Metformin acts in the gut and induces gut-liver crosstalk.

Metformin is the most prescribed drug for DM2, but its site and mechanism of action are still not well established. Here, we investigated the effects of metformin on basolateral intestinal glucose uptake (BIGU), and its consequences on hepatic glucose production (HGP). In diabetic patients and mice, the primary site of metformin action was the gut, increasing BIGU, evaluated through PET-CT. In mice and CaCo2 cells, this increase in BIGU resulted from an increase in GLUT1 and GLUT2, secondary to ATF4 and AMPK. In hyperglycemia, metformin increased the lactate (reducing pH and bicarbonate in portal vein) and acetate production in the gut, modulating liver pyruvate carboxylase, MPC1/2, and FBP1, establishing a gut-liver crosstalk that reduces HGP. In normoglycemia, metformin-induced increases in BIGU is accompanied by hypoglycemia in the portal vein, generating a counter-regulatory mechanism that avoids reductions or even increases HGP. In summary, metformin increases BIGU and through gut-liver crosstalk influences HGP.

Resistance exercise attenuates IKKε phosphorylation and hepatic fat accumulation of obese mice.

Obesity is associated with low-grade inflammation and disturbances in hepatic metabolism. This study aimed to investigate the effects of resistance exercise on inflammatory signalling related to IκB kinase (IKK) ɛ protein (IKKɛ) and on hepatic fat accumulation in obese mice. Male Swiss mice were distributed into three groups: control (CTL) fed with standard chow; obese (OB) mice induced by a high-fat diet (HFD); obese exercised (OB + RE) mice fed with HFD and submitted to a resistance exercise training. The resistance exercise training protocol consisted of 20 sets/3 ladder climbs for 8 weeks, three times/week on alternate days. The training overload was equivalent to 70% of the maximum load supported by the rodent. Assays were performed to evaluate weight gain, hepatic fat content, fasting glucose, insulin sensitivity, IKKɛ phosphorylation and proteins related to insulin signalling and lipogenesis in the liver. Mice that received the high-fat diet showed greater adiposity, impaired insulin sensitivity, increased fasting glucose and increased hepatic fat accumulation. These results were accompanied by an increase in IKKɛ phosphorylation and lipogenesis-related proteins such as cluster of differentiation 36 (CD36) and fatty acid synthase (FAS) in the liver of obese mice. In contrast, exercised mice showed lower body weight and adiposity evolution throughout the experiment. In addition, resistance exercise suppressed the effects of the high-fat diet by reducing IKKɛ phosphorylation and hepatic fat content. In conclusion, resistance exercise training improves hepatic fat metabolism and glycaemic homeostasis, which are, at least in part, linked to the anti-inflammatory effect of reduced IKKɛ phosphorylation in the liver of obese mice.

Growth hormone enhances the recovery of hypoglycemia via ventromedial hypothalamic neurons.

Growth hormone (GH) is secreted during hypoglycemia, and GH-responsive neurons are found in brain areas containing glucose-sensing neurons that regulate the counter-regulatory response (CRR). However, whether GH modulates the CRR to hypoglycemia via specific neuronal populations is currently unknown. Mice carrying ablation of GH receptor (GHR) either in leptin receptor (LepR)- or steroidogenic factor-1 (SF1)-expressing cells were studied. We also investigated the importance of signal transducer and activator of transcription 5 (STAT5) signaling in SF1 cells for the CRR. GHR ablation in LepR cells led to impaired capacity to recover from insulin-induced hypoglycemia and to a blunted CRR caused by 2-deoxy-d-glucose (2DG) administration. GHR inactivation in SF1 cells, which include ventromedial hypothalamic neurons, also attenuated the CRR. The reduced CRR was prevented by parasympathetic blockers. Additionally, infusion of 2DG produced an abnormal hyperactivity of parasympathetic preganglionic neurons, whereas the 2DG-induced activation of anterior bed nucleus of the stria terminalis neurons was reduced in mice without GHR in SF1 cells. Mice carrying ablation of Stat5a/b genes in SF1 cells showed no defects in the CRR. In summary, GHR expression in SF1 cells is required for a normal CRR, and these effects are largely independent of STAT5 pathway.-Furigo, I. C., de Souza, G. O., Teixeira, P. D. S., Guadagnini, D., Frazão, R., List, E. O., Kopchick, J. J., Prada, P. O., Donato, J., Jr. Growth hormone enhances the recovery of hypoglycemia via ventromedial hypothalamic neurons.

