb gene transcription by cooperating with myocyte enhancer factor 2A (MEF2A) in a PPARß/δ ligand-independent manner. MCK-PPARß/δ muscle was shown to have high glycogen stores, increased levels of GLUT4, and augmented capacity for mitochondrial pyruvate oxidation, suggesting a broad reprogramming of glucose utilization pathways. Lastly, exercise studies demonstrated that MCK-PPARß/δ mice persistently oxidized glucose compared with nontransgenic controls, while exhibiting supranormal performance. These results identify a transcriptional regulatory mechanism that increases capacity for muscle glucose utilization in a pattern that resembles the effects of exercise training.
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Glucose/metabolismo , Músculo Esquelético/metabolismo , Fatores de Regulação Miogênica/metabolismo , PPAR delta/metabolismo , Proteínas Quinases/metabolismo , Quinases Proteína-Quinases Ativadas por AMP , Animais , Células Cultivadas , Feminino , Lactato Desidrogenases/genética , Lactato Desidrogenases/metabolismo , Masculino , Camundongos , Músculo Esquelético/enzimologia , Oxirredução , PPAR alfa/metabolismo , Condicionamento Físico Animal , Ativação TranscricionalRESUMO
Long-acting glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) agonists (GLP-1RA), such as exendin-4 (Ex4), promote weight loss. On the basis of a newly discovered interaction between GLP-1 and oleoylethanolamide (OEA), we tested whether OEA enhances GLP-1RA-mediated anorectic signaling and weight loss. We analyzed the effect of GLP-1+OEA and Ex4+OEA on canonical GLP-1R signaling and other proteins/pathways that contribute to the hypophagic action of GLP-1RA (AMPK, Akt, mTOR, and glycolysis). We demonstrate that OEA enhances canonical GLP-1R signaling when combined with GLP-1 but not with Ex4. GLP-1 and Ex4 promote phosphorylation of mTOR pathway components, but OEA does not enhance this effect. OEA synergistically enhanced GLP-1- and Ex4-stimulated glycolysis but did not augment the hypophagic action of GLP-1 or Ex4 in lean or diet-induced obese (DIO) mice. However, the combination of Ex4+OEA promoted greater weight loss in DIO mice than Ex4 or OEA alone during a 7-day treatment. This was due in part to transient hypophagia and increased energy expenditure, phenotypes also observed in Ex4-treated DIO mice. Thus, OEA augments specific GLP-1RA-stimulated signaling but appears to work in parallel with Ex4 to promote weight loss in DIO mice. Elucidating cooperative mechanisms underlying Ex4+OEA-mediated weight loss could, therefore, be leveraged toward more effective obesity therapies.
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Fármacos Antiobesidade/farmacologia , Endocanabinoides/farmacologia , Exenatida/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Incretinas/farmacologia , Obesidade/tratamento farmacológico , Ácidos Oleicos/farmacologia , Redução de Peso/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Células CHO , Cricetulus , Dieta Hiperlipídica , Modelos Animais de Doenças , Quimioterapia Combinada , Comportamento Alimentar/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Glicólise/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Obesidade/metabolismo , Obesidade/fisiopatologia , Obesidade/psicologia , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
Pharmacological activation of the glucagon-like peptide-1 receptor (GLP-1R) in the ventromedial hypothalamus (VMH) reduces food intake. Here, we assessed whether suppression of food intake by GLP-1R agonists (GLP-1RA) in this region is dependent on AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR). We found that pharmacological inhibition of glycolysis, and thus activation of AMPK, in the VMH attenuates the anorectic effect of the GLP-1R agonist exendin-4 (Ex4), indicating that glucose metabolism and inhibition of AMPK are both required for this effect. Furthermore, we found that Ex4-mediated anorexia in the VMH involved mTOR but not acetyl-CoA carboxylase, two downstream targets of AMPK. We support this by showing that Ex4 activates mTOR signaling in the VMH and Chinese hamster ovary (CHO)-K1 cells. In contrast to the clear acute pharmacological impact of the these receptors on food intake, knockdown of the VMH Glp1r conferred no changes in energy balance in either chow- or high-fat-diet-fed mice, and the acute anorectic and glucose tolerance effects of peripherally dosed GLP-1RA were preserved. These results show that the VMH GLP-1R regulates food intake by engaging key nutrient sensors but is dispensable for the effects of GLP-1RA on nutrient homeostasis.
