Prokineticin 2 (PK2) is a hypothalamic neuropeptide expressed in central nervous system areas known to be involved in food intake. We therefore hypothesized that PK2 plays a role in energy homeostasis.

Uroguanylin is a gastrointestinal hormone primarily involved in fluid and electrolyte handling. It has recently been reported that prouroguanylin, secreted postprandially, is converted to uroguanylin in the brain and activates the receptor guanylate cyclase-C (GC-C) to reduce food intake and prevent obesity. We tested central nervous system administration of two GC-C agonists and found no significant reduction of food intake. We also carefully phenotyped mice lacking the GC-C receptor and found them to have normal body weight, adiposity, and glucose tolerance. Interestingly, uroguanylin knockout mice had a small but significant increase in body weight and adiposity that was accompanied by glucose intolerance. Our data indicate that the modest effects of uroguanylin on energy and glucose homeostasis are not mediated by central GC-C receptors.

Disturbances in the circadian system are associated with the development of type 2 diabetes mellitus. Here, we studied the direct contribution of the suprachiasmatic nucleus (SCN), the central pacemaker in the circadian system, in the development of insulin resistance. Exclusive bilateral SCN lesions in male C57Bl/6J mice, as verified by immunochemistry, showed a small but significant increase in body weight (+17%), which was accounted for by an increase in fat mass. In contrast, mice with collateral damage to the ventromedial hypothalamus and paraventricular nucleus showed severe obesity and insulin resistance. Mice with exclusive SCN ablation revealed a loss of circadian rhythm in activity, oxygen consumption, and food intake. Hyperinsulinemic-euglycemic clamp analysis 8 weeks after lesioning showed that the glucose infusion rate was significantly lower in SCN lesioned mice compared with sham-operated mice (-63%). Although insulin potently inhibited endogenous glucose production (-84%), this was greatly reduced in SCN lesioned mice (-7%), indicating severe hepatic insulin resistance. Our data show that SCN malfunctioning plays an important role in the disturbance of energy balance and suggest that an absence of central clock activity, in a genetically intact animal, may lead to the development of insulin resistance.

Because adipose tissue is highly vascularized, modifying adipose tissue vasculature may provide a novel method for reducing body fat. A peptide sequence that elicits apoptosis of endothelium in white fat potently reduced body weight. We sought to determine how inhibiting adipose tissue vasculature changes key aspects of energy balance regulation and the neuroendocrine system that maintains energy balance.

Decades of research in rodent models has shown that early postnatal overnutrition induces excess adiposity and other components of metabolic syndrome that persist into adulthood. The specific biologic mechanisms explaining the persistence of these effects, however, remain unknown. On postnatal day 1 (P1), mice were fostered in control (C) or small litters (SL). SL mice had increased body weight and adiposity at weaning (P21), which persisted to adulthood (P180). Detailed metabolic studies indicated that female adult SL mice have decreased physical activity and energy expenditure but not increased food intake. Genome-scale DNA methylation profiling identified extensive changes in hypothalamic DNA methylation during the suckling period, suggesting that it is a critical period for developmental epigenetics in the mouse hypothalamus. Indeed, SL mice exhibited subtle and sex-specific changes in hypothalamic DNA methylation that persisted from early life to adulthood, providing a potential mechanistic basis for the sustained physiological effects. Expression profiling in adult hypothalamus likewise provided evidence of widespread sex-specific alterations in gene expression. Together, our data indicate that early postnatal overnutrition leads to a reduction in spontaneous physical activity and energy expenditure in females and suggest that early postnatal life is a critical period during which nutrition can affect hypothalamic developmental epigenetics.