PAFR in adipose tissue macrophages is associated with anti-inflammatory phenotype and metabolic homoeostasis.

Metabolic dysfunction is associated with adipose tissue inflammation and macrophage infiltration. PAFR (platelet-activating factor receptor) is expressed in several cell types and binds to PAF (platelet-activating factor) and oxidized phospholipids. Engagement of PAFR in macrophages drives them towards the anti-inflammatory phenotype. In the present study, we investigated whether genetic deficiency of PAFR affects the phenotype of ATMs (adipose tissue macrophages) and its effect on glucose and insulin metabolism. PARFKO (PAFR-knockout) and WT (wild-type) mice were fed on an SD (standard diet) or an HFD (high-fat diet). Glucose and insulin tolerance tests were performed by blood monitoring. ATMs were evaluated by FACS for phenotypic markers. Gene and protein expression was investigated by real-time reverse transcription-quantitative PCR and Western blotting respectively. Results showed that the epididymal adipose tissue of PAFRKO mice had increased gene expression of Ccr7, Nos2, Il6 and Il12, associated with pro-inflammatory mediators, and reduced expression of the anti-inflammatory Il10. Moreover, the adipose tissue of PAFRKO mice presented more pro-inflammatory macrophages, characterized by an increased frequency of F4/80(+)CD11c(+) cells. Blood monocytes of PAFRKO mice also exhibited a pro-inflammatory phenotype (increased frequency of Ly6C(+) cells) and PAFR ligands were detected in the serum of both PAFRKO and WT mice. Regarding metabolic parameters, compared with WT, PAFRKO mice had: (i) higher weight gain and serum glucose concentration levels; (ii) decreased insulin-stimulated glucose disappearance; (iii) insulin resistance in the liver; (iv) increased expression of Ldlr in the liver. In mice fed on an HFD, some of these changes were potentiated, particularly in the liver. Thus it seems that endogenous ligands of PAFR are responsible for maintaining the anti-inflammatory profile of blood monocytes and ATMs under physiological conditions. In the absence of PAFR signalling, monocytes and macrophages acquire a pro-inflammatory phenotype, resulting in adipose tissue inflammation and metabolic dysfunction.

Gut microbiota is a key modulator of insulin resistance in TLR 2 knockout mice.

Environmental factors and host genetics interact to control the gut microbiota, which may have a role in the development of obesity and insulin resistance. TLR2-deficient mice, under germ-free conditions, are protected from diet-induced insulin resistance. It is possible that the presence of gut microbiota could reverse the phenotype of an animal, inducing insulin resistance in an animal genetically determined to have increased insulin sensitivity, such as the TLR2 KO mice. In the present study, we investigated the influence of gut microbiota on metabolic parameters, glucose tolerance, insulin sensitivity, and signaling of TLR2-deficient mice. We investigated the gut microbiota (by metagenomics), the metabolic characteristics, and insulin signaling in TLR2 knockout (KO) mice in a non-germ free facility. Results showed that the loss of TLR2 in conventionalized mice results in a phenotype reminiscent of metabolic syndrome, characterized by differences in the gut microbiota, with a 3-fold increase in Firmicutes and a slight increase in Bacteroidetes compared with controls. These changes in gut microbiota were accompanied by an increase in LPS absorption, subclinical inflammation, insulin resistance, glucose intolerance, and later, obesity. In addition, this sequence of events was reproduced in WT mice by microbiota transplantation and was also reversed by antibiotics. At the molecular level the mechanism was unique, with activation of TLR4 associated with ER stress and JNK activation, but no activation of the IKKß-IκB-NFκB pathway. Our data also showed that in TLR2 KO mice there was a reduction in regulatory T cell in visceral fat, suggesting that this modulation may also contribute to the insulin resistance of these animals. Our results emphasize the role of microbiota in the complex network of molecular and cellular interactions that link genotype to phenotype and have potential implications for common human disorders involving obesity, diabetes, and even other immunological disorders.

