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.

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.

pRb is an obesity suppressor in hypothalamus and high-fat diet inhibits pRb in this location.

pRb is frequently inactivated in tumours by mutations or phosphorylation. Here, we investigated whether pRb plays a role in obesity. The Arcuate nucleus (ARC) in hypothalamus contains antagonizing POMC and AGRP/NPY neurons for negative and positive energy balance, respectively. Various aspects of ARC neurons are affected in high-fat diet (HFD)-induced obesity mouse model. Using this model, we show that HFD, as well as pharmacological activation of AMPK, induces pRb phosphorylation and E2F target gene de-repression in ARC neurons. Some affected neurons express POMC; and deleting Rb1 in POMC neurons induces E2F target gene de-repression, cell-cycle re-entry, apoptosis, and a hyperphagia-obesity-diabetes syndrome. These defects can be corrected by combined deletion of E2f1. In contrast, deleting Rb1 in the antagonizing AGRP/NPY neurons shows no effects. Thus, pRb-E2F1 is an obesity suppression mechanism in ARC POMC neurons and HFD-AMPK inhibits this mechanism by phosphorylating pRb in this location.

Differential transcriptional profiles of vagal sensory neurons in female and male mice.

Introduction: Differences in metabolic homeostasis, diabetes, and obesity between males and females are evident in rodents and humans. Vagal sensory neurons in the vagus nerve ganglia innervate a variety of visceral organs and use specialized nerve endings to sense interoceptive signals. This visceral organ-brain axis plays a role in relaying interoceptive signals to higher brain centers, as well as in regulating the vago-vagal reflex. I hypothesized that molecularly distinct populations of vagal sensory neurons would play a role in causing differences in metabolic homeostasis between the sexes. Methods: SnRNA-Seq was conducted on dissociated cells from the vagus nerve ganglia using the 10X Genomics Chromium platform. Results: Single-nucleus RNA sequencing analysis of vagal sensory neurons from female and male mice revealed differences in the transcriptional profiles of cells in the vagus nerve ganglia. These differences are linked to the expression of sex-specific genes such as Xist, Tsix, and Ddx3y. Among the 13 neuronal clusters, one-fourth of the neurons in male mice were located in the Ddx3y-enriched VN1 and VN8 clusters, which displayed higher enrichment of Trpv1, Piezo2, Htr3a, and Vip genes. In contrast, 70% of the neurons in females were found in Xist-enriched clusters VN4, 6, 7, 10, 11, and 13, which showed enriched genes such as Fgfr1, Lpar1, Cpe, Esr1, Nrg1, Egfr, and Oprm1. Two clusters of satellite cells were identified, one of which contained oligodendrocyte precursor cells in male mice. A small population of cells expressed Ucp1 and Plin1, indicating that they are epineural adipocytes. Discussion: Understanding the physiological implications of distinct transcriptomic profiles in vagal sensory neurons on energy balance and metabolic homeostasis would help develop sex-specific treatments for obesity and metabolic dysregulation.

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.

The development of hepatic steatosis depends on the presence of liver-innervating parasympathetic cholinergic neurons in mice fed a high-fat diet.

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 cholangiocytes are innervated by parasympathetic nerve terminals 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 promotes browning of inguinal white adipose tissue (ingWAT). The loss of liver-innervating cholinergic neurons increases 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 liver-innervating cholinergic neurons, leading to the reappearance of hepatic steatosis. Deleting liver-innervating cholinergic neurons has a small but significant effect on body weight, which is accompanied by an increase in energy expenditure. Taken together, these data suggest that targeting the parasympathetic cholinergic innervation of the liver is a potential therapeutic approach for enhancing hepatic lipid metabolism in obesity and diabetes.

TXNIP in Agrp neurons regulates adiposity, energy expenditure, and central leptin sensitivity.