The FTO gene harbors variation with the strongest effect on adiposity and obesity risk. Previous data support a role for FTO variation in influencing food intake. We conducted a combined analysis of 16,094 boys and girls aged 1-18 years from 14 studies to examine the following: 1) the association between the FTO rs9939609 variant (or a proxy) and total energy and macronutrient intake; and 2) the interaction between the FTO variant and dietary intake, and the effect on BMI. We found that the BMI-increasing allele (minor allele) of the FTO variant was associated with increased total energy intake (effect per allele = 14.3 kcal/day [95% CI 5.9, 22.7 kcal/day], P = 6.5 × 10(-4)), but not with protein, carbohydrate, or fat intake. We also found that protein intake modified the association between the FTO variant and BMI (interactive effect per allele = 0.08 SD [0.03, 0.12 SD], P for interaction = 7.2 × 10(-4)): the association between FTO genotype and BMI was much stronger in individuals with high protein intake (effect per allele = 0.10 SD [0.07, 0.13 SD], P = 8.2 × 10(-10)) than in those with low intake (effect per allele = 0.04 SD [0.01, 0.07 SD], P = 0.02). Our results suggest that the FTO variant that confers a predisposition to higher BMI is associated with higher total energy intake, and that lower dietary protein intake attenuates the association between FTO genotype and adiposity in children and adolescents.

Insulin plays an important role in the hypothalamic control of energy balance, especially by reducing food intake. Emerging data point to a pivotal role of reactive oxygen species (ROS) in energy homeostasis regulation, but their involvement in the anorexigenic effect of insulin is unknown. Furthermore, ROS signal derived from NADPH oxidase activation is required for physiological insulin effects in peripheral cells. In this study, we investigated the involvement of hypothalamic ROS and NADPH oxidase in the feeding behavior regulation by insulin.

Blockade of the CB1 receptor is one of the promising strategies for the treatment of obesity. Although antagonists suppress food intake and reduce body weight, the role of central versus peripheral CB1 activation on weight loss and related metabolic parameters remains to be elucidated. We therefore specifically assessed and compared the respective potential relevance of central nervous system (CNS) versus peripheral CB1 receptors in the regulation of energy homeostasis and lipid and glucose metabolism in diet-induced obese (DIO) rats.

C-type natriuretic peptide (CNP) and its receptor are abundantly distributed in the brain, especially in the arcuate nucleus (ARC) of the hypothalamus associated with regulating energy homeostasis. To elucidate the possible involvement of CNP in energy regulation, we examined the effects of intracerebroventricular administration of CNP on food intake in mice. The intracerebroventricular administration of CNP-22 and CNP-53 significantly suppressed food intake on 4-h refeeding after 48-h fasting. Next, intracerebroventricular administration of CNP-22 and CNP-53 significantly decreased nocturnal food intake. The increment of food intake induced by neuropeptide Y and ghrelin was markedly suppressed by intracerebroventricular administration of CNP-22 and CNP-53. When SHU9119, an antagonist for melanocortin-3 and melanocortin-4 receptors, was coadministered with CNP-53, the suppressive effect of CNP-53 on refeeding after 48-h fasting was significantly attenuated by SHU9119. Immunohistochemical analysis revealed that intracerebroventricular administration of CNP-53 markedly increased the number of c-Fos-positive cells in the ARC, paraventricular nucleus, dorsomedial hypothalamus, ventromedial hypothalamic nucleus, and lateral hypothalamus. In particular, c-Fos-positive cells in the ARC after intracerebroventricular administration of CNP-53 were coexpressed with α-melanocyte-stimulating hormone immunoreactivity. These results indicated that intracerebroventricular administration of CNP induces an anorexigenic action, in part, via activation of the melanocortin system.