Air pollution accelerates the development of obesity and Alzheimer's disease: the role of leptin and inflammation - a mini-review.

Air pollution is an urgent concern linked to numerous health problems in low- and middle-income countries, where 92% of air pollution-related deaths occur. Particulate matter 2.5 (PM2.5) is the most harmful component of air pollutants, increasing inflammation and changing gut microbiota, favoring obesity, type 2 diabetes, and Alzheimer's Disease (AD). PM2.5 contains lipopolysaccharides (LPS), which can activate the Toll-like receptor 4 (TLR4) signaling pathway. This pathway can lead to the release of pro-inflammatory markers, including interleukins, and suppressor of cytokine signaling-3 (SOCS3), which inhibits leptin action, a hormone that keeps the energy homeostasis. Leptin plays a role in preventing amyloid plaque deposition and hyperphosphorylation of tau-protein (p-tau), mechanisms involved in the neurodegeneration in AD. Approximately 50 million people worldwide are affected by dementia, with a significant proportion living in low-and middle-income countries. This number is expected to triple by 2050. This mini-review focuses on the potential impact of PM2.5 exposure on the TLR4 signaling pathway, its contribution to leptin resistance, and dysbiosis that exacerbates the link between obesity and AD.

Multiple metabolic signals in the CeA regulate feeding: The role of AMPK.

BACKGROUND: The central nucleus of the amygdala (CeA) is part of the dopaminergic reward system and controls energy balance. Recently, a cluster of neurons was identified as responsive to the orexigenic effect of ghrelin and fasting. However, the signaling pathway by which ghrelin and fasting induce feeding is unknown. AMP-activated protein kinase (AMPK) is a cellular energy sensor, and its Thr172 phosphorylation (AMPKThr172) in the mediobasal hypothalamus regulates food intake. However, whether the expression and activation of AMPK in CeA could be one of the intracellular signaling activated in response to ghrelin and fasting eliciting food intake is unknown. AIM: To evaluate the activation of AMPK into CeA in response to ghrelin, fasting, and 2-deoxy-D-glucose (2DG) and whether feeding accompanied these changes. In addition, to investigate whether the inhibition of AMPK into CeA could decrease food intake. METHODS: On a chow diet, eight-week-old Wistar male rats were stereotaxically implanted with a cannula in the CeA to inject several modulators of AMPKα1/2Thr172 phosphorylation, and we performed physiological and molecular assays. KEY FINDINGS: Fasting increased, and refeeding reduced AMPKThr172 in the CeA. Intra-CeA glucose injection decreased feeding, whereas injection of 2DG, a glucoprivation inductor, in the CeA, increased food intake and blood glucose, despite faint increases in AMPKThr172. Intra-CeA ghrelin injection increased food intake and AMPKThr172. To further confirm the role of AMPK in the CeA, chronic injection of Melanotan II (MTII) in CeA reduced body mass and food intake over seven days together with a slight decrease in AMPKThr172. SIGNIFICANCE: Our findings identified that AMPK might be part of the signaling machinery in the CeA, which responds to nutrients and hormones contributing to feeding control. The results can contribute to understanding the pathophysiological mechanisms of altered feeding behavior/consumption, such as binge eating of caloric-dense, palatable food.

Development of a genetically encoded sensor for probing endogenous nociceptin opioid peptide release.

Nociceptin/orphanin-FQ (N/OFQ) is a recently appreciated critical opioid peptide with key regulatory functions in several central behavioral processes including motivation, stress, feeding, and sleep. The functional relevance of N/OFQ action in the mammalian brain remains unclear due to a lack of high-resolution approaches to detect this neuropeptide with appropriate spatial and temporal resolution. Here we develop and characterize NOPLight, a genetically encoded sensor that sensitively reports changes in endogenous N/OFQ release. We characterized the affinity, pharmacological profile, spectral properties, kinetics, ligand selectivity, and potential interaction with intracellular signal transducers of NOPLight in vitro. Its functionality was established in acute brain slices by exogeneous N/OFQ application and chemogenetic induction of endogenous N/OFQ release from PNOC neurons. In vivo studies with fibre photometry enabled direct recording of NOPLight binding to exogenous N/OFQ receptor ligands, as well as detection of endogenous N/OFQ release within the paranigral ventral tegmental area (pnVTA) during natural behaviors and chemogenetic activation of PNOC neurons. In summary, we show here that NOPLight can be used to detect N/OFQ opioid peptide signal dynamics in tissue and freely behaving animals.

