Claudia R. Vianna

Glucose sensing by POMC neurons regulates glucose homeostasis and is impaired in obesity

A subset of neurons in the brain, known as ‘glucose-excited’ neurons, depolarize and increase their firing rate in response to increases in extracellular glucose. Similar to insulin secretion by pancreatic β-cells, glucose excitation of neurons is driven by ATP-mediated closure of ATP-sensitive potassium (KATP) channels. Although β-cell-like glucose sensing in neurons is well established, its physiological relevance and contribution to disease states such as type 2 diabetes remain unknown. To address these issues, we disrupted glucose sensing in glucose-excited pro-opiomelanocortin (POMC) neurons via transgenic expression of a mutant Kir6.2 subunit (encoded by the Kcnj11 gene) that prevents ATP-mediated closure of KATP channels. Here we show that this genetic manipulation impaired the whole-body response to a systemic glucose load, demonstrating a role for glucose sensing by POMC neurons in the overall physiological control of blood glucose. We also found that glucose sensing by POMC neurons became defective in obese mice on a high-fat diet, suggesting that loss of glucose sensing by neurons has a role in the development of type 2 diabetes. The mechanism for obesity-induced loss of glucose sensing in POMC neurons involves uncoupling protein 2 (UCP2), a mitochondrial protein that impairs glucose-stimulated ATP production. UCP2 negatively regulates glucose sensing in POMC neurons. We found that genetic deletion of Ucp2 prevents obesity-induced loss of glucose sensing, and that acute pharmacological inhibition of UCP2 reverses loss of glucose sensing. We conclude that obesity-induced, UCP2-mediated loss of glucose sensing in glucose-excited neurons might have a pathogenic role in the development of type 2 diabetes.

Direct leptin action on POMC neurons regulates glucose homeostasis and hepatic insulin sensitivity in mice

Leptin action on its receptor (LEPR) stimulates energy expenditure and reduces food intake, thereby lowering body weight. One leptin-sensitive target cell mediating these effects on energy balance is the proopiomelano-cortin (POMC) neuron. Recent evidence suggests that the action of leptin on POMC neurons regulates glucose homeostasis independently of its effects on energy balance. Here, we have dissected the physiological impact of direct leptin action on POMC neurons using a mouse model in which endogenous LEPR expression was prevented by a LoxP-flanked transcription blocker (loxTB), but could be reactivated by Cre recombinase. Mice homozygous for the Lepr(loxTB) allele were obese and exhibited defects characteristic of LEPR deficiency. Reexpression of LEPR only in POMC neurons in the arcuate nucleus of the hypothalamus did not reduce food intake, but partially normalized energy expenditure and modestly reduced body weight. Despite the moderate effects on energy balance and independent of changes in body weight, restoring LEPR in POMC neurons normalized blood glucose and ameliorated hepatic insulin resistance, hyperglucagonemia, and dyslipidemia. Collectively, these results demonstrate that direct leptin action on POMC neurons does not reduce food intake, but is sufficient to normalize glucose and glucagon levels in mice otherwise lacking LEPR.

Hypothalamic POMC neurons promote cannabinoid-induced feeding

Hypothalamic pro-opiomelanocortin (POMC) neurons promote satiety. Cannabinoid receptor 1 (CB1R) is critical for the central regulation of food intake. Here we test whether CB1R-controlled feeding in sated mice is paralleled by decreased activity of POMC neurons. We show that chemical promotion of CB1R activity increases feeding, and notably, CB1R activation also promotes neuronal activity of POMC cells. This paradoxical increase in POMC activity was crucial for CB1R-induced feeding, because designer-receptors-exclusively-activated-by-designer-drugs (DREADD)-mediated inhibition of POMC neurons diminishes, whereas DREADD-mediated activation of POMC neurons enhances CB1R-driven feeding. The Pomc gene encodes both the anorexigenic peptide α-melanocyte-stimulating hormone, and the opioid peptide β-endorphin. CB1R activation selectively increases β-endorphin but not α-melanocyte-stimulating hormone release in the hypothalamus, and systemic or hypothalamic administration of the opioid receptor antagonist naloxone blocks acute CB1R-induced feeding. These processes involve mitochondrial adaptations that, when blocked, abolish CB1R-induced cellular responses and feeding. Together, these results uncover a previously unsuspected role of POMC neurons in the promotion of feeding by cannabinoids.