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Ingestão de Alimentos/fisiologia , Alimentos , Receptor do Peptídeo Semelhante ao Glucagon 1/fisiologia , Sensação/fisiologia , Núcleo Hipotalâmico Ventromedial/fisiologia , Acetil-CoA Carboxilase/metabolismo , Adenilato Quinase/metabolismo , Animais , Composição Corporal/efeitos dos fármacos , Células CHO , Cricetulus , Relação Dose-Resposta a Droga , Ingestão de Alimentos/efeitos dos fármacos , Exenatida , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Glicólise/efeitos dos fármacos , Homeostase/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/farmacologia , Sensação/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Peçonhas/farmacologia , Núcleo Hipotalâmico Ventromedial/metabolismoRESUMO
Sweet taste receptors (STRs) on the tongue mediate gustatory sweet sensing, but their expression in the gut, pancreas, and adipose tissue suggests a physiological contribution to whole body nutrient sensing and metabolism. However, little is known about the function and contribution of these sugar sensors during metabolic stress induced by overnutrition and subsequent obesity. Here, we investigated the effects of high-fat/low-carbohydrate (HF/LC) diet on glucose homeostasis and energy balance in mice with global disruption of the sweet taste receptor protein T1R2. We assessed body composition, energy balance, glucose homeostasis, and tissue-specific nutrient metabolism in T1R2 knockout (T1R2-KO) mice fed a HF/LC diet for 12 wk. HF/LC diet-fed T1R2-KO mice gained a similar amount of body mass as did WT mice, but had reduced fat mass and increased lean mass relative to WT mice. T1R2-KO mice were also hyperphagic and hyperactive. Ablation of the T1R2 sugar sensor protected mice from HF/LC diet-induced hyperinsulinemia and altered substrate utilization, including increased rates of glucose oxidation and decreased liver triglyceride (TG) accumulation, despite normal intestinal fat absorption. Finally, STRs (T1r2/T1r3) were upregulated in the adipose tissue of WT mice in response to HF/LC diet, and their expression positively correlated with fat mass and glucose intolerance. The chemosensory receptor T1R2, plays an important role in glucose homeostasis during diet-induced obesity through the regulation of yet to be identified molecular mechanisms that alter energy disposal and utilization in peripheral tissues.
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Glicemia/metabolismo , Composição Corporal/genética , Dieta com Restrição de Carboidratos , Dieta Hiperlipídica , Metabolismo Energético/genética , Intolerância à Glucose/genética , Obesidade/genética , Receptores Acoplados a Proteínas G/genética , Tecido Adiposo/metabolismo , Aminoácidos , Animais , Peso Corporal/genética , Cromo , Intolerância à Glucose/metabolismo , Homeostase , Hiperinsulinismo/metabolismo , Insulina/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Knockout , Ácidos Nicotínicos , Obesidade/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptores Acoplados a Proteínas G/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Receptor 3 Toll-Like/metabolismo , Triglicerídeos/metabolismo , Regulação para CimaRESUMO
The incretin hormone glucagon-like peptide-1 (Glp1) is cardioprotective in models of ischemia-reperfusion injury, myocardial infarction and gluco/lipotoxicity. Inflammation is a factor in these models, yet it is unknown whether Glp1 receptor (Glp1r) agonists are protective against cardiac inflammation. We tested the hypothesis that the Glp1r agonist Exendin-4 (Ex4) is cardioprotective in mice with cardiac-specific monocyte chemoattractant protein-1 overexpression. These MHC-MCP1 mice exhibit increased cardiac monocyte infiltration, endoplasmic reticulum (ER) stress, apoptosis, fibrosis and left ventricular dysfunction. Ex4 treatment for 8 weeks improved cardiac function and reduced monocyte infiltration, fibrosis and apoptosis in MHC-MCP1 mice. Ex4 enhanced expression of the ER chaperone glucose-regulated protein-78 (GRP78), decreased expression of the pro-apoptotic ER stress marker CCAAT/-enhancer-binding protein homologous protein (CHOP) and increased expression of the ER calcium regulator Sarco/Endoplasmic Reticulum Calcium ATPase-2a (SERCA2a). These findings suggest that the Glp1r is a viable target for treating cardiomyopathies associated with stimulation of pro-inflammatory factors.