Thioredoxin interacting protein (TXNIP) has recently been described as a key regulator of energy metabolism through pleiotropic actions that include nutrient sensing in the mediobasal hypothalamus (MBH). However, the role of TXNIP in neurochemically specific hypothalamic subpopulations and the circuits downstream from MBH TXNIP engaged to regulate energy homeostasis remain unexplored. To evaluate the metabolic role of TXNIP activity specifically within arcuate Agrp neurons, we generated Agrp-specific TXNIP gain-of-function and loss-of-function mouse models using Agrp-Ires-cre mice, TXNIP (flox/flox) mice, and a lentivector expressing the human TXNIP isoform conditionally in the presence of Cre recombinase. Overexpression of TXNIP in Agrp neurons predisposed to diet-induced obesity and adipose tissue storage by decreasing energy expenditure and spontaneous locomotion, without affecting food intake. Conversely, Agrp neuronal TXNIP deletion protected against diet-induced obesity and adipose tissue storage by increasing energy expenditure and spontaneous locomotion, also without affecting food intake. TXNIP overexpression in Agrp neurons did not primarily affect glycemic control, whereas deletion of TXNIP in Agrp neurons improved fasting glucose levels and glucose tolerance independently of its effects on body weight and adiposity. Bidirectional manipulation of TXNIP expression induced reciprocal changes in central leptin sensitivity and the neural regulation of lipolysis. Together, these results identify a critical role for TXNIP in Agrp neurons in mediating diet-induced obesity through the regulation of energy expenditure and adipose tissue metabolism, independently of food intake. They also reveal a previously unidentified role for Agrp neurons in the brain-adipose axis.

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.

Cross-talk between P2X4 and gamma-aminobutyric acid, type A receptors determines synaptic efficacy at a central synapse.

The essence of neuronal function is to generate outputs in response to synaptic potentials. Synaptic integration at postsynaptic sites determines neuronal outputs in the CNS. Using immunohistochemical and electrophysiological approaches, we first reveal that steroidogenic factor 1 (SF-1) green fluorescent protein (GFP)-positive neurons in the ventromedial nucleus of the hypothalamus express P2X4 subunits that are activated by exogenous ATP. Increased membrane expression of P2X4 channels by using a peptide competing with P2X4 intracellular endocytosis motif enhances neuronal excitability of SF-1 GFP-positive neurons. This increased excitability is inhibited by a P2X receptor antagonist. Furthermore, increased surface P2X4 receptor expression significantly decreases the frequency and the amplitude of GABAergic postsynaptic currents of SF-1 GFP-positive neurons. Co-immunopurification and pulldown assays reveal that P2X4 receptors complex with aminobutyric acid, type A (GABA(A)) receptors and demonstrate that two amino acids in the carboxyl tail of the P2X4 subunit are crucial for its physical association with GABA(A) receptors. Mutation of these two residues prevents the physical association, thereby blocking cross-inhibition between P2X4 and GABA(A) receptors. Moreover, disruption of the physical coupling using competitive peptides containing the identified motif abolishes current inhibition between P2X4 and GABA(A) receptors in recombinant system and P2X4 receptor-mediated GABAergic depression in SF-1 GFP-positive neurons. Our present work thus provides evidence for cross-talk between excitatory and inhibitory receptors that appears to be crucial in determining GABAergic synaptic strength at a central synapse.

Endogenous BDNF regulates inhibitory synaptic transmission in the ventromedial nucleus of the hypothalamus.

Output from steroidogenic factor-1 (SF-1) neurons in the ventromedial nucleus of the hypothalamus (VMH) is anorexigenic. SF-1 neurons express brain-derived neurotrophic factor (BDNF) that contributes to the regulation of food intake and body weight. Here I show that regulation of GABAergic inputs onto SF-1 neurons by endogenous BDNF determines the anorexigenic outcome from the VMH. Single-cell RT-PCR analysis reveals that one-third of SF-1 neurons express BDNF and that only a subset of BDNF-expressing SF-1 neurons coexpresses the melanocortin receptor type 4. Whole cell patch-clamp analysis of SF-1 neurons in the VMH shows that exogenous BDNF significantly increases the frequency of spontaneous GABAergic inhibitory postsynaptic currents (sIPSCs). This enhancement of GABA drive readily decreases the excitability of SF-1 neurons. However, treatment with BDNF has no significant effect on the frequency of TTX-independent GABAergic IPSCs. Moreover, TrkB receptors are not localized at the postsynaptic sites of GABAergic synapses on SF-1 neurons as there is no change in the amplitude of miniature IPSCs in the presence of BDNF. Dual patch-clamp recordings in mouse hypothalamic slices reveal that stimulation of one SF-1 neuron induces an increase in sIPSC frequency onto the neighboring SF-1 neuron. More importantly, this effect is blocked by a tyrosine kinase inhibitor. Hence, this increased GABA drive onto SF-1 neurons may, in part, explain the cellular mechanisms that mediate the anorexigenic effects of BDNF.

