Proopiomelanocortin Neurons Research Articles

Glucose-dependent insulinotropic polypeptide counteracts diet-induced obesity along with reduced feeding, elevated plasma leptin and activation of leptin-responsive and proopiomelanocortin neurons in the arcuate nucleus.

To clarify the effects of glucose-dependent insulinotropic polypeptide (GIP) receptor agonists (GIPRAs) on feeding and body weight. Acute and subchronic effects of subcutaneous GIPFA-085, a long-acting GIPRA, on blood glucose, food intake, body weight, respiratory exchange ratio and plasma leptin levels were measured in diet-induced obese (DIO) mice and/or functional leptin-deficient ob/ob mice. The effects of GIPFA-085 on the hypothalamic arcuate nucleus (ARC) neurons from lean and DIO mice were studied by measuring cytosolic Ca2+ concentration ([Ca2+ ]i ). Single bolus GIPFA-085 (30, 300 nmol/kg) dose-dependently reduced blood glucose in glucose tolerance tests, elevated plasma leptin levels at 0.5-6hours and inhibited food intake at 2-24 hours after injection in DIO mice. Daily GIPFA-085 (300 nmol/kg) inhibited food intake and increased fat utilization on day 1, and reduced body weight gain on days 3-12 of treatment in DIO, but not ob/ob, mice. GIPFA-085 increased [Ca2+ ]i in the ARC leptin-responsive and proopiomelanocortin (POMC) neurons. GIPFA-085 and leptin cooperated to increase [Ca2+ ]i in ARC neurons and inhibit food intake. GIPFA-085 acutely inhibits feeding and increases lipid utilization, and sustainedly lowers body weight in DIO mice via mechanisms involving rises in leptin and activation of ARC leptin-responsive and POMC neurons. This study highlights the therapeutic potential of GIPRAs for treating obesity and diabetes.

A critical update on the leptin-melanocortin system.

The discovery of leptin in 1994 was an "eureka moment" in the field of neurometabolism that provided new opportunities to better understand the central control of energy balance and glucose metabolism. Rapidly, a prevalent model in the field emerged that pro-opiomelanocortin (POMC) neurons were key in promoting leptin's anorexigenic effects and that the arcuate nucleus of the hypothalamus (ARC) was a key region for the regulation of energy homeostasis. While this model inspired many important discoveries, a growing body of literature indicates that this model is now outdated. In this review, we re-evaluate the hypothalamic leptin-melanocortin model in light of recent advances that directly tackle previous assumptions, with a particular focus on the ARC. We discuss how segregated and heterogeneous these neurons are, and examine how the development of modern approaches allowing spatiotemporal, intersectional, and chemogenetic manipulations of melanocortin neurons has allowed a better definition of the complexity of the leptin-melanocortin system. We review the importance of leptin in regulating glucose homeostasis, but not food intake, through direct actions on ARC POMC neurons. We further highlight how non-POMC, GABAergic neurons mediate leptin's direct effects on energy balance and influence POMC neurons.

Leptin Increases: Physiological Roles in the Control of Sympathetic Nerve Activity, Energy Balance, and the Hypothalamic–Pituitary–Thyroid Axis

It is well established that decreases in plasma leptin levels, as with fasting, signal starvation and elicit appropriate physiological responses, such as increasing the drive to eat and decreasing energy expenditure. These responses are mediated largely by suppression of the actions of leptin in the hypothalamus, most notably on arcuate nucleus (ArcN) orexigenic neuropeptide Y neurons and anorexic pro-opiomelanocortin neurons. However, the question addressed in this review is whether the effects of increased leptin levels are also significant on the long-term control of energy balance, despite conventional wisdom to the contrary. We focus on leptin's actions (in both lean and obese individuals) to decrease food intake, increase sympathetic nerve activity, and support the hypothalamic-pituitary-thyroid axis, with particular attention to sex differences. We also elaborate on obesity-induced inflammation and its role in the altered actions of leptin during obesity.

Hypothalamic Grb10 enhances leptin signalling and promotes weight loss.