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Cardiotônicos/farmacologia , Quimiocina CCL2/metabolismo , Miócitos Cardíacos/metabolismo , Peptídeos/farmacologia , Peçonhas/farmacologia , Disfunção Ventricular/tratamento farmacológico , Animais , Células Cultivadas , Quimiocina CCL2/genética , Avaliação Pré-Clínica de Medicamentos , Chaperona BiP do Retículo Endoplasmático , Exenatida , Expressão Gênica , Receptor do Peptídeo Semelhante ao Glucagon 1 , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Hipertrofia Ventricular Esquerda/metabolismo , Hipertrofia Ventricular Esquerda/fisiopatologia , Mediadores da Inflamação/metabolismo , Masculino , Camundongos Transgênicos , Receptores de Glucagon/agonistas , Volume Sistólico , Disfunção Ventricular/metabolismo , Disfunção Ventricular/fisiopatologiaRESUMO
Obesity is characterized by dysregulated homeostatic mechanisms resulting in positive energy balance, yet when this dysregulation occurs is unknown. We assessed the time course of alterations to behaviors promoting weight gain in male and female mice switched to obesogenic 60% or 45% high fat diet (HFD). Switching mice to obesogenic diets promotes transient bouts of hyperphagia during the first 2 weeks followed by persistent caloric hyperphagia. Energy expenditure increases but not sufficiently to offset increased caloric intake, resulting in a sustained net positive energy balance. Hyperphagia is associated with consumption of calorically larger meals (impaired satiation) more frequently (impaired satiety) particularly during the light-cycle. Running wheel exercise delays weight gain in 60% HFD-fed male mice by enhancing satiation and increasing energy expenditure. However, exercise effects on satiation are no longer apparent after 2 weeks, coinciding with weight gain. Thus, exposure to obesogenic diets engages homeostatic regulatory mechanisms for â¼2 weeks that ultimately fail, and consequent weight gain is characterized by impaired satiation and satiety. Insights into the etiology of obesity can be obtained by investigating changes to satiation and satiety mechanisms during the initial â¼2 weeks of HFD exposure. What is already known about this subject?: Obesity is associated with dysregulated homeostatic mechanisms.Increased caloric consumption contributes to obesity.Obese rodents tend to eat larger, more frequent meals. What are the new findings in your manuscript?: Exposure to obesogenic diets promotes transient attempts to maintain weight homeostasis.After â¼2 weeks, caloric hyperphagia exceeds increased energy expenditure, promoting weight gain.This is associated with consumption of larger, more frequent meals. How might your results change the direction of research or the focus of clinical practice?: Our findings suggest that molecular studies focusing on mechanisms that regulate meal size and frequency, particularly those engaged during the first â¼2 weeks of obesogenic diet feeding that eventually fail, can provide unique insight into the etiology of obesity.
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OBJECTIVE: Obesity is characterized by dysregulated homeostatic mechanisms resulting in positive energy balance; however, when this dysregulation occurs is unknown. We assessed the time course of alterations to behaviors promoting weight gain in male and female mice switched to an obesogenic high-fat diet (HFD). METHODS: Male and female C57BL/6J mice were housed in metabolic chambers and were switched from chow to a 60% or 45% HFD for 4 and 3 weeks, respectively. Food intake, meal patterns, energy expenditure (EE), and body weight were continuously measured. A separate cohort of male mice was switched from chow to a 60% HFD and was given access to locked or unlocked running wheels. RESULTS: Switching mice to obesogenic diets promotes transient bouts of hyperphagia during the first 2 weeks followed by persistent caloric hyperphagia. EE increases but not sufficiently enough to offset increased caloric intake, resulting in a sustained net positive energy balance. Hyperphagia is associated with consumption of calorically larger meals (impaired satiation) more frequently (impaired satiety), particularly during the light cycle. Running wheel exercise delays weight gain in male mice fed a 60% HFD by enhancing satiation and increasing EE. However, exercise effects on satiation are no longer apparent after 2 weeks, coinciding with weight gain. CONCLUSIONS: Exposure to obesogenic diets engages homeostatic regulatory mechanisms for ~2 weeks that ultimately fail, and consequent weight gain is characterized by impaired satiation and satiety. Insights into the etiology of obesity can be obtained by investigating changes to satiation and satiety mechanisms during the initial ~2 weeks of HFD exposure.
