Liver-innervating vagal sensory neurons play an indispensable role in the development of hepatic steatosis and anxiety-like behavior in mice fed a high-fat diet.

Background and Aims: The visceral organ-brain axis, mediated by vagal sensory neurons in the vagal nerve ganglion, is essential for maintaining various physiological functions. In this study, we investigated the impact of liver-projecting vagal sensory neurons on energy balance, hepatic steatosis, and anxiety-like behavior in mice under obesogenic conditions. Methods: We performed single-nucleus RNA sequencing of vagal sensory neurons innervating the liver. Based on our snRNA-Seq results, we used the Avil CreERT2 strain to identify vagal sensory neurons that innervate the liver. Results: A small subset of polymodal sensory neurons innervating the liver was located in the left and right ganglia, projecting centrally to the nucleus of the tractus solitarius, area postrema, and dorsal motor nucleus of the vagus, and peripherally to the periportal areas in the liver. Male and female control mice developed diet-induced obesity (DIO) during high-fat diet feeding. Deleting liver-projecting advillin-positive vagal sensory neurons prevented DIO in male and female mice, and these outcomes are associated with increased energy expenditure. Although males and females exhibited improved glucose homeostasis following disruption of liver-projecting vagal sensory neurons, only male mice displayed increased insulin sensitivity. The loss of liver-projecting vagal sensory neurons limited the progression of hepatic steatosis in male and female mice fed a steatogenic diet. Finally, mice lacking liver-innervating vagal sensory neurons exhibited less anxiety-like behavior compared to the control mice. Conclusions: The liver-brain axis contributes to the regulation of energy balance, glucose tolerance, hepatic steatosis, and anxiety-like behavior depending on the nutrient status in healthy and obesogenic conditions.

Loss of the brain-liver axis prevents hepatic steatosis in mice.

Hepatic lipid metabolism is regulated by the autonomic nervous system of the liver, with the sympathetic innervation being extensively studied, while the parasympathetic efferent innervation is less understood despite its potential importance. In this study, we investigate the consequences of disrupted brain-liver communication on hepatic lipid metabolism in mice exposed to obesogenic conditions. We found that a subset of hepatocytes and the bile duct are innervated by parasympathetic nerves originating from the dorsal motor nucleus of the vagus. The elimination of the brain-liver axis by deleting parasympathetic cholinergic neurons innervating the liver prevents hepatic steatosis and promots browning of inguinal white adipose tissue (ingWAT). The loss of the brain-liver axis also raises hepatic Cyp7b1 expression and fasting serum bile acid levels. Furthermore, knockdown of the G protein-coupled bile acid receptor 1 gene in ingWAT reverses the beneficial effects of the loss of the brain-liver axis, leading to the reappearance of hepatic steatosis in the experimental groups. However, deleting the brain-liver axis has a small but significant effect on body weight, which is accompanied by an increase in energy expenditure. Therefore, altering parasympathetic cholinergic innervation of the liver could offer a potential therapeutic approach for enhancing hepatic lipid metabolism in obesity and diabetes.

The gut signals to AGRP-expressing cells of the pituitary to control glucose homeostasis.

Glucose homeostasis can be improved after bariatric surgery, which alters bile flow and stimulates gut hormone secretion, particularly FGF15/19. FGFR1 expression in AGRP-expressing cells is required for bile acids' ability to improve glucose control. We show that the mouse Agrp gene has 3 promoter/enhancer regions that direct transcription of each of their own AGRP transcripts. One of these Agrp promoters/enhancers, Agrp-B, is regulated by bile acids. We generated an Agrp-B knockin FLP/knockout allele. AGRP-B-expressing cells are found in endocrine cells of the pars tuberalis and coexpress diacylglycerol lipase B - an endocannabinoid biosynthetic enzyme - distinct from pars tuberalis thyrotropes. AGRP-B expression is also found in the folliculostellate cells of the pituitary's anterior lobe. Mice without AGRP-B were protected from glucose intolerance induced by high-fat feeding but not from excess weight gain. Chemogenetic inhibition of AGRP-B cells improved glucose tolerance by enhancing glucose-stimulated insulin secretion. Inhibition of the AGRP-B cells also caused weight loss. The improved glucose tolerance and reduced body weight persisted up to 6 weeks after cessation of the DREADD-mediated inhibition, suggesting the presence of a biological switch for glucose homeostasis that is regulated by long-term stability of food availability.