Leptin acts on hypothalamic neurons expressing agouti-related protein (AgRP) or pro-opiomelanocortin (POMC) to suppress appetite and increase energy expenditure, but the intracellular mechanisms that modulate central leptin signalling are not fully understood. Here we show that growth factor receptor-bound protein 10 (Grb10), an adaptor protein that binds to the insulin receptor and negatively regulates its signalling pathway, can interact with the leptin receptor and enhance leptin signalling. Ablation of Grb10 in AgRP neurons promotes weight gain, while overexpression of Grb10 in AgRP neurons reduces body weight in male and female mice. In parallel, deletion or overexpression of Grb10 in POMC neurons exacerbates or attenuates diet-induced obesity, respectively. Consistent with its role in leptin signalling, Grb10 in AgRP and POMC neurons enhances the anorexic and weight-reducing actions of leptin. Grb10 also exaggerates the inhibitory effects of leptin on AgRP neurons via ATP-sensitive potassium channel-mediated currents while facilitating the excitatory drive of leptin on POMC neurons through transient receptor potential channels. Our study identifies Grb10 as a potent leptin sensitizer that contributes to the maintenance of energy homeostasis by enhancing the response of AgRP and POMC neurons to leptin.

Deconstruction of a hypothalamic astrocyte-white adipocyte sympathetic axis that regulates lipolysis in mice

The role of non-neuronal glial cells in the regulation of adipose sympathetic nerve activity and adipocyte functions such as white adipose tissue lipid lipolysis is poorly understood. Here, we combine chemo/optogenetic manipulations of medio-basal hypothalamic astrocytes, real-time fiber photometry monitoring of white adipose tissue norepinephrine (NE) contents and nerve activities, electrophysiological recordings of local sympathetic inputs to inguinal white adipose tissue (iWAT), and adipose tissue lipid lipolytic assays to define the functional roles of hypothalamic astrocytes in the regulation of iWAT sympathetic outflow and lipolysis. Our results show that astrocyte stimulation elevates iWAT NE contents, excites sympathetic neural inputs and promotes lipolysis. Mechanistically, we find that sympathetic paravertebral ganglia (PG) partake in those astrocyte effects. We also find that astrocyte stimulation excites pro-opiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARH), and chemogenetic inhibition of POMC neurons blunts the effects induced by astrocyte stimulation. While we cannot exclude potential roles played by other cell populations such as microglia, our findings in this study reveal a central astrocyte-peripheral adipocyte axis modulating sympathetic drive to adipose tissues and adipocyte functions, one that might serve as a target for therapeutic intervention in the treatment of obesity.

Increased intrinsic and synaptic excitability of hypothalamic POMC neurons underlies chronic stress-induced behavioral deficits

Chronic stress exposure induces maladaptive behavioral responses and increases susceptibility to neuropsychiatric conditions. However, specific neuronal populations and circuits that are highly sensitive to stress and trigger maladaptive behavioral responses remain to be identified. Here we investigate the patterns of spontaneous activity of proopiomelanocortin (POMC) neurons in the arcuate nucleus (ARC) of the hypothalamus following exposure to chronic unpredictable stress (CUS) for 10 days, a stress paradigm used to induce behavioral deficits such as anhedonia and behavioral despair [1, 2]. CUS exposure increased spontaneous firing of POMC neurons in both male and female mice, attributable to reduced GABA-mediated synaptic inhibition and increased intrinsic neuronal excitability. While acute activation of POMC neurons failed to induce behavioral changes in non-stressed mice of both sexes, subacute (3 days) and chronic (10 days) repeated activation of POMC neurons was sufficient to induce anhedonia and behavioral despair in males but not females under non-stress conditions. Acute activation of POMC neurons promoted susceptibility to subthreshold unpredictable stress in both male and female mice. Conversely, acute inhibition of POMC neurons was sufficient to reverse CUS-induced anhedonia and behavioral despair in both sexes. Collectively, these results indicate that chronic stress induces both synaptic and intrinsic plasticity of POMC neurons, leading to neuronal hyperactivity. Our findings suggest that POMC neuron dysfunction drives chronic stress-related behavioral deficits.