SIRT1 deacetylase in POMC neurons is required for homeostatic defenses against diet-induced obesity

Feeding on high-calorie (HC) diets induces serious metabolic imbalances, including obesity. Understanding the mechanisms against excessive body weight gain is critical for developing effective antiobesity strategies. Here we show that lack of nicotinamide adenine dinucleotide (NAD(+))-dependent deacetylase SIRT1 in pro-opiomelanocortin (POMC) neurons causes hypersensitivity to diet-induced obesity due to reduced energy expenditure. The ability of leptin to properly engage the phosphoinositide 3-kinase (PI3K) signaling in POMC neurons and elicit remodeling of perigonadal white adipose tissue (WAT) is severely compromised in mutant mice. Also, electrophysiological and histomorphomolecular analyses indicate a selective reduction in sympathetic nerve activity and brown-fat-like characteristics in perigonadal WAT of mutant mice, suggesting a physiologically important role for POMC neurons in controlling this visceral fat depot. In summary, our results provide direct genetic evidence that SIRT1 in POMC neurons is required for normal autonomic adaptations against diet-induced obesity.

Central administration of resveratrol improves diet-induced diabetes.

Resveratrol is a natural polyphenolic compound that activates nicotinamide adenosine dinucleotide-dependent deacetylase SIRT1. Resveratrol has recently been shown to exert potent antidiabetic actions when orally delivered to animal models of type 2 diabetes. However, the tissue(s) mediating these beneficial effects is unknown. Because SIRT1 is expressed in central nervous system (CNS) neurons known to control glucose and insulin homeostasis, we hypothesized that resveratrol antidiabetic effects are mediated by the brain. Here, we report that long-term intracerebroventricular infusion of resveratrol normalizes hyperglycemia and greatly improves hyperinsulinemia in diet-induced obese and diabetic mice. It is noteworthy that these effects are independent of changes in body weight, food intake, and circulating leptin levels. In addition, CNS resveratrol delivery improves hypothalamic nuclear factor-kappaB inflammatory signaling by reducing acetylated-RelA/p65 and total RelA/p65 protein contents, and inhibitor of nuclear factor-kappaB alpha and IkappaB kinase beta mRNA levels. Furthermore, this treatment leads to reduced hepatic phosphoenolpyruvate carboxykinase 1 mRNA and protein levels and ameliorates pyruvate-induced hyperglycemia in this mouse model of type 2 diabetes. Collectively, our results unveiled a previously unrecognized key role for the CNS in mediating the antidiabetic actions of resveratrol.

Hepatocyte Toll-like receptor 4 regulates obesity-induced inflammation and insulin resistance.

Chronic low-grade inflammation is a hallmark of obesity and thought to contribute to the development of obesity-related insulin resistance. Toll-like receptor 4 (Tlr4) is a key mediator of pro-inflammatory responses. Mice lacking Tlr4s are protected from diet-induced insulin resistance and inflammation; however which Tlr4 expressing cells mediate this effect is unknown. Here we show that mice deficient in hepatocyte Tlr4 (Tlr4LKO) exhibit improved glucose tolerance, enhanced insulin sensitivity, and ameliorated hepatic steatosis despite the development of obesity after a high fat diet (HFD) challenge. Furthermore, Tlr4LKO mice have reduced macrophage content in white adipose tissue, as well as decreased tissue and circulating inflammatory markers. In contrast, the loss of Tlr4 activity in myeloid cells has little effect on insulin sensitivity. Collectively, these data indicate that the activation of Tlr4 on hepatocytes contributes to obesity-associated inflammation and insulin resistance, and suggest that targeting hepatocyte Tlr4 might be a useful therapeutic strategy for the treatment of type 2 diabetes.

SIRT1 deacetylase in SF1 neurons protects against metabolic imbalance

Chronic feeding on high-calorie diets causes obesity and type 2 diabetes mellitus (T2DM), illnesses that affect hundreds of millions. Thus, understanding the pathways protecting against diet-induced metabolic imbalance is of paramount medical importance. Here, we show that mice lacking SIRT1 in steroidogenic factor 1 (SF1) neurons are hypersensitive to dietary obesity owing to maladaptive energy expenditure. Also, mutant mice have increased susceptibility to developing dietary T2DM due to insulin resistance in skeletal muscle. Mechanistically, these aberrations arise, in part, from impaired metabolic actions of the neuropeptide orexin-A and the hormone leptin. Conversely, mice overexpressing SIRT1 in SF1 neurons are more resistant to diet-induced obesity and insulin resistance due to increased energy expenditure and enhanced skeletal muscle insulin sensitivity. Our results unveil important protective roles of SIRT1 in SF1 neurons against dietary metabolic imbalance.