Magel2 knockdown in hypothalamic POMC neurons innervating the medial amygdala reduces susceptibility to diet-induced obesity.

Hyperphagia and obesity profoundly affect the health of children with Prader-Willi syndrome (PWS). The Magel2 gene among the genes in the Prader-Willi syndrome deletion region is expressed in proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC). Knockout of the Magel2 gene disrupts POMC neuronal circuits and functions. Here, we report that loss of the Magel2 gene exclusively in ARCPOMC neurons innervating the medial amygdala (MeA) causes a reduction in body weight in both male and female mice fed with a high-fat diet. This anti-obesity effect is associated with an increased locomotor activity. There are no significant differences in glucose and insulin tolerance in mice without the Magel2 gene in ARCPOMC neurons innervating the MeA. Plasma estrogen levels are higher in female mutant mice than in controls. Blockade of the G protein-coupled estrogen receptor (GPER), but not estrogen receptor-α (ER-α), reduces locomotor activity in female mutant mice. Hence, our study provides evidence that knockdown of the Magel2 gene in ARCPOMC neurons innervating the MeA reduces susceptibility to diet-induced obesity with increased locomotor activity through activation of central GPER.

Overexpressing the hydroxycarboxylic acid receptor 1 in mouse brown adipose tissue restores glucose tolerance and insulin sensitivity in diet-induced obese mice.

Interscapular brown adipose tissue (BAT) plays an important role in controlling glucose homeostasis. Increased glucose entry and glycolysis in BAT result in lactate production and release. The adipose tissue expresses the lactate receptor hydrocarboxylic acid receptor 1 (HCAR1), markedly downregulated in male diet-induced obese (DIO) and ob/ob mice. In this study, we examined the role of HCAR1 in BAT in controlling glucose homeostasis in male DIO mice. We overexpressed HCAR1 in BAT by injecting adeno-associated viruses (AAVs) expressing HCAR1 into the BAT pads of male DIO C57BL/6J mice. Overexpressing HCAR1 in BAT resulted in augmented glucose uptake by BAT in response to treatment with the HCAR1 agonist. HCAR1 overexpression elevated BAT temperature associated with increased thermogenic gene expression in BAT. HCAR1 overexpression prevented body weight gain in male DIO mice. Importantly, mice overexpressing HCAR1 in BAT exhibited improved glucose tolerance and insulin sensitivity. HCAR1 overexpression upregulated the Slc2a4 gene expression and promoted GLUT4 trafficking to the plasma membrane. In addition, mice overexpressing HCAR1 displayed a decrease in hormone-sensitive lipase (HSL) phosphorylation and increased lipogenic enzyme gene expression in BAT. Unlike DIO mice, overexpressing HCAR1 in BAT of mice fed a low-fat diet did not change body weight gain and glucose homeostasis. Taken together, our results support the interpretation that HCAR1 expressed in BAT promotes glucose entry and reduces lipolysis in BAT of male DIO mice. As activation of HCAR1 in BAT restores body weight, glucose tolerance, and insulin sensitivity in male DIO mice, our study suggests that interoceptive lactate detection via HCAR1 in BAT can regulate glucose and lipid substrate utilization and/or availability to promote healthy metabolism.NEW & NOTEWORTHY HCAR1 expressed in BAT can promote glucose entry and reduce lipolysis, resulting in body weight loss and increased insulin sensitivity. Hence, targeting HCAR1 in BAT would provide an alternative way to control body weight and euglycemia in individuals with obesity.

Optogenetic stimulation of the liver-projecting melanocortinergic pathway promotes hepatic glucose production.

The central melanocortin system plays a fundamental role in the control of feeding and body weight. Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC) also regulate overall glucose homeostasis via insulin-dependent and -independent pathways. Here, we report that a subset of ARC POMC neurons innervate the liver via preganglionic parasympathetic acetylcholine (ACh) neurons in the dorsal motor nucleus of the vagus (DMV). Optogenetic stimulation of this liver-projecting melanocortinergic pathway elevates blood glucose levels that is associated with increased expression of hepatic gluconeogenic enzymes in female and male mice. Pharmacological blockade and knockdown of the melanocortin-4 receptor gene in the DMV abolish this stimulation-induced effect. Activation of melanocortin-4 receptors inhibits DMV cholinergic neurons and optogenetic inhibition of liver-projecting parasympathetic cholinergic fibers increases blood glucose levels. This elevated blood glucose is not due to altered pancreatic hormone release. Interestingly, insulin-induced hypoglycemia increases ARC POMC neuron activity. Hence, this liver-projecting melanocortinergic circuit that we identified may play a critical role in the counterregulatory response to hypoglycemia.