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Dieta Hiperlipídica , Ingestão de Energia , Metabolismo Energético , Comportamento Alimentar , Hiperfagia , Camundongos Endogâmicos C57BL , Obesidade , Aumento de Peso , Animais , Masculino , Camundongos , Dieta Hiperlipídica/efeitos adversos , Feminino , Obesidade/etiologia , Obesidade/metabolismo , Hiperfagia/etiologia , Comportamento Alimentar/fisiologia , Fatores de Tempo , Condicionamento Físico Animal , Saciação , Ingestão de Alimentos/fisiologiaRESUMO
During postnatal life, the adipocyte-derived hormone leptin is required for proper targeting of neural inputs to the paraventricular nucleus of the hypothalamus (PVH) and impacts the activity of neurons containing agouti-related peptide (AgRP) in the arcuate nucleus of the hypothalamus. Activity-dependent developmental mechanisms are known to play a defining role during postnatal organization of neural circuits, but whether leptin-mediated postnatal neuronal activity specifies neural projections to the PVH or impacts downstream connectivity is largely unexplored. Here, we blocked neuronal activity of AgRP neurons during a discrete postnatal period and evaluated development of AgRP inputs to defined regions in the PVH, as well as descending projections from PVH oxytocin neurons to the dorsal vagal complex (DVC) and assessed their dependence on leptin or postnatal AgRP neuronal activity. In leptin-deficient mice, AgRP inputs to PVH neurons were significantly reduced, as well as oxytocin-specific neuronal targeting by AgRP. Moreover, downstream oxytocin projections from the PVH to the DVC were also impaired, despite the lack of leptin receptors found on PVH oxytocin neurons. Blocking AgRP neuron activity specifically during early postnatal life reduced the density of AgRP inputs to the PVH, as well as the density of projections from PVH oxytocin neurons to the DVC, and these innervation deficits were associated with dysregulated autonomic function. These findings suggest that postnatal targeting of descending PVH oxytocin projections to the DVC requires leptin-mediated AgRP neuronal activity, and represents a novel activity-dependent mechanism for hypothalamic specification of metabolic circuitry, with consequences for autonomic regulation. Significance statement: Hypothalamic neural circuits maintain homeostasis by coordinating endocrine signals with autonomic responses and behavioral outputs to ensure that physiological responses remain in tune with environmental demands. The paraventricular nucleus of the hypothalamus (PVH) plays a central role in metabolic regulation, and the architecture of its neural inputs and axonal projections is a defining feature of how it receives and conveys neuroendocrine information. In adults, leptin regulates multiple aspects of metabolic physiology, but it also functions during development to direct formation of circuits controlling homeostatic functions. Here we demonstrate that leptin acts to specify the input-output architecture of PVH circuits through an activity-dependent, transsynaptic mechanism, which represents a novel means of sculpting neuroendocrine circuitry, with lasting effects on how the brain controls energy balance.
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Hyperglycemia-induced cardiomyocyte apoptosis contributes to diabetic cardiomyopathy. Glucagon-like peptide-1 (Glp1) receptor (Glp1r) agonists improve cardiac function and survival in response to ischemia-reperfusion and myocardial infarction. The present studies assessed whether Glp1r activation exerts direct cardioprotective effects in response to hyperglycemia. Treatment with the Glp1r agonist Exendin-4 attenuated apoptosis in neonatal rat ventricular cardiomyocytes cultured in high (33 mM) glucose. This protective effect was mimicked by the cAMP inducer forskolin. The Exendin-4 protective effect was blocked by the Glp1r antagonist Exendin(9-39) or the PKA antagonist H-89. Exendin-4 also protected cardiomyocytes from hydrogen peroxide (H2O2)-induced cell death. Cardiomyocyte protection by Exendin-4 was not due to reduced reactive oxygen species levels. Instead, Exendin-4 treatment reduced endoplasmic reticulum (ER) stress, demonstrated by decreased expression of glucose-regulated protein-78 (GRP78) and CCAT/enhancer-binding homologous protein (CHOP). Reduced ER stress was not due to activation of the unfolded protein response, indicating that Exendin-4 directly prevents ER stress. Exendin-4 treatment selectively protected cardiomyocytes from thapsigargin- but not tunicamycin-induced death. This suggests that Exendin-4 attenuates thapsigargin-mediated inhibition of the sarco/endoplasmic reticulum Ca(2+) ATPase-2a (SERCA2a). High glucose attenuates SERCA2a function by reducing SERCA2a mRNA and protein levels, but Exendin-4 treatment prevented this reduction. Exendin-4 treatment also enhanced phosphorylation of the SERCA2a regulator phospholamban (PLN), which would be expected to stimulate SERCA2a activity. In sum, Glp1r activation attenuates high glucose-induced cardiomyocyte apoptosis in association with decreased ER stress and markers of enhanced SERCA2a activity. These findings identify a novel mechanism whereby Glp1-based therapies could be used as treatments for diabetic cardiomyopathy.