UCP2-dependent redox sensing in POMC neurons regulates feeding

Paradoxically, glucose, the primary driver of satiety, activates a small population of anorexigenic pro-opiomelanocortin (POMC) neurons. Here, we show that lactate levels in the circulation and in the cerebrospinal fluid are elevated in the fed state and the addition of lactate to glucose activates the majority of POMC neurons while increasing cytosolic NADH generation, mitochondrial respiration, and extracellular pyruvate levels. Inhibition of lactate dehydrogenases diminishes mitochondrial respiration, NADH production, and POMC neuronal activity. However, inhibition of the mitochondrial pyruvate carrier has no effect. POMC-specific downregulation of Ucp2 (Ucp2PomcKO), a molecule regulated by fatty acid metabolism and shown to play a role as transporter in the malate-aspartate shuttle, abolishes lactate- and glucose-sensing of POMC neurons. Ucp2PomcKO mice have impaired glucose metabolism and are prone to obesity on a high-fat diet. Altogether, our data show that lactate through redox signaling and blocking mitochondrial glucose utilization activates POMC neurons to regulate feeding and glucose metabolism.

Role of astroglial ACBP in energy metabolism flexibility and feeding responses to metabolic challenges in male mice.

Acyl-CoA binding protein (ACBP), also known as diazepam binding inhibitor (DBI), has recently emerged as a hypothalamic and brainstem gliopeptide regulating energy balance. Previous work has shown that the ACBP-derived octadecaneuropeptide exerts strong anorectic action via proopiomelanocortin (POMC) neuron activation and the melanocortin-4 receptor. Importantly, targeted ACBP loss-of-function in astrocytes promotes hyperphagia and diet-induced obesity while its overexpression in arcuate astrocytes reduces feeding and body weight. Despite this knowledge, the role of astroglial ACBP in adaptive feeding and metabolic responses to acute metabolic challenges has not been investigated. Using different paradigms, we found that ACBP deletion in glial fibrillary acidic protein (GFAP)-positive astrocytes does not affect weight loss when obese male mice are transitioned from a high fat diet to a chow diet, nor metabolic parameters in mice fed with a normal chow diet (e.g., energy expenditure, body temperature) during fasting, cold exposure and at thermoneutrality. In contrast, astroglial ACBP deletion impairs meal pattern and feeding responses during refeeding after a fast and during cold exposure, thereby showing that ACBP is required to stimulate feeding in states of increased energy demand. These findings challenge the general view that astroglial ACBP exerts anorectic effects and suggest that regulation of feeding by ACBP is dependent on metabolic status.

Human loss-of-function variants in the serotonin 2C receptor associated with obesity and maladaptive behavior

Serotonin reuptake inhibitors and receptor agonists are used to treat obesity, anxiety and depression. Here we studied the role of the serotonin 2C receptor (5-HT2CR) in weight regulation and behavior. Using exome sequencing of 2,548 people with severe obesity and 1,117 control individuals without obesity, we identified 13 rare variants in the gene encoding 5-HT2CR (HTR2C) in 19 unrelated people (3 males and 16 females). Eleven variants caused a loss of function in HEK293 cells. All people who carried variants had hyperphagia and some degree of maladaptive behavior. Knock-in male mice harboring a human loss-of-function HTR2C variant developed obesity and reduced social exploratory behavior; female mice heterozygous for the same variant showed similar deficits with reduced severity. Using the 5-HT2CR agonist lorcaserin, we found that depolarization of appetite-suppressing proopiomelanocortin neurons was impaired in knock-in mice. In conclusion, we demonstrate that 5-HT2CR is involved in the regulation of human appetite, weight and behavior. Our findings suggest that melanocortin receptor agonists might be effective in treating severe obesity in individuals carrying HTR2C variants. We suggest that HTR2C should be included in diagnostic gene panels for severe childhood-onset obesity.

A neural circuit for the suppression of feeding under persistent pain.