The prevalence of obesity-related diabetes is increasing worldwide. Here we report the identification of a pentapeptide, GLP-1(32-36)amide (LVKGRamide), derived from the glucoincretin hormone GLP-1, that increases basal energy expenditure and curtails the development of obesity, insulin resistance, diabetes, and hepatic steatosis in diet-induced obese mice. The pentapeptide inhibited weight gain, reduced fat mass without change in energy intake, and increased basal energy expenditure independent of physical activity. Analyses of tissues from peptide-treated mice reveal increased expression of UCP-1 and UCP-3 in brown adipose tissue and increased UCP-3 and inhibition of acetyl-CoA carboxylase in skeletal muscle, findings consistent with increased fatty acid oxidation and thermogenesis. In palmitate-treated C2C12 skeletal myotubes, GLP-1(32-36)amide activated AMPK and inhibited acetyl-CoA carboxylase, suggesting activation of fat metabolism in response to energy depletion. By mass spectroscopy, the pentapeptide is rapidly formed from GLP-1(9-36)amide, the major form of GLP-1 in the circulation of mice. These findings suggest that the reported insulin-like actions of GLP-1 receptor agonists that occur independently of the GLP-1 receptor might be mediated by the pentapeptide, and the previously reported nonapeptide (FIAWLVKGRamide). We propose that by increasing basal energy expenditure, GLP-1(32-36)amide might be a useful treatment for human obesity and associated metabolic disorders.

Hypothalamic "metabolic-sensing" neurons sense glucose and fatty acids (FAs) and play an integral role in the regulation of glucose, energy homeostasis, and the development of obesity and diabetes. Using pharmacologic agents, we previously found that ~50% of these neurons responded to oleic acid (OA) by using the FA translocator/receptor FAT/CD36 (CD36). For further elucidation of the role of CD36 in neuronal FA sensing, ventromedial hypothalamus (VMH) CD36 was depleted using adeno-associated viral (AAV) vector expressing CD36 short hairpin RNA (shRNA) in rats. Whereas their neuronal glucosensing was unaffected by CD36 depletion, the percent of neurons that responded to OA was decreased specifically in glucosensing neurons. A similar effect was seen in total-body CD36-knockout mice. Next, weanling rats were injected in the VMH with CD36 AAV shRNA. Despite significant VMH CD36 depletion, there was no effect on food intake, body weight gain, or total carcass adiposity on chow or 45% fat diets. However, VMH CD36-depleted rats did have increased plasma leptin and subcutaneous fat deposition and markedly abnormal glucose tolerance. These results demonstrate that CD36 is a critical factor in both VMH neuronal FA sensing and the regulation of energy and glucose homeostasis.

Evidence links the hypothalamic fatty acid synthase (FAS) pathway to the regulation of food intake and body weight. This includes pharmacological inhibitors that potently reduce feeding and body weight. The mammalian target of rapamycin (mTOR) is an intracellular fuel sensor whose activity in the hypothalamus is also linked to the regulation of energy balance. The purpose of these experiments was to determine whether hypothalamic mTOR complex 1 (mTORC1) signaling is involved in mediating the effects of FAS inhibitors.

Transcription factor forkhead box O1 (FoxO1) regulates energy expenditure (EE), food intake, and hepatic glucose production. These activities have been mapped to specific hypothalamic neuronal populations using cell type-specific knockout experiments in mice. To parse out the integrated output of FoxO1-dependent transcription from different neuronal populations and multiple hypothalamic regions, we used transgenic mice expressing Cre recombinase from the Nkx2.1 promoter to ablate loxP-flanked Foxo1 alleles from a majority of hypothalamic neurons (Foxo1KO(Nkx2.1) mice). This strategy resulted in the expected inhibition of FoxO1 expression, but only produced a transient reduction of body weight as well as a decreased body length. The transient decrease of body weight in male mice was accompanied by decreased fat mass. Male Foxo1KO(Nkx2.1) mice show food intake similar to that in wild-type controls, and, although female knockout mice eat less, they do so in proportion to a reduced body size. EE is unaffected in Foxo1KO(Nkx2.1) mice, although small increases in body temperature are present. Unlike other neuron-specific Foxo1 knockout mice, Foxo1KO(Nkx2.1) mice are not protected from diet-induced obesity. These studies indicate that, unlike the metabolic effects of highly restricted neuronal subsets (proopiomelanocortin, neuropeptide Y/agouti-related peptide, and steroidogenic factor 1), those of neurons derived from the Nkx2.1 lineage either occur in a FoxO1-independent fashion or are compensated for through developmental plasticity.