Assuntos
Apoptose/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Miócitos Cardíacos/fisiologia , Peptídeos/farmacologia , Receptores de Glucagon/metabolismo , Peçonhas/farmacologia , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Células Cultivadas , Colforsina/farmacologia , Cardiomiopatias Diabéticas/metabolismo , Ativação Enzimática , Exenatida , Receptor do Peptídeo Semelhante ao Glucagon 1 , Glucose/metabolismo , Proteínas de Choque Térmico HSP70 , Proteínas de Choque Térmico/metabolismo , Peróxido de Hidrogênio/farmacologia , Hiperglicemia/metabolismo , Hipoglicemiantes/farmacologia , Isoquinolinas/farmacologia , Proteínas de Membrana , Estresse Oxidativo/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/biossíntese , Ratos , Receptores de Glucagon/agonistas , Receptores de Glucagon/antagonistas & inibidores , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/antagonistas & inibidores , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/genética , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Sulfonamidas/farmacologia , Tapsigargina/farmacologia , Fator de Transcrição CHOP/biossíntese , Fator de Transcrição CHOP/metabolismo , Tunicamicina/farmacologia , Resposta a Proteínas não DobradasRESUMO
Glucagon-like peptide-1 (GLP-1) suppresses food intake via activation of a central (i.e., brain) GLP-1 receptor (GLP-1R). Central AMP-activated protein kinase (AMPK) is a nutrient-sensitive regulator of food intake that is inhibited by anorectic signals. The anorectic effect elicited by hindbrain GLP-1R activation is attenuated by the AMPK stimulator AICAR. This suggests that central GLP-1R activation suppresses food intake via inhibition of central AMPK. The present studies examined the mechanism(s) by which central GLP-1R activation inhibits AMPK. Supporting previous findings, AICAR attenuated the anorectic effect elicited by intracerebroventricular (icv) administration of the GLP-1R agonist exendin-4 (Ex-4). We demonstrate that Ex-4 stimulates glycolysis and suppresses AMPK phosphorylation in a glucose-dependent manner in hypothalamic GT1-7 cells. This suggests that inhibition of AMPK and food intake by Ex-4 requires central glucose metabolism. Supporting this, the glycolytic inhibitor 2-deoxyglucose (2-DG) attenuated the anorectic effect of Ex-4. However, icv glucose did not enhance the suppression of food intake by Ex-4. AICAR had no effect on Ex-4-mediated reduction in locomotor activity. We also tested whether other carbohydrates affect the anorectic response to Ex-4. Intracerebroventricular pretreatment with the sucrose metabolite fructose, an AMPK activator, attenuated the anorectic effect of Ex-4. This potentially explains the increased food intake observed in sucrose-fed mice. In summary, we propose a model whereby activation of the central GLP-1R reduces food intake via glucose metabolism-dependent inhibition of central AMPK. We also suggest that fructose stimulates food intake by impairing central GLP-1R action. This has significant implications given the correlation between sugar consumption and obesity.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Anorexia/metabolismo , Regulação do Apetite/fisiologia , Frutose/metabolismo , Glucose/metabolismo , Hipotálamo/metabolismo , Receptores de Glucagon/metabolismo , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/efeitos dos fármacos , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacologia , Animais , Antimetabólitos/farmacologia , Regulação do Apetite/efeitos dos fármacos , Linhagem Celular , Desoxiglucose/farmacologia , Ingestão de Líquidos/efeitos dos fármacos , Ingestão de Líquidos/fisiologia , Ingestão de Alimentos/efeitos dos fármacos , Ingestão de Alimentos/fisiologia , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/fisiologia , Exenatida , Frutose/administração & dosagem , Receptor do Peptídeo Semelhante ao Glucagon 1 , Hipoglicemiantes/farmacologia , Hipotálamo/efeitos dos fármacos , Masculino , Camundongos , Camundongos Knockout , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Peptídeos/farmacologia , Receptores de Glucagon/efeitos dos fármacos , Receptores de Glucagon/genética , Ribonucleotídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Peçonhas/farmacologiaRESUMO
The canonical target of the glucagon-like peptide-1 receptor (GLP-1R), Protein Kinase A (PKA), has been shown to stimulate mechanistic Target of Rapamycin Complex 1 (mTORC1) by phosphorylating the mTOR-regulating protein Raptor at Ser791 following ß-adrenergic stimulation. The objective of these studies is to test whether GLP-1R agonists similarly stimulate mTORC1 via PKA phosphorylation of Raptor at Ser791 and whether this contributes to the weight loss effect of the therapeutic GLP-1R agonist liraglutide. We measured phosphorylation of the mTORC1 signaling target ribosomal protein S6 in Chinese Hamster Ovary cells expressing GLP-1R (CHO-Glp1r) treated with liraglutide in combination with PKA inhibitors. We also assessed liraglutide-mediated phosphorylation of the PKA substrate RRXS*/T* motif in CHO-Glp1r cells expressing Myc-tagged wild-type (WT) Raptor or a PKA-resistant (Ser791Ala) Raptor mutant. Finally, we measured the body weight response to liraglutide in WT mice and mice with a targeted knock-in of PKA-resistant Ser791Ala Raptor. Liraglutide increased phosphorylation of S6 and the PKA motif in WT Raptor in a PKA-dependent manner but failed to stimulate phosphorylation of the PKA motif in Ser791Ala Raptor in CHO-Glp1r cells. Lean Ser791Ala Raptor knock-in mice were resistant to liraglutide-induced weight loss but not setmelanotide-induced (melanocortin-4 receptor-dependent) weight loss. Diet-induced obese Ser791Ala Raptor knock-in mice were not resistant to liraglutide-induced weight loss; however, there was weight-dependent variation such that there was a tendency for obese Ser791Ala Raptor knock-in mice of lower relative body weight to be resistant to liraglutide-induced weight loss compared to weight-matched controls. Together, these findings suggest that PKA-mediated phosphorylation of Raptor at Ser791 contributes to liraglutide-induced weight loss.