Activation of the ARCPOMC→MeA Projection Reduces Food Intake.

Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC) plays an essential role in the control of food intake and energy expenditure. Melanocortin-4 receptors (MC4Rs) are expressed in key areas that are implicated in regulating energy homeostasis. Although the importance of MC4Rs in the paraventricular hypothalamus (PVH) has been well documented, the role of MC4Rs in the medial amygdala (MeA) on feeding remains controversial. In this study, we specifically examine the role of a novel ARCPOMC→MeA neural circuit in the regulation of short-term food intake. To map a local melanocortinergic neural circuit, we use monosynaptic anterograde as well as retrograde viral tracers and perform double immunohistochemistry to determine the identity of the neurons receiving synaptic input from POMC neurons in the ARC. To investigate the role of the ARCPOMC→MeA projection on feeding, we optogenetically stimulate channelrhodopsin-2 (ChR2)-expressing POMC fibers in the MeA. Anterograde viral tracing studies reveal that ARC POMC neurons send axonal projections to estrogen receptor-α (ER-α)- and MC4R-expressing neurons in the MeA. Retrograde viral tracing experiments show that the neurons projecting to the MeA is located mainly in the lateral part of the ARC. Optogenetic stimulation of the ARCPOMC→MeA pathway reduces short-term food intake. This anorectic effect is blocked by treatment with the MC4R antagonist SHU9119. In addition to the melanocortinergic local circuits within the hypothalamus, this extrahypothalamic ARCPOMC→MeA neural circuit would play a role in regulating short-term food intake.

Hydrocarboxylic acid receptor 1 in BAT regulates glucose uptake in mice fed a high-fat diet.

Interscapular brown adipose tissue (BAT) has the capability to take up glucose from the circulation. Despite the important role of BAT in the control of glucose homeostasis, the metabolic fate and function of glucose in BAT remain elusive as there is clear dissociation between glucose uptake and BAT thermogenesis. Interestingly, intracellular glycolysis and lactate production appear to be required for glucose uptake by BAT. Here, we specifically examine whether activation of lactate receptors in BAT plays a key role in regulating glucose homeostasis in mice fed a high-fat diet (HFD). When C57BL/6J mice are given HFD for 5 weeks at 28°C, male, but not female, mice gain body weight and develop hyperglycemia. Importantly, high-fat feeding upregulates expression of the lactate receptor hydroxycarboxylic acid receptor 1 (HCAR1) in female C57BL/6J mice, whereas male C57BL/6J mice show reduced HCAR1 expression in BAT. Treatment with the HCAR1 agonist lowers systemic glucose levels in male DIO mice. This reduction is associated with increased glucose uptake in BAT. Therefore, our results suggest that HCAR1 in BAT may contribute to the development of hyperglycemia in male C57BL/6J DIO mice.

Activation of temperature-sensitive TRPV1-like receptors in ARC POMC neurons reduces food intake.

Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC) respond to numerous hormonal and neural signals, resulting in changes in food intake. Here, we demonstrate that ARC POMC neurons express capsaicin-sensitive transient receptor potential vanilloid 1 receptor (TRPV1)-like receptors. To show expression of TRPV1-like receptors in ARC POMC neurons, we use single-cell reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, electrophysiology, TRPV1 knock-out (KO), and TRPV1-Cre knock-in mice. A small elevation of temperature in the physiological range is enough to depolarize ARC POMC neurons. This depolarization is blocked by the TRPV1 receptor antagonist and by Trpv1 gene knockdown. Capsaicin-induced activation reduces food intake that is abolished by a melanocortin receptor antagonist. To selectively stimulate TRPV1-like receptor-expressing ARC POMC neurons in the ARC, we generate an adeno-associated virus serotype 5 (AAV5) carrying a Cre-dependent channelrhodopsin-2 (ChR2)-enhanced yellow fluorescent protein (eYFP) expression cassette under the control of the two neuronal POMC enhancers (nPEs). Optogenetic stimulation of TRPV1-like receptor-expressing POMC neurons decreases food intake. Hypothalamic temperature is rapidly elevated and reaches to approximately 39 °C during treadmill running. This elevation is associated with a reduction in food intake. Knockdown of the Trpv1 gene exclusively in ARC POMC neurons blocks the feeding inhibition produced by increased hypothalamic temperature. Taken together, our findings identify a melanocortinergic circuit that links acute elevations in hypothalamic temperature with acute reductions in food intake.