Pharmacological activation of the hypothalamic glucagon-like peptide 1 (GLP-1) receptor (GLP-1R) promotes weight loss and improves glucose tolerance. This demonstrates that the hypothalamic GLP-1R is sufficient but does not show whether it is necessary for the effects of exogenous GLP-1R agonists (GLP-1RA) or endogenous GLP-1 on these parameters. To address this, we crossed mice harboring floxed Glp1r alleles to mice expressing Nkx2.1-Cre to knock down Glp1r expression throughout the hypothalamus (GLP-1RKDΔNkx2.1cre). We also generated mice lacking Glp1r expression specifically in two GLP-1RA-responsive hypothalamic feeding nuclei/cell types, the paraventricular nucleus (GLP-1RKDΔSim1cre) and proopiomelanocortin neurons (GLP-1RKDΔPOMCcre). Chow-fed GLP-1RKDΔNkx2.1cre mice exhibited increased food intake and energy expenditure with no net effect on body weight. When fed a high-fat diet, these mice exhibited normal food intake but elevated energy expenditure, yielding reduced weight gain. None of these phenotypes were observed in GLP-1RKDΔSim1cre and GLP-1RKDΔPOMCcre mice. The acute anorectic and glucose tolerance effects of peripherally dosed GLP-1RA exendin-4 and liraglutide were preserved in all mouse lines. Chronic liraglutide treatment reduced body weight in chow-fed GLP-1RKDΔNkx2.1cre mice, but this effect was attenuated with high-fat diet feeding. In sum, classic homeostatic control regions are sufficient but not individually necessary for the effects of GLP-1RA on nutrient homeostasis.

Maternal obesity and gestational diabetes mellitus (GDM) are associated with obesity and diabetes risk in offspring. We tested whether maternal insulin resistance, which frequently coexists with GDM and obesity, could independently contribute to dysregulation of offspring metabolism. Female mice haploinsufficient for insulin receptor substrate-1 (IRS1-het) are hyperinsulinemic and insulin resistant during pregnancy, despite normal plasma glucose and body weight, and thus serve as a model of isolated maternal insulin resistance. Wild-type (WT) offspring of IRS1-het dams insulin resistance-exposed [IR-exposed] were compared with WT offspring of WT dams. Despite no differences in adiposity, male IR-exposed pups were glucose intolerant (P = 0.04) and hyperinsulinemic (1.3-fold increase, P = 0.02) by 1 month of age and developed progressive fasting hyperglycemia. Moreover, male IR-exposed pups challenged with high-fat diet exhibited insulin resistance. Liver lipidomic analysis of 3-week-old IR-exposed males revealed increases in the 16:1n7 fraction of several lipid classes, suggesting increased Scd1 activity. By 6 months of age, IR-exposed males had increased lipid accumulation in liver as well as increased plasma refed fatty acids, consistent with disrupted lipid metabolism. Our results indicate that isolated maternal insulin resistance, even in the absence of hyperglycemia or obesity, can promote metabolic perturbations in male offspring.

The recognition of sensory signals from within the body (interoceptive) and from the external environment (exteroceptive), along with the integration of these cues by the central nervous system, plays a crucial role in maintaining metabolic balance. This orchestration is vital for regulating processes related to both food intake and energy expenditure. Animal model studies indicate that manipulating specific populations of neurons in the central nervous system which influence these processes can effectively modify energy balance. This body of work presents an opportunity for the development of innovative weight loss therapies for the treatment of obesity and type 2 diabetes. In this overview, we delve into the sensory cues and the neuronal populations responsible for their integration, exploring their potential in the development of weight loss treatments for obesity and type 2 diabetes. This article is the first in a series of Perspectives that report on research funded by the American Diabetes Association Pathway to Stop Diabetes program.