In humans, persistent pain often leads to decreased appetite. However, the neural circuits underlying this behaviour remain unclear. Here, we show that a circuit arising from glutamatergic neurons in the anterior cingulate cortex (GluACC) projects to glutamatergic neurons in the lateral hypothalamic area (GluLHA) to blunt food intake in a mouse model of persistent pain. In turn, these GluLHA neurons project to pro-opiomelanocortin neurons in the hypothalamic arcuate nucleus (POMCArc), a well-known neuronal population involved in decreasing food intake. In vivo calcium imaging and multi-tetrode electrophysiological recordings reveal that the GluACC → GluLHA → Arc circuit is activated in mouse models of persistent pain and is accompanied by decreased feeding behaviour in both males and females. Inhibition of this circuit using chemogenetics can alleviate the feeding suppression symptoms. Our study indicates that the GluACC → GluLHA → Arc circuit is involved in driving the suppression of feeding under persistent pain through POMC neuronal activity. This previously unrecognized pathway could be explored as a potential target for pain-associated diseases.

CRISPR/SaCas9 mutagenesis of stromal interaction molecule 1 in proopiomelanocortin neurons increases glutamatergic excitability and protects against diet-induced obesity

ObjectiveProopiomelanocortin (POMC) neurons are the key anorexigenic hypothalamic neuron for integrating metabolic cues to generate the appropriate output for maintaining energy homeostasis and express the requisite channels as a perfect synaptic integrator in this role. Similar to the metabolic hormones leptin and insulin, glutamate also excites POMC neurons via group I metabotropic glutamate receptors (mGluR1 and 5, mGluR1/5) that activate Transient Receptor Potential Canonical (TRPC 5) Channels to cause depolarization. A key modulator of TRPC 5 channel activity is stromal interaction molecule 1 (STIM1), which is involved in recruitment of TRPC 5 channels from receptor-operated to store-operated calcium entry following depletion of calcium from the endoplasmic reticulum. MethodsWe used a single adeno-associated viral (AAV) vector containing a recombinase-dependent Staphylococcus aureus Cas9 (SaCas) and a single guide RNA (sgRNA) to mutate Stim1 in POMCCre neurons in male mice, verified by qPCR of Stim1 mRNA expression in single POMC neurons. Whole-cell patch clamp experiments were conducted to validate the effects of Stim1 mutagenesis. Body weight and food intake were measured in male mice to assess disruptions in energy balance. ResultsReduced Stim1 expression augmented the efficacy of the mGluR1/5 agonist 3, 5-Dihydroxyphenylglycine (DHPG) to depolarize POMC neurons via a Gαq-coupled signaling pathway, which is an essential part of excitatory glutamatergic input in regulating energy homeostasis. The TRPC 5 channel blockers HC070 and Pico145 antagonized the excitatory effects of DHPG. As proof of principle, mutagenesis of Stim1 in POMC neurons reduced food intake, attenuated weight gain, reduced body fat and fat pad mass in mice fed a high fat diet. ConclusionsUsing CRISPR technology we have uncovered a critical role of STIM1 in modulating glutamatergic activation of TRPC 5 channels in POMC neurons, which ultimately is important for maintaining energy balance.

Exogenous Melatonin Regulates Puberty and the Hypothalamic GnRH-GnIH System in Female Mice.

In recent years, the age of children entering puberty is getting lower and the incidence of central precocious puberty is increasing. It is known that melatonin plays an increasingly important role in regulating animal reproduction, but the specific role and mechanism of melatonin in regulating the initiation of puberty remain unclear. The purpose of the current study was to investigate the effect of subcutaneous melatonin injection on pubertal development in female mice and its mechanism of action. Female mice that were 22 days old received 1 mg/kg doses of melatonin subcutaneously every day for 10, 15 and 20 days. The vaginal opening was checked daily. Hematoxylin and eosin (HE) stain was used to determine the growth of the uterus and ovaries. Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of follicle-stimulating hormone (FSH), gonadotropin-inhibiting hormone (GnIH), and gonadotropin-releasing hormone (GnRH) in serum. By using RT-PCR and Western blotting, the mRNA and protein expression of the hypothalamus GnRH, GnIH, Kisspeptin (Kp), Proopiomelanocortin (POMC), Neuropeptide Y (NPY), as well as G protein-coupled receptor 147 (GPR147) were identified. The findings demonstrated that melatonin could suppress ovarian follicle and uterine wall growth as well as delay vaginal opening, decrease serum levels of GnRH and FSH and increase levels of GnIH. Melatonin increased GnIH and GPR147 expression in the hypothalamus in comparison to the saline group, while decreasing the expression of GnRH, Kisspeptin, POMC, and NPY. In conclusion, exogenous melatonin can inhibit the onset of puberty in female mice by modulating the expression of hypothalamic GnRH, GnIH, Kisspeptin, POMC and NPY neurons and suppressing the hypothalamic-pituitary-gonadal axis.