Intracellular glycolysis in brown adipose tissue is essential for optogenetically induced nonshivering thermogenesis in mice.

Release of fatty acids from lipid droplets upon activation of the sympathetic nervous system (SNS) is a key step in nonshivering thermogenesis in brown adipose tissue (BAT). However, intracellular lipolysis appears not to be critical for cold-induced thermogenesis. As activation of the SNS increases glucose uptake, we studied whether intracellular glycolysis plays a role in BAT thermogenesis. To stimulate BAT-innervating sympathetic nerves in vivo, we expressed channelrhodopsin-2 (ChR2) in catecholaminergic fibers by crossbreeding tyrosine hydroxylase-Cre mice with floxed-stop ChR2 mice. Acute optogenetic stimulation of sympathetic efferent fibers of BAT increased body temperature and lowered blood glucose levels that were completely abolished by the ß-adrenergic receptor antagonist. Knockdown of the Ucp1 gene in BAT blocked the effects of optogenetic stimulation on body temperature and glucose uptake. Inhibition of glucose uptake in BAT and glycolysis abolished optogenetically induced thermogenesis. Stimulation of sympathetic nerves upregulated expression of the lactate dehydrogenase-A and -B genes in BAT. Optogenetic stimulation failed to induce thermogenesis following treatment with the LDH inhibitor. Pharmacological blockade and genetic deletion of the monocarboxylate transporter 1 completely abolished the effects of sympathetic activation. Our results suggest that intracellular glycolysis and lactate shuttle play an important role in regulating acute thermogenesis in BAT.

A gut-brain axis regulating glucose metabolism mediated by bile acids and competitive fibroblast growth factor actions at the hypothalamus.

OBJECTIVE: Bile acids have been implicated as important regulators of glucose metabolism via activation of FXR and GPBAR1. We have previously shown that FGF19 can modulate glucose handling by suppressing the activity of hypothalamic AGRP/NPY neurons. As bile acids stimulate the release of FGF19/FGF15 into the circulation, we pursued the potential of bile acids to improve glucose tolerance via a gut-brain axis involving FXR and FGF15/FGF19 within enterocytes and FGF receptors on hypothalamic AGRP/NPY neurons. METHODS: A 5-day gavage of taurocholic acid, mirroring our previous protocol of a 5-day FGF19 treatment, was performed. Oral glucose tolerance tests in mice with genetic manipulations of FGF signaling and melanocortin signaling were used to define a gut-brain axis responsive to bile acids. RESULTS: The taurocholic acid gavage led to increased serum concentrations of taurocholic acid as well as increases of FGF15 mRNA in the ileum and improved oral glucose tolerance in obese (ob/ob) mice. In contrast, lithocholic acid, an FXR antagonist but a potent agonist for GPBAR1, did not improve glucose tolerance. The positive response to taurocholic acid is dependent upon an intact melanocortinergic system as obese MC4R-null mice or ob/ob mice without AGRP did not show improvements in glucose tolerance after taurocholate gavage. We also tested the FGF receptor isoform necessary for the bile acid response, using AGRP:Fgfr1-/- and AGRP:Fgfr2-/- mice. While the absence of FGFR1 in AGRP/NPY neurons did not alter glucose tolerance after taurocholate gavage, manipulations of Fgfr2 caused bidirectional changes depending upon the experimental model. We hypothesized the existence of an endogenous hypothalamic FGF, most likely FGF17, that acted as a chronic activator of AGRP/NPY neurons. We developed two short peptides based on FGF8 and FGF17 that should antagonize FGF17 action. Both of these peptides improved glucose homeostasis after a 4-day course of central and peripheral injections. Significantly, daily average blood glucose from continuous glucose monitoring was reduced in all tested animals but glucose concentrations remained in the euglycemia range. CONCLUSIONS: We have defined a gut-brain axis that regulates glucose metabolism mediated by antagonistic fibroblast growth factors. From the intestine, bile acids stimulate FGF15 secretion, leading to activation of the FGF receptors in hypothalamic AGRP/NPY neurons. FGF receptor intracellular signaling subsequently silences AGRP/NPY neurons, leading to improvements of glucose tolerance that are likely mediated by the autonomic nervous system. Finally, short peptides that antagonize homodimeric FGF receptor signaling within the hypothalamus have beneficial effects on glucose homeostasis without inducing hypoglycemia. These peptides could provide a new mode of regulating glucose metabolism.