Oxyntomodulin (OXM) is a glucagon-like peptide 1 (GLP-1) receptor (GLP1R)/glucagon receptor (GCGR) dual agonist peptide that reduces body weight in obese subjects through increased energy expenditure and decreased energy intake. The metabolic effects of OXM have been attributed primarily to GLP1R agonism. We examined whether a long acting GLP1R/GCGR dual agonist peptide exerts metabolic effects in diet-induced obese mice that are distinct from those obtained with a GLP1R-selective agonist.

Awake and fed thermogenesis (AFT) is the energy expenditure (EE) of the nonactive fed condition above the minimum metabolic requirement during sleep and is composed of the thermic effect of food and the cost of being awake. AFT was estimated from whole-room 24-h EE measures in 509 healthy subjects (368 Native Americans and 141 whites) while subjects consumed a eucaloric diet. Follow-up data were available for 290 Native Americans (median follow-up time: 6.6 years). AFT accounted for ~10% of 24-h EE and explained a significant portion of deviations from expected energy requirements. Energy intake was the major determinant of AFT. AFT, normalized as a percentage of intake, was inversely related to age and fasting glucose concentration and showed a nonlinear relationship with waist circumference and BMI. Spline analysis demonstrated that AFT becomes inversely related to BMI at an inflection point of 29 kg/m(2). The residual variance of AFT, after accounting for covariates, predicted future weight change only in subjects with a BMI >29 kg/m(2). AFT may influence daily energy balance, is reduced in obese individuals, and predicts future weight gain in these subjects. Once central adiposity develops, a blunting of AFT may occur that then contributes to further weight gain.

Effectors of the phosphoinositide 3-kinase (PI3K) signal transduction pathway contribute to the hypothalamic regulation of energy and glucose homeostasis in divergent ways. Here we show that central nervous system (CNS) action of the PI3K signaling intermediate atypical protein kinase C (aPKC) constrains food intake, weight gain, and glucose intolerance in both rats and mice. Pharmacological inhibition of CNS aPKC activity acutely increases food intake and worsens glucose tolerance in chow-fed rodents and causes excess weight gain during high-fat diet (HFD) feeding. Similarly, selective deletion of the aPKC isoform Pkc-λ in proopiomelanocortin (POMC) neurons disrupts leptin action, reduces melanocortin content in the paraventricular nucleus, and markedly increases susceptibility to obesity, glucose intolerance, and insulin resistance specifically in HFD-fed male mice. These data implicate aPKC as a novel regulator of energy and glucose homeostasis downstream of the leptin-PI3K pathway in POMC neurons.

Dietary methionine restriction (MR) produces a rapid and persistent remodeling of white adipose tissue (WAT), an increase in energy expenditure (EE), and enhancement of insulin sensitivity. Recent work established that hepatic expression of FGF21 is robustly increased by MR. Fgf21-/- mice were used to test whether FGF21 is an essential mediator of the physiological effects of dietary MR. The MR-induced increase in energy intake and EE and activation of thermogenesis in WAT and brown adipose tissue were lost in Fgf21-/- mice. However, dietary MR produced a comparable reduction in body weight and adiposity in both genotypes because of a negative effect of MR on energy intake in Fgf21-/- mice. Despite the similar loss in weight, dietary MR produced a more significant increase in in vivo insulin sensitivity in wild-type than in Fgf21-/- mice, particularly in heart and inguinal WAT. In contrast, the ability of MR to regulate lipogenic and integrated stress response genes in liver was not compromised in Fgf21-/- mice. Collectively, these findings illustrate that FGF21 is a critical mediator of the effects of dietary MR on EE, remodeling of WAT, and increased insulin sensitivity but not of its effects on hepatic gene expression.