Dopamine-inhibited POMCDrd2+ neurons in the ARC acutely regulate feeding and body temperature.

Dopamine acts on neurons in the arcuate nucleus (ARC) of the hypothalamus, which controls homeostatic feeding responses. Here we demonstrate a differential enrichment of dopamine receptor 1 (Drd1) expression in food intake-promoting agouti related peptide (AgRP)/neuropeptide Y (NPY) neurons and a large proportion of Drd2-expressing anorexigenic proopiomelanocortin (POMC) neurons. Owing to the nature of these receptors, this translates into a predominant activation of AgRP/NPY neurons upon dopamine stimulation and a larger proportion of dopamine-inhibited POMC neurons. Employing intersectional targeting of Drd2-expressing POMC neurons, we reveal that dopamine-mediated POMC neuron inhibition is Drd2 dependent and that POMCDrd2+ neurons exhibit differential expression of neuropeptide signaling mediators compared with the global POMC neuron population, which manifests in enhanced somatostatin responsiveness of POMCDrd2+ neurons. Selective chemogenetic activation of POMCDrd2+ neurons uncovered their ability to acutely suppress feeding and to preserve body temperature in fasted mice. Collectively, the present study provides the molecular and functional characterization of POMCDrd2+ neurons and aids our understanding of dopamine-dependent control of homeostatic energy-regulatory neurocircuits.

The orphan G protein-coupled receptor, GPR139, is expressed in the hypothalamus and is involved in the regulation of body mass, blood glucose, and insulin

GPR139 is an orphan G-protein-coupled receptor that is expressed in restricted areas of the nervous system, including the hypothalamus. In this study, we hypothesized that GPR139 could be involved in the regulation of energy balance and metabolism. In the first part of the study, we confirmed that GPR139 is expressed in the hypothalamus and particularly in proopiomelanocortin and agouti-related peptide neurons of the mediobasal hypothalamus. Using a lentivirus with a short-hairpin RNA, we inhibited the expression of GPR139 bilaterally in the mediobasal hypothalamus of mice. The intervention promoted a 40% reduction in the hypothalamic expression of GPR139, which was accompanied by an increase in body mass, a reduction in fasting blood glucose levels, and an increase in insulin levels. In the hypothalamus, inhibition of GPR139 was accompanied by a reduction in the expression of orexin. As previous studies using a pharmacological antagonist of orexin showed a beneficial impact on type 2 diabetes and glucose metabolism, we propose that the inhibition of hypothalamic GPR139 could be acting indirectly through the orexin system to control systemic glucose and insulin. In conclusion, this study advances the characterization of GPR139 in the hypothalamus, demonstrating its involvement in the regulation of body mass, blood insulin, and glycemia.

OR08-4 Endocrine Mechanisms of an Orphan G Protein-Coupled Receptor Regulating Metabolic Homeostasis