Assuntos
Receptor do Peptídeo Semelhante ao Glucagon 1 , Liraglutida , Proteína Regulatória Associada a mTOR , Redução de Peso , Animais , Cricetinae , Camundongos , Células CHO , Cricetulus , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Liraglutida/farmacologia , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Obesidade/tratamento farmacológico , Fosforilação , Proteína Regulatória Associada a mTOR/metabolismoRESUMO
Negative regulation of exocytosis from secretory cells is accomplished through inhibitory signals from Gi/o GPCRs by Gßγ subunit inhibition of 2 mechanisms: decreased calcium entry and direct interaction of Gßγ with soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (SNARE) plasma membrane fusion machinery. Previously, we disabled the second mechanism with a SNAP25 truncation (SNAP25Δ3) that decreased Gßγ affinity for the SNARE complex, leaving exocytotic fusion and modulation of calcium entry intact and removing GPCR-Gßγ inhibition of SNARE-mediated exocytosis. Here, we report substantial metabolic benefit in mice carrying this mutation. Snap25Δ3/Δ3 mice exhibited enhanced insulin sensitivity and beiging of white fat. Metabolic protection was amplified in Snap25Δ3/Δ3 mice challenged with a high-fat diet. Glucose homeostasis, whole-body insulin action, and insulin-mediated glucose uptake into white adipose tissue were improved along with resistance to diet-induced obesity. Metabolic protection in Snap25Δ3/Δ3 mice occurred without compromising the physiological response to fasting or cold. All metabolic phenotypes were reversed at thermoneutrality, suggesting that basal autonomic activity was required. Direct electrode stimulation of sympathetic neuron exocytosis from Snap25Δ3/Δ3 inguinal adipose depots resulted in enhanced and prolonged norepinephrine release. Thus, the Gßγ-SNARE interaction represents a cellular mechanism that deserves further exploration as an additional avenue for combating metabolic disease.
Assuntos
Subunidades beta da Proteína de Ligação ao GTP , Subunidades gama da Proteína de Ligação ao GTP , Insulinas , Camundongos , Animais , Cálcio/metabolismo , Subunidades beta da Proteína de Ligação ao GTP/genética , Subunidades beta da Proteína de Ligação ao GTP/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/genética , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Exocitose/fisiologia , Proteínas SNARE/genética , Dieta , Obesidade/genética , Adipócitos/metabolismo , Insulinas/metabolismo , Insulina/metabolismoRESUMO
Glucagon-like peptide-1 receptor (GLP-1R) agonists and fibroblast growth factor 21 (FGF21) confer similar metabolic benefits. Studies report that GLP-1RA induce FGF21. Here, we investigated the mechanisms engaged by the GLP-1R agonist liraglutide to increase FGF21 levels and the metabolic relevance of liraglutide-induced FGF21. We show that liraglutide increases FGF21 levels via neuronal GLP-1R activation. We also demonstrate that lack of liver Fgf21 expression confers partial resistance to liraglutide-induced weight loss. Since FGF21 reduces carbohydrate intake, we tested whether the contribution of FGF21 to liraglutide-induced weight loss is dependent on dietary carbohydrate content. In control and liver Fgf21 knockout (Liv Fgf21 -/- ) mice fed calorically matched diets with low- (LC) or high-carbohydrate (HC) content, we found that only HC-fed Liv Fgf21 -/- mice were resistant to liraglutide-induced weight loss. Similarly, liraglutide-induced weight loss was partially impaired in Liv Fgf21 -/- mice fed a high-fat, high-sugar (HFHS) diet. Lastly, we show that loss of neuronal ß-klotho expression also diminishes liraglutide-induced weight loss in mice fed a HC or HFHS diet, indicating that FGF21 mediates liraglutide-induced weight loss via neuronal FGF21 action. Our findings support a novel role for a GLP-1R-FGF21 axis in regulating body weight in the presence of high dietary carbohydrate content.