Serotonin 2C receptors in pro-opiomelanocortin neurons regulate energy and glucose homeostasis

Energy and glucose homeostasis are regulated by central serotonin 2C receptors. These receptors are attractive pharmacological targets for the treatment of obesity; however, the identity of the serotonin 2C receptor-expressing neurons that mediate the effects of serotonin and serotonin 2C receptor agonists on energy and glucose homeostasis are unknown. Here, we show that mice lacking serotonin 2C receptors (Htr2c) specifically in pro-opiomelanocortin (POMC) neurons had normal body weight but developed glucoregulatory defects including hyperinsulinemia, hyperglucagonemia, hyperglycemia, and insulin resistance. Moreover, these mice did not show anorectic responses to serotonergic agents that suppress appetite and developed hyperphagia and obesity when they were fed a high-fat/high-sugar diet. A requirement of serotonin 2C receptors in POMC neurons for the maintenance of normal energy and glucose homeostasis was further demonstrated when Htr2c loss was induced in POMC neurons in adult mice using a tamoxifen-inducible POMC-cre system. These data demonstrate that serotonin 2C receptor-expressing POMC neurons are required to control energy and glucose homeostasis and implicate POMC neurons as the target for the effect of serotonin 2C receptor agonists on weight-loss induction and improved glycemic control.

Leptin signaling in Kiss1 neurons arises after pubertal development.

The adipocyte-derived hormone leptin is required for normal pubertal maturation in mice and humans and, therefore, leptin has been recognized as a crucial metabolic cue linking energy stores and the onset of puberty. Several lines of evidence have suggested that leptin acts via kisspeptin expressing neurons of the arcuate nucleus to exert its effects. Using conditional knockout mice, we have previously demonstrated that deletion of leptin receptors (LepR) from kisspeptin cells cause no puberty or fertility deficits. However, developmental adaptations and system redundancies may have obscured the physiologic relevance of direct leptin signaling in kisspeptin neurons. To overcome these putative effects, we re-expressed endogenous LepR selectively in kisspeptin cells of mice otherwise null for LepR, using the Cre-loxP system. Kiss1-Cre LepR null mice showed no pubertal development and no improvement of the metabolic phenotype, remaining obese, diabetic and infertile. These mice displayed decreased numbers of neurons expressing Kiss1 gene, similar to prepubertal control mice, and an unexpected lack of re-expression of functional LepR. To further assess the temporal coexpression of Kiss1 and Lepr genes, we generated mice with the human renilla green fluorescent protein (hrGFP) driven by Kiss1 regulatory elements and crossed them with mice that express Cre recombinase from the Lepr locus and the R26-tdTomato reporter gene. No coexpression of Kiss1 and LepR was observed in prepubertal mice. Our findings unequivocally demonstrate that kisspeptin neurons are not the direct target of leptin in the onset of puberty. Leptin signaling in kisspeptin neurons arises only after completion of sexual maturation.

Leptin engages a hypothalamic neurocircuitry to permit survival in the absence of insulin

The dogma that life without insulin is incompatible has recently been challenged by results showing the viability of insulin-deficient rodents undergoing leptin monotherapy. Yet, the mechanisms underlying these actions of leptin are unknown. Here, the metabolic outcomes of intracerebroventricular (i.c.v.) administration of leptin in mice devoid of insulin and lacking or re-expressing leptin receptors (LEPRs) only in selected neuronal groups were assessed. Our results demonstrate that concomitant re-expression of LEPRs only in hypothalamic γ-aminobutyric acid (GABA) and pro-opiomelanocortin (POMC) neurons is sufficient to fully mediate the lifesaving and antidiabetic actions of leptin in insulin deficiency. Our analyses indicate that enhanced glucose uptake by brown adipose tissue and soleus muscle, as well as improved hepatic metabolism, underlies these effects of leptin. Collectively, our data elucidate a hypothalamic-dependent pathway enabling life without insulin and hence pave the way for developing better treatments for diseases of insulin deficiency.