Abstract G protein-coupled receptors (GPCRs) are the largest protein family and amenable for pharmacological manipulation. GPCRs in the neuroendocrine and enteroendocrine systems respond to various biological cues and in turn exert crucial roles in maintaining whole body metabolic homeostasis. For example, GPCRs in the gastrointestinal tract are involved in maintaining glucose and energy homeostasis by regulating the release of gut hormones in response to luminal dietary nutrients as well as microbial metabolites. We recently identified that an orphan GPCR, Gpr17, was co-expressed in glucagon-like peptide-1 (GLP-1)-expressing EECs in human and rodent intestinal epithelium. We reported that acute genetic ablation of Gpr17 in intestinal epithelium (iKO) improved oral glucose tolerance and glucose-stimulated insulin secretion (GSIS) in female and male mice through increased secretion of incretin hormone in response to nutrient ingestion (Cell Reports, 2022). In the neuroendocrine system, we previously identified Gpr17 as the transcription target of Forkhead box protein O1 (FoxO1) in the hypothalamus and generated hypothalamic neuron specific Gpr17 knockout animals (Cell, 2012). Our results demonstrated that agouti-related peptide (AgRP) neuron Gpr17 knockout mice had increased satiety during fasting-refeeding challenge (Diabetes, 2015) and that proopiomelanocortin (POMC) neuron Gpr17 knockout mice had increased alpha-melanocyte stimulating hormone (aMSH) processing and increased firing of POMC neurons (Nutrition & Diabetes, 2019). However, how Gpr17 may regulate energy balance especially regarding leptin hormone signaling is unclear. To address this question, we used genetic knockout approach to generate Gpr17 conditional knockout in leptin receptor expressing neurons in the brain. We found Gpr17 deficiency in leptin receptor expressing neurons increased leptin sensitivity in animals and resulted in reduced food intake, which corresponded with reduced inflammatory response in the hypothalamus. Moreover, male Gpr17 whole body knockout mice have improved intraperitoneal glucose tolerance, and increased insulin sensitivity in hyperinsulinemic-euglycemic clamp studies. In conclusion, our collective body of work showed that ablation of Gpr17 signaling in the neuroendocrine and enteroendocrine systems led to improved neurohormonal regulation to maintain metabolic homeostasis, indicating functionally targeting this GPCR may lead to better therapeutic outcome of diabetes and obesity. Presentation: Saturday, June 11, 2022 12:15 p.m. - 12:30 p.m.

RF24 | PSUN95 Defining Predictors of Weight Loss Response to Lorcaserin

Abstract The need for personalized medicine in obesity is pressing, but we are currently unable to predict individual responses to weight loss (WL) medications. The melanocortin (MC) system consisting of proopiomelanocortin (POMC) and agouti related protein (AgRP) neurons and brain MC-Rs plays a critical role in regulating energy homeostasis and is targeted by lorcaserin (LOR), a 5HT2cR agonist previously FDA approved for WL. We therefore investigated the short and long-term effects of LOR on the MC system as assessed by cerebrospinal fluid (CSF) neuropeptide measurements and other parameters in order to identify potential biomarkers to predict WL response. Methods In phase-1 of our two-phase study, thirty subjects with obesity were randomized to receive placebo or LOR for 7-days and were then crossed over to 7-days of LOR or placebo after a 3-week washout period. Subjects then continued to phase-2 and were treated with LOR for 6-months. The study was terminated early as LOR was withdrawn from the market and only 19 subjects completed 6M. Anthropometrics, plasma and CSF were collected and test meals were administered after both placebo and LOR during phase-1 and at the end of phase-2. POMC prohormone and the POMC-derived peptide, β-endorphin (β-EP), were measured in CSF by in house ELISA and RIA. The MC-R antagonist, AgRP, was measured by ELISA in CSF and plasma as both may reflect brain AgRP. Results During phase-1 there was a decline in CSF POMC (p=0.001) and an increase in CSF β-EP (p=0.0017) resulting in an increase in the ratio of β-EP /POMC (processed peptide/prohormone) (p<0.0001) after 7-days of LOR vs placebo. Serum insulin and HOMA-IR also decreased despite no WL during phase-1 (p<0.005). After 6M of LOR, average WL was 6.9%, with 11/19 subjects achieving >5% and 7/19 >10% WL. Leptin, insulin and HOMA-IR declined. CSF POMC remained lower and β-EP and β-EP/POMC remained higher after 6M vs 7-day placebo (baseline), whereas AgRP increased only at 6M. Anthropometrics and caloric intake during test meals were not significantly different between LOR and placebo in phase-1 and did not predict WL at 6M. Phase-1 changes in POMC or β-EP did not predict WL. However, baseline CSF POMC and POMC/AgRP ratio correlated negatively with WL and were significantly lower in subjects with >10% WL. A CSF POMC cutoff of < 220 fmol/ml at baseline was found to predict 10% WL at 6M (p=0.045 Fisher). Conclusion In this study we show that the melanocortin system is impacted after 1 week of LOR. Furthermore lower melanocortin activity at baseline predicted a better weight loss response to drug treatment. Assessment of melanocortin activity could thus provide a way to personalize the pharmacotherapy of obesity with possible future alternative selective 5HT2cR agonists. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m., Sunday, June 12, 2022 1:00 p.m. - 1:05 p.m.