RESUMO
Muscle fitness and mass deteriorate under the conditions of obesity and aging for reasons yet to be fully elucidated. Herein, we describe a novel pathway linking peripheral nutrient sensing and skeletal muscle function through the sweet taste receptor TAS1R2 and the involvement of ERK2-PARP1-NAD signaling axis. Muscle-specific deletion of TAS1R2 (mKO) in mice produced elevated NAD levels due to suppressed PARP1 activity, improved mitochondrial function, increased muscle mass and strength, and prolonged running endurance. Deletion of TAS1R2 in obese or aged mice also ameliorated the decline in muscle mass and fitness arising from these conditions. Remarkably, partial loss-of-function of TAS1R2 (rs35874116) in older, obese humans recapitulated the healthier muscle phenotype displayed by mKO mice in response to exercise training. Our findings show that inhibition of the TAS1R2 signaling in skeletal muscle is a promising therapeutic approach to preserve muscle mass and function.
RESUMO
OBJECTIVE: Glucagon-like peptide-1 receptor (GLP-1R) agonists (GLP-1RA) and fibroblast growth factor-21 (FGF21) confer similar metabolic benefits. GLP-1RA induce FGF21, leading us to investigate mechanisms engaged by the GLP-1RA liraglutide to increase FGF21 levels and the metabolic relevance of liraglutide-induced FGF21. METHODS: Circulating FGF21 levels were measured in fasted male C57BL/6J, neuronal GLP-1R knockout, ß-cell GLP-1R knockout, and liver peroxisome proliferator-activated receptor alpha knockout mice treated acutely with liraglutide. To test the metabolic relevance of liver FGF21 in response to liraglutide, chow-fed control and liver Fgf21 knockout (LivFgf21-/-) mice were treated with vehicle or liraglutide in metabolic chambers. Body weight and composition, food intake, and energy expenditure were measured. Since FGF21 reduces carbohydrate intake, we measured body weight in mice fed matched diets with low- (LC) or high-carbohydrate (HC) content and in mice fed a high-fat, high-sugar (HFHS) diet. This was done in control and LivFgf21-/- mice and in mice lacking neuronal ß-klotho (Klb) expression to disrupt brain FGF21 signaling. RESULTS: Liraglutide increases FGF21 levels independently of decreased food intake via neuronal GLP-1R activation. Lack of liver Fgf21 expression confers resistance to liraglutide-induced weight loss due to attenuated reduction of food intake in chow-fed mice. Liraglutide-induced weight loss was impaired in LivFgf21-/- mice when fed HC and HFHS diets but not when fed a LC diet. Loss of neuronal Klb also attenuated liraglutide-induced weight loss in mice fed HC or HFHS diets. CONCLUSIONS: Our findings support a novel role for a GLP-1R-FGF21 axis in regulating body weight in a dietary carbohydrate-dependent manner.
Assuntos
Receptor do Peptídeo Semelhante ao Glucagon 1 , Liraglutida , Animais , Masculino , Camundongos , Carboidratos , Dieta Hiperlipídica , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Liraglutida/farmacologia , Camundongos Endogâmicos C57BL , Redução de PesoRESUMO
Glucagon-like peptide-1 (GLP-1) receptor knockout (Glp1r(-/-)) mice exhibit impaired hepatic insulin action. High fat (HF)-fed Glp1r(-/-) mice exhibit improved, rather than the expected impaired, hepatic insulin action. This is due to decreased lipogenic gene expression and triglyceride accumulation. The present studies overcome these secondary adaptations by acutely modulating GLP-1R action in HF-fed wild-type mice. The central GLP-1R was targeted given its role as a regulator of hepatic insulin action. We hypothesized that acute inhibition of the central GLP-1R impairs hepatic insulin action beyond the effects of HF feeding. We further hypothesized that activation of the central GLP-1R improves hepatic insulin action in HF-fed mice. Insulin action was assessed in conscious, unrestrained mice using the hyperinsulinemic euglycemic clamp. Mice received intracerebroventricular (icv) infusions of artificial cerebrospinal fluid, GLP-1, or the GLP-1R antagonist exendin-9 (Ex-9) during the clamp. Intracerebroventricular Ex-9 impaired the suppression of hepatic glucose production by insulin, whereas icv GLP-1 improved it. Neither treatment affected tissue glucose uptake. Intracerebroventricular GLP-1 enhanced activation of hepatic Akt and suppressed hypothalamic AMP-activated protein kinase. Central GLP-1R activation resulted in lower hepatic triglyceride levels but did not affect muscle, white adipose tissue, or plasma triglyceride levels during hyperinsulinemia. In response to oral but not intravenous glucose challenges, activation of the central GLP-1R improved glucose tolerance. This was associated with higher insulin levels. Inhibition of the central GLP-1R had no effect on oral or intravenous glucose tolerance. These results show that inhibition of the central GLP-1R deteriorates hepatic insulin action in HF-fed mice but does not affect whole body glucose homeostasis. Contrasting this, activation of the central GLP-1R improves glucose homeostasis in HF-fed mice by increasing insulin levels and enhancing hepatic insulin action.