Acute exercise suppresses hypothalamic PTP1B protein level and improves insulin and leptin signaling in obese rats.

Hypothalamic inflammation is associated with insulin and leptin resistance, hyperphagia, and obesity. In this scenario, hypothalamic protein tyrosine phosphatase 1B (PTP1B) has emerged as the key phosphatase induced by inflammation that is responsible for the central insulin and leptin resistance. Here, we demonstrated that acute exercise reduced inflammation and PTP1B protein level/activity in the hypothalamus of obese rodents. Exercise disrupted the interaction between PTP1B with proteins involved in the early steps of insulin (IRß and IRS-1) and leptin (JAK2) signaling, increased the tyrosine phosphorylation of these molecules, and restored the anorexigenic effects of insulin and leptin in obese rats. Interestingly, the anti-inflammatory action and the reduction of PTP1B activity mediated by exercise occurred in an interleukin-6 (IL-6)-dependent manner because exercise failed to reduce inflammation and PTP1B protein level after the disruption of hypothalamic-specific IL-6 action in obese rats. Conversely, intracerebroventricular administration of recombinant IL-6 reproduced the effects of exercise, improving hypothalamic insulin and leptin action by reducing the inflammatory signaling and PTP1B activity in obese rats at rest. Taken together, our study reports that physical exercise restores insulin and leptin signaling, at least in part, by reducing hypothalamic PTP1B protein level through the central anti-inflammatory response.

IL-6 and IL-10 anti-inflammatory activity links exercise to hypothalamic insulin and leptin sensitivity through IKKbeta and ER stress inhibition.

Overnutrition caused by overeating is associated with insulin and leptin resistance through IKKbeta activation and endoplasmic reticulum (ER) stress in the hypothalamus. Here we show that physical exercise suppresses hyperphagia and associated hypothalamic IKKbeta/NF-kappaB activation by a mechanism dependent upon the pro-inflammatory cytokine interleukin (IL)-6. The disruption of hypothalamic-specific IL-6 action blocked the beneficial effects of exercise on the re-balance of food intake and insulin and leptin resistance. This molecular mechanism, mediated by physical activity, involves the anti-inflammatory protein IL-10, a core inhibitor of IKKbeta/NF-kappaB signaling and ER stress. We report that exercise and recombinant IL-6 requires IL-10 expression to suppress hyperphagia-related obesity. Moreover, in contrast to control mice, exercise failed to reverse the pharmacological activation of IKKbeta and ER stress in C3H/HeJ mice deficient in hypothalamic IL-6 and IL-10 signaling. Hence, inflammatory signaling in the hypothalamus links beneficial physiological effects of exercise to the central action of insulin and leptin.

CLK2 in GABAergic neurons is critical in regulating energy balance and anxiety-like behavior in a gender-specific fashion.