Prostaglandin PGE2 Receptor EP4 Regulates Microglial Phagocytosis and Increases Susceptibility to Diet-Induced Obesity.

In rodents, susceptibility to diet-induced obesity requires microglial activation, but the molecular components of this pathway remain incompletely defined. Prostaglandin PGE2 levels increase in the mediobasal hypothalamus during high-fat-diet (HFD) feeding, and the PGE2 receptor EP4 regulates microglial activation state and phagocytic activity, suggesting a potential role for microglial EP4 signaling in obesity pathogenesis. To test the role of microglial EP4 in energy balance regulation, we analyzed the metabolic phenotype in a microglia-specific EP4 knockout (MG-EP4 KO) mouse model. Microglial EP4 deletion markedly reduced weight gain and food intake in response to HFD feeding. Corresponding with this lean phenotype, insulin sensitivity was also improved in HFD-fed MG-EP4 KO mice, though glucose tolerance remained surprisingly unaffected. Mechanistically, EP4-deficient microglia showed an attenuated phagocytic state marked by reduced CD68 expression and fewer contacts with pro-opiomelanocortin (POMC) neuron processes. These cellular changes observed in the MG-EP4 KO mice corresponded with an increased density of POMC neurites extending into the paraventricular nucleus (PVN). These findings reveal that microglial EP4 signaling promotes body weight gain and insulin resistance during HFD feeding. Furthermore, the data suggest that curbing microglial phagocytic function may preserve POMC cytoarchitecture and PVN input to limit overconsumption during diet-induced obesity.

The vital role of arcuate nociceptin/orphanin FQ neurones in mounting an oestradiol-dependent adaptive response to negative energy balance via inhibition of nearby proopiomelanocortin neurones.

We tested the hypothesis that N/OFQ neurones in the arcuate nucleus (N/OFQARC ) inhibit proopiomelanocortin (POMCARC ) neurones in a diet- and hormone-dependent manner to promote a more extensive rebound hyperphagia upon re-feeding following an 18h fast. We utilized intact male or ovariectomized (OVX) female mice subjected to ad libitum-feeding or fasting conditions. N/OFQARC neurones under negative energy balance conditions displayed heightened sensitivity as evidenced by a decreased rheobase threshold, increased firing frequency, and increased burst duration and frequency compared to ad libitum-feeding conditions. Stimulation of N/OFQARC neurones more robustly inhibited POMCARC neurones under fasting conditions compared to ad libitum-feeding conditions. N/OFQARC inhibition of POMCARC neurones is hormone dependent as chemostimulation of N/OFQARC neurones from fasted males and OVX females produced a sizable outward current in POMCARC neurones. Oestradiol (E2 ) markedly attenuated the N/OFQ-induced POMCARC outward current. Additionally, N/OFQ tonically inhibits POMCARC neurones to a greater degree under fasting conditions than in ad libitum-feeding conditions as evidenced by the abrogation of N/OFQ-nociceptin opioid peptide (NOP) receptor signalling and inhibition of N/OFQ release via chemoinhibition of N/OFQARC neurones. Intra-arcuate nucleus application of N/OFQ further elevated the hyperphagic response and increased meal size during the 6 h re-feed period, and these effects were mimicked by chemostimulation of N/OFQARC neurones in vivo. E2 attenuated the robust N/OFQ-induced rebound hyperphagia seen in vehicle-treated OVX females. These data demonstrate that N/OFQARC neurones play a vital role in mitigating the impact of negative energy balance by inhibiting the excitability of anorexigenic neural substrates, an effect that is diminished by E2 in females. KEY POINTS: Nociceptin/orphanin FQ (N/OFQ) promotes increased energy intake and decreased energy expenditure under conditions of positive energy balance in a sex- and hormone-dependent manner. Here it is shown that under conditions of negative energy balance, i.e. fasting, N/OFQ inhibits anorexigenic proopiomelanocortin (POMC) neurones to a greater degree compared to homeostatic conditions due to fasting-induced hyperexcitability of N/OFQ neurones. Additionally, N/OFQ promotes a sustained increase in rebound hyperphagia and increase in meal size during the re-feed period following a fast. These results promote greater understanding of how energy balance influences the anorexigenic circuitry of the hypothalamus, and aid in understanding the neurophysiological pathways implicated in eating disorders promoting cachexia.