Cholinergic neurons in the dorsomedial hypothalamus regulate food intake.

OBJECTIVE: Central cholinergic neural circuits play a role in the regulation of feeding behavior. The dorsomedial hypothalamus (DMH) is considered the appetite-stimulating center and contains cholinergic neurons. Here, we study the role of DMH cholinergic neurons in the control of food intake. METHODS: To selectively stimulate DMH cholinergic neurons, we expressed stimulatory designer receptors exclusively activated by designer drugs (DREADDs) and channelrhodopsins in DMH cholinergic neurons by injection of adeno-associated virus (AAV) vectors into the DMH of choline acetyltransferase (ChAT)-IRES-Cre mice. We also generated transgenic mice expressing channelrhodopsins in cholinergic neurons with the Cre-LoxP technique. To delete the Chat gene exclusively in the DMH, we injected an AAV carrying a Cre recombinase transgene into the DMH of floxed ChAT mice. Food intake was measured with and without selective stimulation of DMH cholinergic neurons. RESULTS: Mice lacking the Chat gene in the DMH show reduced body weight as compared to control. Chemogenetic activation of DMH cholinergic neurons promotes food intake. This orexigenic effect is further supported by experiments of optogenetic stimulation of DMH cholinergic neurons. DMH cholinergic neurons innervate pro-opiomelanocortin neurons in the arcuate nucleus of the hypothalamus (ARC). Treatment with acetylcholine (ACh) enhances GABAergic inhibitory transmission to ARC POMC neurons that is blocked by the muscarinic receptor antagonist. Direct activation of cholinergic fibers in the ARC readily stimulates food intake that is also abolished by the muscarinic receptor antagonist. CONCLUSION: ACh released from DMH cholinergic neurons regulates food intake and body weight. This effect is mediated in part through regulation of ARC POMC neurons. Activation of muscarinic receptors on GABAergic axon terminals enhances inhibitory tone to ARC POMC neurons. Hence, this novel DMHACh â†’ ARCPOMC pathway plays an important role in the control of food intake and body weight.

Single-Cell Gene Expression Analysis of Cholinergic Neurons in the Arcuate Nucleus of the Hypothalamus.

The cholinoceptive system in the hypothalamus, in particular in the arcuate nucleus (ARC), plays a role in regulating food intake. Neurons in the ARC contain multiple neuropeptides, amines, and neurotransmitters. To study molecular and neurochemical heterogeneity of ARC neurons, we combine single-cell qRT-PCR and single-cell whole transcriptome amplification methods to analyze expression patterns of our hand-picked 60 genes in individual neurons in the ARC. Immunohistochemical and single-cell qRT-PCR analyses show choline acetyltransferase (ChAT)-expressing neurons in the ARC. Gene expression patterns are remarkably distinct in each individual cholinergic neuron. Two-thirds of cholinergic neurons express tyrosine hydroxylase (Th) mRNA. A large subset of these Th-positive cholinergic neurons is GABAergic as they express the GABA synthesizing enzyme glutamate decarboxylase and vesicular GABA transporter transcripts. Some cholinergic neurons also express the vesicular glutamate transporter transcript gene. POMC and POMC-processing enzyme transcripts are found in a subpopulation of cholinergic neurons. Despite this heterogeneity, gene expression patterns in individual cholinergic cells appear to be highly regulated in a cell-specific manner. In fact, membrane receptor transcripts are clustered with their respective intracellular signaling and downstream targets. This novel population of cholinergic neurons may be part of the neural circuitries that detect homeostatic need for food and control the drive to eat.

Cholinergic neurons in the dorsomedial hypothalamus regulate mouse brown adipose tissue metabolism.