Introduction: Cdc2-like kinase (CLK2) is a member of CLK kinases expressed in hypothalamic neurons and is activated in response to refeeding, leptin, or insulin. Diet-induced obesity and leptin receptor-deficient db/db mice lack CLK2 signal in the hypothalamic neurons. The neurotransmiter gamma-aminobutyric acid (GABA) is among the most prevalent in the central nervous system (CNS), particularly in the hypothalamus. Given the abundance of GABA-expressing neurons and their potential influence on regulating energy and behavioral homeostasis, we aimed to explore whether the deletion of CLK2 in GABAergic neurons alters energy homeostasis and behavioral and cognitive functions in both genders of mice lacking CLK2 in Vgat-expressing neurons (Vgat-Cre; Clk2loxP/loxP) on chow diet. Methods: We generated mice lacking Clk2 in Vgat-expressing neurons (Vgat-Cre; Clk2loxP/loxP) by mating Clk2loxP/loxP mice with Vgat-IRES-Cre transgenic mice and employed behavior, and physiological tests, and molecular approaches to investigate energy metabolism and behavior phenotype of both genders. Results and discussion: We showed that deletion of CLK2 in GABAergic neurons increased adiposity and food intake in females. The mechanisms behind these effects were likely due, at least in part, to hypothalamic insulin resistance and upregulation of hypothalamic Npy and Agrp expression. Besides normal insulin and pyruvate sensitivity, Vgat-Cre; Clk2loxP/loxP females were glucose intolerant. Male Vgat-Cre; Clk2loxP/loxP mice showed an increased energy expenditure (EE). Risen EE may account for avoiding weight and fat mass gain in male Vgat-Cre; Clk2loxP/loxP mice. Vgat-Cre; Clk2loxP/loxP mice had no alteration in cognition or memory functions in both genders. Interestingly, deleting CLK2 in GABAergic neurons changed anxiety-like behavior only in females, not males. These findings suggest that CLK2 in GABAergic neurons is critical in regulating energy balance and anxiety-like behavior in a gender-specific fashion and could be a molecular therapeutic target for combating obesity associated with psychological disorders in females.

Maternal exposure to air pollution alters energy balance transiently according to gender and changes gut microbiota.

Introduction: The timing of maternal exposure to air pollution is crucial to define metabolic changes in the offspring. Here we aimed to determine the most critical period of maternal exposure to particulate matter (PM2.5) that impairs offspring's energy metabolism and gut microbiota composition. Methods: Unexposed female and male C57BL/6J mice were mated. PM2.5 or filtered air (FA) exposure occurred only in gestation (PM2.5/FA) or lactation (FA/PM2.5). We studied the offspring of both genders. Results: PM2.5 exposure during gestation increased body weight (BW) at birth and from weaning to young in male adulthood. Leptin levels, food intake, Agrp, and Npy levels in the hypothalamus were also increased in young male offspring. Ikbke, Tnf increased in male PM2.5/FA. Males from FA/PM2.5 group were protected from these phenotypes showing higher O2 consumption and Ucp1 in the brown adipose tissue. In female offspring, we did not see changes in BW at weaning. However, adult females from PM2.5/FA displayed higher BW and leptin levels, despite increased energy expenditure and thermogenesis. This group showed a slight increase in food intake. In female offspring from FA/PM2.5, BW, and leptin levels were elevated. This group displayed higher energy expenditure and a mild increase in food intake. To determine if maternal exposure to PM2.5 could affect the offspring's gut microbiota, we analyzed alpha diversity by Shannon and Simpson indexes and beta diversity by the Linear Discriminant Analysis (LDA) in offspring at 30 weeks. Unlike males, exposure during gestation led to higher adiposity and leptin maintenance in female offspring at this age. Gestation exposure was associated with decreased alpha diversity in the gut microbiota in both genders. Discussion: Our data support that exposure to air pollution during gestation is more harmful to metabolism than exposure during lactation. Male offspring had an unfavorable metabolic phenotype at a young age. However, at an older age, only females kept more adiposity. Ultimately, our data highlight the importance of controlling air pollution, especially during gestation.

Development of a genetically-encoded sensor for probing endogenous nociceptin opioid peptide release.

Nociceptin/orphanin-FQ (N/OFQ) is a recently appreciated critical opioid peptide with key regulatory functions in several central behavioral processes including motivation, stress, feeding, and sleep. The functional relevance of N/OFQ action in the mammalian brain remains unclear due to a lack of high-resolution approaches to detect this neuropeptide with appropriate spatial and temporal resolution. Here we develop and characterize NOPLight, a genetically encoded sensor that sensitively reports changes in endogenous N/OFQ release. We characterized the affinity, pharmacological profile, spectral properties, kinetics, ligand selectivity, and potential interaction with intracellular signal transducers of NOPLight in vitro. Its functionality was established in acute brain slices by exogeneous N/OFQ application and chemogenetic induction of endogenous N/OFQ release from PNOC neurons. In vivo studies with fiber photometry enabled a direct recording of binding by N/OFQ receptor ligands, as well as the detection of natural or chemogenetically-evoked endogenous N/OFQ release within the paranigral ventral tegmental area (pnVTA). In summary, we show that NOPLight can be used to detect N/OFQ opioid peptide signal dynamics in tissue and freely-behaving animals.