Involvement of POMC neurons in LEAP2 regulation of food intake and body weight.

Liver-expressed antimicrobial peptide 2 (LEAP2) is a newly discovered antagonist of the growth hormone secretagogue receptor (GHSR) and is considered the first endogenous peptide that can antagonize the metabolic actions of ghrelin. The effects of ghrelin administration on feeding behavior, body weight, and energy metabolism involve the activation of orexigenic neurons in the arcuate nucleus (ARC) of the hypothalamus. It is unclear, however, if LEAP2 applied directly to the ARC of the hypothalamus affects these metabolic processes. Here, we show that overexpression of LEAP2 in the ARC through adeno-associated virus (AAV) reduced food intake and body weight in wild-type (WT) mice fed chow and a high-fat diet (HFD) and improved metabolic disorders. LEAP2 overexpression in the ARC overrides both central and peripheral ghrelin action on a chow diet. Interestingly, this AAV-LEAP2 treatment increased proopiomelanocortin (POMC) expression while agouti-related peptide (AGRP)/neuropeptide Y (NPY) and GHSR levels remained unchanged in the hypothalamus. Additionally, intracerebroventricular (i.c.v.) administration of LEAP2 decreased food intake, increased POMC neuronal activity, and repeated LEAP2 administration to mice induced body weight loss. Using chemogenetic manipulations, we found that inhibition of POMC neurons abolished the anorexigenic effect of LEAP2. These results demonstrate that central delivery of LEAP2 leads to appetite-suppressing and body weight reduction, which might require activation of POMC neurons in the ARC.

Pro-opiomelanocortin neurons in the nucleus of the solitary tract mediate endorphinergic endogenous analgesia in mice.

The nucleus of the solitary tract (NTS) contains pro-opiomelanocortin (POMC) neurons that are 1 of the 2 major sources of β-endorphin in the brain. The functional role of these NTS POMC neurons in nociceptive and cardiorespiratory function is debated. We have shown that NTS POMC optogenetic activation produces bradycardia and transient apnoea in a working heart-brainstem preparation and chemogenetic activation with an engineered ion channel (PSAM) produced opioidergic analgesia in vivo. To better define the role of the NTS POMC neurons in behaving animals, we adopted in vivo optogenetics (ChrimsonR) and excitatory/inhibitory chemogenetic DREADD (hM3Dq/hM4Di) strategies in POMC-Cre mice. We show that optogenetic activation of NTS POMC neurons produces time-locked, graded, transient bradycardia and bradypnoea in anaesthetised mice that is naloxone sensitive (1 mg/kg, i.p.), suggesting a role of β-endorphin. Both optogenetic and chemogenetic activation of NTS POMC neurons produces sustained thermal analgesia in behaving mice that can be blocked by naloxone. It also produced analgesia in an inflammatory pain model (carrageenan) but not in a neuropathic pain model (tibial nerve transection). Inhibiting NTS POMC neurons does not produce any effect on basal nociception but inhibits stress-induced analgesia (unlike inhibition of arcuate POMC neurons). Activation of NTS POMC neuronal populations in conscious mice did not cause respiratory depression, anxiety, or locomotor deficit (in open field) or affective preference. These findings indicate that NTS POMC neurons play a key role in the generation of endorphinergic endogenous analgesia and can also regulate cardiorespiratory function.