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.

Mediobasal hypothalamic leucine sensing regulates food intake through activation of a hypothalamus-brainstem circuit.

In response to nutrient stimuli, the mediobasal hypothalamus (MBH) drives multiple neuroendocrine and behavioral mechanisms to regulate energy balance. While central leucine reduces food intake and body weight, the specific neuroanatomical sites of leucine sensing, downstream neural substrates, and neurochemical effectors involved in this regulation remain largely unknown. Here we demonstrate that MBH leucine engages a neural energy regulatory circuit by stimulating POMC (proopiomelanocortin) neurons of the MBH, oxytocin neurons of the paraventricular hypothalamus, and neurons within the brainstem nucleus of the solitary tract to acutely suppress food intake by reducing meal size. We identify central p70 S6 kinase and Erk1/2 pathways as intracellular effectors required for this response. Activation of endogenous leucine intracellular metabolism produced longer-term reductions in meal number. Our data identify a novel, specific hypothalamus-brainstem circuit that links amino acid availability and nutrient sensing to the control of food intake.

Direct innervation of GnRH neurons by metabolic- and sexual odorant-sensing leptin receptor neurons in the hypothalamic ventral premammillary nucleus.

Leptin acts via its receptor (LepRb) on specific CNS neurons to signal the adequacy of long-term energy stores, thereby permitting the expenditure of resources on energy-intensive processes such as reproduction. The ventral premammillary nucleus of the hypothalamus (PMv), which has been implicated in the stimulation of gonadotropin release by olfactory cues, contains numerous LepRb neurons, suggesting a potential role for LepRb PMv neurons in transmitting both metabolic and odorant signals to the neuroendocrine reproductive system. Indeed, Fos immunoreactivity and electrophysiologic recordings revealed the direct activation of LepRb PMv neurons by leptin, and exposure to odors from mice of the opposite sex promoted Fos immunoreactivity (Fos-IR) in many LepRb PMv neurons. To determine the regions innervated by the LepRb PMv neurons, we used two novel cre-activated tract-tracing systems in Lepr(cre) animals; data from these systems and from standard tracing techniques revealed that LepRb PMv neurons project to a subset of the regions, including the preoptic area, that are innervated by the PMv as a whole. Furthermore, the retrograde accumulation in LepRb PMv neurons of a trans-synaptic tracer from GnRH neurons revealed the direct innervation of GnRH neurons by many LepRb PMv neurons. Thus, LepRb PMv neurons sense metabolic and sexual odorant cues and project to the rostral hypothalamus to directly innervate GnRH neurons. These results are consistent with a role for LepRb PMv neurons in regulating the reproductive axis in response to metabolic and odorant stimuli.

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.

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.

Integration of endocannabinoid and leptin signaling in an appetite-related neural circuit.

Recently developed therapeutics for obesity, targeted against cannabinoid receptors, result in decreased appetite and sustained weight loss. Prior studies have demonstrated CB1 receptors (CB1Rs) and leptin modulation of cannabinoid synthesis in hypothalamic neurons. Here, we show that depolarization of perifornical lateral hypothalamus (LH) neurons elicits a CB1R-mediated suppression of inhibition in local circuits thought to be involved in appetite and "natural reward." The depolarization-induced decrease in inhibitory tone to LH neurons is blocked by leptin. Leptin inhibits voltage-gated calcium channels in LH neurons via the activation of janus kinase 2 (JAK2) and of mitogen-activated protein kinase (MAPK). Leptin-deficient mice are characterized by both an increase in steady-state voltage-gated calcium currents in LH neurons and a CB1R-mediated depolarization-induced suppression of inhibition that is 6-fold longer than that in littermate controls. Our data provide direct electrophysiological support for the involvement of endocannabinoids and leptin as modulators of hypothalamic circuits underlying motivational aspects of feeding behavior.