OBJECTIVE: Brown adipose tissue (BAT) thermogenesis is critical in maintaining body temperature. The dorsomedial hypothalamus (DMH) integrates cutaneous thermosensory signals and regulates adaptive thermogenesis. Here, we study the function and synaptic connectivity of input from DMH cholinergic neurons to sympathetic premotor neurons in the raphe pallidus (Rpa). METHODS: In order to selectively manipulate DMH cholinergic neuron activity, we generated transgenic mice expressing channelrhodopsin fused to yellow fluorescent protein (YFP) in cholinergic neurons (choline acetyltransferase (ChAT)-Cre::ChR2-YFP) with the Cre-LoxP technique. In addition, we used an adeno-associated virus carrying the Cre recombinase gene to delete the floxed Chat gene in the DMH. Physiological studies in response to optogenetic stimulation of DMH cholinergic neurons were combined with gene expression and immunocytochemical analyses. RESULTS: A subset of DMH neurons are ChAT-immunopositive neurons. The activity of these neurons is elevated by warm ambient temperature. A phenotype-specific neuronal tracing shows that DMH cholinergic neurons directly project to serotonergic neurons in the Rpa. Optical stimulation of DMH cholinergic neurons decreases BAT activity, which is associated with reduced body core temperature. Furthermore, elevated DMH cholinergic neuron activity decreases the expression of BAT uncoupling protein 1 (Ucp1) and peroxisome proliferator-activated receptor γ coactivator 1 α (Pgc1α) mRNAs, markers of BAT activity. Injection of M2-selective muscarinic receptor antagonists into the 4th ventricle abolishes the effect of optical stimulation. Single cell qRT-PCR analysis of retrogradely identified BAT-projecting neurons in the Rpa shows that all M2 receptor-expressing neurons contain tryptophan hydroxylase 2. In animals lacking the Chat gene in the DMH, exposure to warm temperature reduces neither BAT Ucp1 nor Pgc1α mRNA expression. CONCLUSION: DMH cholinergic neurons directly send efferent signals to sympathetic premotor neurons in the Rpa. Elevated cholinergic input to this area reduces BAT activity through activation of M2 mAChRs on serotonergic neurons. Therefore, the direct DMH(ACh)-Rpa(5-HT) pathway may mediate physiological heat-defense responses to elevated environmental temperature.

Interplay between glucose and leptin signalling determines the strength of GABAergic synapses at POMC neurons.

Regulation of GABAergic inhibitory inputs and alterations in POMC neuron activity by nutrients and adiposity signals regulate energy and glucose homeostasis. Thus, understanding how POMC neurons integrate these two signal molecules at the synaptic level is important. Here we show that leptin's action on GABA release to POMC neurons is influenced by glucose levels. Leptin stimulates the JAK2-PI3K pathway in both presynaptic GABAergic terminals and postsynaptic POMC neurons. Inhibition of AMPK activity in presynaptic terminals decreases GABA release at 10 mM glucose. However, postsynaptic TRPC channel opening by the PI3K-PLC signalling pathway in POMC neurons enhances spontaneous GABA release via activation of presynaptic MC3/4 and mGlu receptors at 2.5 mM glucose. High-fat feeding blunts AMPK-dependent presynaptic inhibition, whereas PLC-mediated GABAergic feedback inhibition remains responsive to leptin. Our data indicate that the interplay between glucose and leptin signalling in glutamatergic POMC neurons is critical for determining the strength of inhibitory tone towards POMC neurons.

Apelin-13 enhances arcuate POMC neuron activity via inhibiting M-current.

The hypothalamus is a key element of the neural circuits that control energy homeostasis. Specific neuronal populations within the hypothalamus are sensitive to a variety of homeostatic indicators such as circulating nutrient levels and hormones that signal circulating glucose and body fat content. Central injection of apelin secreted by adipose tissues regulates feeding and glucose homeostasis. However, the precise neuronal populations and cellular mechanisms involved in these physiological processes remain unclear. Here we examine the electrophysiological impact of apelin-13 on proopiomelanocortin (POMC) neuron activity. Approximately half of POMC neurons examined respond to apelin-13. Apelin-13 causes a dose-dependent depolarization. This effect is abolished by the apelin (APJ) receptor antagonist. POMC neurons from animals pre-treated with pertussis toxin still respond to apelin, whereas the Gßγ signaling inhibitor gallein blocks apelin-mediated depolarization. In addition, the effect of apelin is inhibited by the phospholipase C and protein kinase inhibitors. Furthermore, single-cell qPCR analysis shows that POMC neurons express the APJ receptor, PLC-ß isoforms, and KCNQ subunits (2, 3 and 5) which contribute to M-type current. Apelin-13 inhibits M-current that is blocked by the KCNQ channel inhibitor. Therefore, our present data indicate that apelin activates APJ receptors, and the resultant dissociation of the Gαq heterotrimer triggers a Gßγ-dependent activation of PLC-ß signaling that inhibits M-current.