Subchronic exposure to 1,2-naphthoquinone induces adipose tissue inflammation and changes the energy homeostasis of mice, partially due to TNFR1 and TLR4.

Air pollution affects energy homeostasis detrimentally. Yet, knowledge of how each isolated pollutant can impact energy metabolism remains incomplete. The present study was designed to investigate the distinct effects of 1,2-naphthoquinone (1,2-NQ) on energy metabolism since this pollutant increases at the same rate as diesel combustion. In particular, we aimed to determine in vivo effects of subchronic exposure to 1,2-NQ on metabolic and inflammatory parameters of wild-type mice (WT) and to explore the involvement of tumor necrosis factor receptor 1 (TNFR1) and toll-like receptor 4 (TLR4) in this process. Males WT, TNFR1KO, and TLR4KO mice at eight weeks of age received 1,2-NQ or vehicle via nebulization five days a week for 17 weeks. In WT mice, 1,2-NQ slightly decreased the body mass compared to vehicle-WT. This effect was likely due to a mild food intake reduction and increased energy expenditure (EE) observed after six weeks of exposure. After nine weeks of exposure, we observed higher fasting blood glucose and impaired glucose tolerance, whereas insulin sensitivity was slightly improved compared to vehicle-WT. After 17 weeks of 1,2-NQ exposure, WT mice displayed an increased percentage of M1 and a decreased (p = 0.057) percentage of M2 macrophages in adipose tissue. The deletion of TNFR1 and TLR4 abolished most of the metabolic impacts caused by 1,2-NQ exposure, except for the EE and insulin sensitivity, which remained high in these mice under 1,2-NQ exposure. Our study demonstrates for the first time that subchronic exposure to 1,2-NQ affects energy metabolism in vivo. Although 1,2-NQ increased EE and slightly reduced feeding and body mass, the WT mice displayed higher inflammation in adipose tissue and impaired fasting blood glucose and glucose tolerance. Thus, in vivo subchronic exposure to 1,2-NQ is harmful, and TNFR1 and TLR4 are partially involved in these outcomes.

Identification of intracellular peptides in rat adipose tissue: Insights into insulin resistance.

Intracellular peptides generated by the proteasome and oligopeptidases have been suggested to function in signal transduction and to improve insulin resistance in mice fed a high-caloric diet. The aim of this study was to identify specific intracellular peptides in the adipose tissue of Wistar rats that could be associated with the physiological and therapeutic control of glucose uptake. Using semiquantitative mass spectrometry and LC/MS/MS analyses, we identified ten peptides in the epididymal adipose tissue of the Wistar rats; three of these peptides were present at increased levels in rats that were fed a high-caloric Western diet (WD) compared with rats fed a control diet (CD). The results of affinity chromatography suggested that in the cytoplasm of epididymal adipose tissue from either WD or CD rats, distinctive proteins bind to these peptides. However, despite the observed increase in the WD animals, the evaluated peptides increased insulin-stimulated glucose uptake in 3T3-L1 adipocytes treated with palmitate. Thus, intracellular peptides from the adipose tissue of Wistar rats can bind to specific proteins and facilitate insulin-induced glucose uptake in 3T3-L1 adipocytes.

Acute exercise reduces feeding by activating IL-6/Tubby axis in the mouse hypothalamus.

Background: Acute exercise contributes to decreased feeding through leptin and interleukin/Janus kinase 2/signal transducers and activators of transcription 3 (IL-6/JAK2/STAT3) signaling. Considering the pleiotropic use of substrates by JAK2 and that JAK2 can phosphorylate the Tubby protein (TUB) in CHO-IR cells, we speculated that acute exercise can activate the IL-6/JAK2/TUB pathway to decrease food intake. Aims: We investigated whether acute exercise induced tyrosine phosphorylation and the association of TUB and JAK2 in the hypothalamus and if IL-6 is involved in this response, whether acute exercise increases the IL-6/TUB axis to regulate feeding, and if leptin has an additive effect over this mechanism. Methods: We applied a combination of genetic, pharmacological, and molecular approaches. Key findings: The in vivo experiments showed that acute exercise increased the tyrosine phosphorylation and association of JAK2/TUB in the hypothalamus, which reduced feeding. This response was dependent on IL-6. Leptin had no additive effect on this mechanism. Significance: The results of this study suggest a novel hypothalamic pathway by which IL-6 released by exercise regulates feeding and reinforces the beneficial effects of exercise.