Transcription factors in the development of medial hypothalamic structures.

The hypothalamus has historically been subdivided into nuclei, agglomerations of cell bodies that are visually distinct in histological sections. Regulatory functions of metabolism have been assigned to the various hypothalamic nuclei principally by analysis of animals with lesions of individual nuclei but also via various means of stimulation, such as cooling or heating probes. Biochemical and molecular specificity of these studies became possible with the identification and synthesis of neurotransmitters as well as the means to manipulate the expression of endogenous neurotransmitters and their receptors by genetic means . The arcuate nucleus (ARC) is likely to be the primary site for neurons that sense circulating fuels and energy reserves (POMC/CART neurons, NPY/AGRP neurons), whereas the paraventricular nucleus (PVN) receives input from the ARC and harbors many of the releasing factors (CRF, TRH, vasopressin, and oxytocin) that control pituitary hormone release. The ventromedial nucleus (VMN) receives input from the ARC and plays a critical role in energy balance in parallel with the ARC. The VMN and PVN also send descending projections to the autonomic nervous system and other pathways that control ingestive behavior and metabolism. Developmental analyses have revealed that the neurons that comprise the hypothalamic nuclei arise by differentiation and migration from stem cells within the ventricular zone. Based on recent work, it is becoming clear that coordination between numerous transcription factors that determine specification, survival, and migration is necessary for the formation of the hypothalamus, with each nucleus being determined by its own unique set of factors. In this minireview, we will provide a selective view of the roles that transcription factors play in the developing hypothalamus.

Steroidogenic factor 1 regulates expression of the cannabinoid receptor 1 in the ventromedial hypothalamic nucleus.

The nuclear receptor steroidogenic factor 1 (SF-1) plays essential roles in the development and function of the ventromedial hypothalamic nucleus (VMH). Considerable evidence links the VMH and SF-1 with the regulation of energy homeostasis. Here, we demonstrate that SF-1 colocalizes in VMH neurons with the cannabinoid receptor 1 (CB1R) and that a specific CB1R agonist modulates electrical activity of SF-1 neurons in hypothalamic slice preparations. We further show that SF-1 directly regulates CB1R gene expression via a SF-1-responsive element at -101 in its 5'-flanking region. Finally, we show that knockout mice with selective inactivation of SF-1 in the brain have decreased expression of CB1R in the region of the VMH and exhibit a blunted response to systemically administered CB1R agonists. These studies suggest that SF-1 directly regulates the expression of CB1R, which has been implicated in the regulation of energy homeostasis and anxiety-like behavior.

Collective and individual functions of leptin receptor modulated neurons controlling metabolism and ingestion.

Two known types of leptin-responsive neurons reside within the arcuate nucleus: the agouti gene-related peptide (AgRP)/neuropeptide Y (NPY) neuron and the proopiomelanocortin (POMC) neuron. By deleting the leptin receptor gene (Lepr) specifically in AgRP/NPY and/or POMC neurons of mice, we examined the several and combined contributions of these neurons to leptin action. Body weight and adiposity were increased by Lepr deletion from AgRP and POMC neurons individually, and simultaneous deletion in both neurons (A+P LEPR-KO mice) further increased these measures. Young (periweaning) A+P LEPR-KO mice exhibit hyperphagia and decreased energy expenditure, with increased weight gain, oxidative sparing of triglycerides, and increased fat accumulation. Interestingly, however, many of these abnormalities were attenuated in adult animals, and high doses of leptin partially suppress food intake in the A+P LEPR-KO mice. Although mildly hyperinsulinemic, the A+P LEPR-KO mice displayed normal glucose tolerance and fertility. Thus, AgRP/NPY and POMC neurons each play mandatory roles in aspects of leptin-regulated energy homeostasis, high leptin levels in adult mice mitigate the importance of leptin-responsiveness in these neurons for components of energy balance, suggesting the presence of other leptin-regulated pathways that partially compensate for the lack of leptin action on the POMC and AgRP/NPY neurons.