Assuntos
Peptídeo 1 Semelhante ao Glucagon/metabolismo , Hipotálamo/metabolismo , Resistência à Insulina , Insulina/metabolismo , Fígado/metabolismo , Pâncreas/metabolismo , Receptores de Glucagon/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Dieta Hiperlipídica/efeitos adversos , Peptídeo 1 Semelhante ao Glucagon/administração & dosagem , Receptor do Peptídeo Semelhante ao Glucagon 1 , Técnica Clamp de Glucose , Glicogenólise/efeitos dos fármacos , Hipotálamo/efeitos dos fármacos , Hipotálamo/enzimologia , Infusões Intraventriculares , Insulina/sangue , Secreção de Insulina , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/agonistas , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/metabolismo , Especificidade de Órgãos , Pâncreas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Glucagon/agonistas , Receptores de Glucagon/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacosRESUMO
OBJECTIVE: While stress typically reduces caloric intake (hypophagia) in chow-fed rodents, presentation of palatable, high calorie substances during stress can increase caloric consumption (i.e. "comfort feeding") and promote obesity. However, little is known about how obesity itself affects feeding behavior in response to stress and the mechanisms that can influence stress-associated feeding in the context of obesity. METHODS: We assessed food intake and other metabolic parameters in lean and obese male and female mice following acute restraint stress. We also measured real-time activity of glucagon-like peptide-1 (Glp1) receptor (Glp1r)-expressing neurons in the dorsal lateral septum (dLS) during stress in lean and obese mice using fiber photometry. Glp1r activation in various brain regions, including the dLS, promotes hypophagia in response to stress. Finally, we used inhibitory Designer Receptors Activated Exclusively by Designer Drugs (DREADDs) to test whether activation of Glp1r-expressing neurons in the LS is required for stress-induced hypophagia. RESULTS: Lean male mice display the expected hypophagic response following acute restraint stress, but obese male mice are resistant to this acute stress-induced hypophagia. Glp1r-positive neurons in the dLS are robustly activated during acute restraint stress in lean but not in obese male mice. This raises the possibility that activation of dLS Glp1r neurons during restraint stress contributes to subsequent hypophagia. Supporting this, we show that chemogenetic inhibition of LS Glp1r neurons attenuates acute restraint stress hypophagia in male mice. Surprisingly, we show that both lean and obese female mice are resistant to acute restraint stress-induced hypophagia and activation of dLS Glp1r neurons. CONCLUSIONS: These results suggest that dLS Glp1r neurons contribute to the hypophagic response to acute restraint stress in male mice, but not in female mice, and that obesity disrupts this response in male mice. Broadly, these findings show sexually dimorphic mechanisms and feeding behaviors in lean vs. obese mice in response to acute stress.
Assuntos
Dieta Hiperlipídica , Receptor do Peptídeo Semelhante ao Glucagon 1 , Animais , Dieta Hiperlipídica/efeitos adversos , Feminino , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Masculino , Camundongos , Camundongos Obesos , Neurônios/metabolismo , Obesidade/metabolismoRESUMO
Diabetic patients show elevated plasma IL18 concentrations. IL18 has two receptors: the IL18 receptor (IL18r) and the Na-Cl co-transporter (NCC). Here, we report that IL18 is expressed on islet α cells, NCC on ß cells, and IL18r on acinar cells in human and mouse pancreases. The deficiency of these receptors reduces islet size, ß cell proliferation, and insulin secretion but increases ß cell apoptosis and exocrine macrophage accumulation after diet-induced glucose intolerance or streptozotocin-induced hyperglycemia. Together with the glucagon-like peptide-1 (GLP1), IL18 uses the NCC and GLP1 receptors on ß cells to trigger ß cell development and insulin secretion. IL18 also uses the IL18r on acinar cells to block hyperglycemic pancreas macrophage expansion. The ß cell-selective depletion of the NCC or acinar-cell-selective IL18r depletion reduces glucose tolerance and insulin sensitivity with impaired ß cell proliferation, enhanced ß cell apoptosis and macrophage expansion, and inflammation in mouse hyperglycemic pancreas. IL18 uses NCC, GLP1r, and IL18r to maintain islet ß cell function and homeostasis.