Evidence for a neuromuscular circuit involving hypothalamic interleukin-6 in the control of skeletal muscle metabolism.

Hypothalamic interleukin-6 (IL6) exerts a broad metabolic control. Here, we demonstrated that IL6 activates the ERK1/2 pathway in the ventromedial hypothalamus (VMH), stimulating AMPK/ACC signaling and fatty acid oxidation in mouse skeletal muscle. Bioinformatics analysis revealed that the hypothalamic IL6/ERK1/2 axis is closely associated with fatty acid oxidation- and mitochondrial-related genes in the skeletal muscle of isogenic BXD mouse strains and humans. We showed that the hypothalamic IL6/ERK1/2 pathway requires the α2-adrenergic pathway to modify fatty acid skeletal muscle metabolism. To address the physiological relevance of these findings, we demonstrated that this neuromuscular circuit is required to underpin AMPK/ACC signaling activation and fatty acid oxidation after exercise. Last, the selective down-regulation of IL6 receptor in VMH abolished the effects of exercise to sustain AMPK and ACC phosphorylation and fatty acid oxidation in the muscle after exercise. Together, these data demonstrated that the IL6/ERK axis in VMH controls fatty acid metabolism in the skeletal muscle.

Exercise alters the mitochondrial proteostasis and induces the mitonuclear imbalance and UPRmt in the hypothalamus of mice.

The maintenance of mitochondrial activity in hypothalamic neurons is determinant to the control of energy homeostasis in mammals. Disturbs in the mitochondrial proteostasis can trigger the mitonuclear imbalance and mitochondrial unfolded protein response (UPRmt) to guarantee the mitochondrial integrity and function. However, the role of mitonuclear imbalance and UPRmt in hypothalamic cells are unclear. Combining the transcriptomic analyses from BXD mice database and in vivo experiments, we demonstrated that physical training alters the mitochondrial proteostasis in the hypothalamus of C57BL/6J mice. This physical training elicited the mitonuclear protein imbalance, increasing the mtCO-1/Atp5a ratio, which was accompanied by high levels of UPRmt markers in the hypothalamus. Also, physical training increased the maximum mitochondrial respiratory capacity in the brain. Interestingly, the transcriptomic analysis across several strains of the isogenic BXD mice revealed that hypothalamic mitochondrial DNA-encoded genes were negatively correlated with body weight and several genes related to the orexigenic response. As expected, physical training reduced body weight and food intake. Interestingly, we found an abundance of mt-CO1, a mitochondrial DNA-encoded protein, in NPY-producing neurons in the lateral hypothalamus nucleus of exercised mice. Collectively, our data demonstrated that physical training altered the mitochondrial proteostasis and induced the mitonuclear protein imbalance and UPRmt in hypothalamic cells.

Modulation of hypothalamic S6K1 and S6K2 alters feeding behavior and systemic glucose metabolism.

The mTOR/S6Ks signaling is one of the intracellular pathways important for metabolic control, acting both peripherally and centrally. In the hypothalamus, mTOR/S6Ks axis mediates the action of leptin and insulin and can modulate the expression of neuropeptides. We analyzed the role of different S6Ks isoforms in the hypothalamic regulation of metabolism. We observed decreased food intake and decreased expression of agouti-related peptide (AgRP) following intracerebroventricular (icv) injections of adenoviral-mediated overexpression of three different S6Ks isoforms. Moreover, mice overexpressing p70-S6K1 in undefined periventricular hypothalamic neurons presented changes in glucose metabolism, as an increase in gluconeogenesis. To further evaluate the hypothalamic role of a less-studied S6K isoform, p54-S6K2, we used a Cre-LoxP approach to specifically overexpress it in AgRP neurons. Our findings demonstrate the potential participation of S6K2 in AgRP neurons regulating feeding behavior.