Central action of FGF19 reduces hypothalamic AGRP/NPY neuron activity and improves glucose metabolism.

Tight control of glucose excursions has been a long-standing goal of treatment for patients with type 2 diabetes mellitus in order to ameliorate the morbidity and mortality associated with hyperglycemia. Fibroblast growth factor (FGF) 19 is a hormone-like enterokine released postprandially that emerged as a potential therapeutic agent for metabolic disorders, including diabetes and obesity. Remarkably, FGF19 treatment has hypoglycemic actions that remain potent in models of genetic and acquired insulin resistance. Here, we provided evidence that the central nervous system responds to FGF19 administered in the periphery. Then, in two mouse models of insulin resistance, leptin-deficiency and high-fat diet feeding, third intra-cerebro-ventricular infusions of FGF19 improved glycemic status, reduced insulin resistance and potentiated insulin signaling in the periphery. In addition, our study highlights a new mechanism of central FGF19 action, involving the suppression of AGRP/NPY neuronal activity. Overall, our work unveils novel regulatory pathways induced by FGF19 that will be useful in the design of novel strategies to control diabetes in obesity.

Clusterin and LRP2 are critical components of the hypothalamic feeding regulatory pathway.

Hypothalamic feeding circuits are essential for the maintenance of energy balance. There have been intensive efforts to discover new biological molecules involved in these pathways. Here we report that central administration of clusterin, also called apolipoprotein J, causes anorexia, weight loss and activation of hypothalamic signal transduction-activated transcript-3 in mice. In contrast, inhibition of hypothalamic clusterin action results in increased food intake and body weight, leading to adiposity. These effects are likely mediated through the mutual actions of the low-density lipoprotein receptor-related protein-2, a potential receptor for clusterin, and the long-form leptin receptor. In response to clusterin, the low-density lipoprotein receptor-related protein-2 binding to long-form leptin receptor is greatly enhanced in cultured neuronal cells. Furthermore, long-form leptin receptor deficiency or hypothalamic low-density lipoprotein receptor-related protein-2 suppression in mice leads to impaired hypothalamic clusterin signalling and actions. Our study identifies the hypothalamic clusterin-low-density lipoprotein receptor-related protein-2 axis as a novel anorexigenic signalling pathway that is tightly coupled with long-form leptin receptor-mediated signalling.

Overnight fasting regulates inhibitory tone to cholinergic neurons of the dorsomedial nucleus of the hypothalamus.

The dorsomedial nucleus of the hypothalamus (DMH) contributes to the regulation of overall energy homeostasis by modulating energy intake as well as energy expenditure. Despite the importance of the DMH in the control of energy balance, DMH-specific genetic markers or neuronal subtypes are poorly defined. Here we demonstrate the presence of cholinergic neurons in the DMH using genetically modified mice that express enhanced green florescent protein (eGFP) selectively in choline acetyltransferase (Chat)-neurons. Overnight food deprivation increases the activity of DMH cholinergic neurons, as shown by induction of fos protein and a significant shift in the baseline resting membrane potential. DMH cholinergic neurons receive both glutamatergic and GABAergic synaptic input, but the activation of these neurons by an overnight fast is due entirely to decreased inhibitory tone. The decreased inhibition is associated with decreased frequency and amplitude of GABAergic synaptic currents in the cholinergic DMH neurons, while glutamatergic synaptic transmission is not altered. As neither the frequency nor amplitude of miniature GABAergic or glutamatergic postsynaptic currents is affected by overnight food deprivation, the fasting-induced decrease in inhibitory tone to cholinergic neurons is dependent on superthreshold activity of GABAergic inputs. This study reveals that cholinergic neurons in the DMH readily sense the availability of nutrients and respond to overnight fasting via decreased GABAergic inhibitory tone. As such, altered synaptic as well as neuronal activity of DMH cholinergic neurons may play a critical role in the regulation of overall energy homeostasis.