Leptin and insulin act on POMC neurons to promote the browning of white fat.

The primary task of white adipose tissue (WAT) is the storage of lipids. However, "beige" adipocytes also exist in WAT. Beige adipocytes burn fat and dissipate the energy as heat, but their abundance is diminished in obesity. Stimulating beige adipocyte development, or WAT browning, increases energy expenditure and holds potential for combating metabolic disease and obesity. Here, we report that insulin and leptin act together on hypothalamic neurons to promote WAT browning and weight loss. Deletion of the phosphatases PTP1B and TCPTP enhanced insulin and leptin signaling in proopiomelanocortin neurons and prevented diet-induced obesity by increasing WAT browning and energy expenditure. The coinfusion of insulin plus leptin into the CNS or the activation of proopiomelanocortin neurons also increased WAT browning and decreased adiposity. Our findings identify a homeostatic mechanism for coordinating the status of energy stores, as relayed by insulin and leptin, with the central control of WAT browning.

Sympathetic Innervation of White Adipose Tissue: to Beige or Not to Beige?

Obesity research progresses in understanding neuronal circuits and adipocyte biology to regulate metabolism. However, the interface of neuro-adipocyte interaction is less studied. We summarize the current knowledge of adipose tissue innervation and interaction with adipocytes and emphasize adipocyte transitions from white to brown adipocytes and vice versa. We further highlight emerging concepts for the differential neuronal regulation of brown/beige versus white adipocyte and the interdependence of both for metabolic regulation.

Maternal cold exposure induces distinct transcriptome changes in the placenta and fetal brown adipose tissue in mice.

BACKGROUND: Brown adipose tissue (BAT) is specialized to dissipate energy in the form of heat. BAT-mediated heat production in rodents and humans is critical for effective temperature adaptation of newborns to the extrauterine environment immediately after birth. However, very little is known about whether and how fetal BAT development is modulated in-utero in response to changes in maternal thermal environment during pregnancy. Using BL6 mice, we evaluated the impact of different maternal environmental temperatures (28 °C and 18 °C) on the transcriptome of the placenta and fetal BAT to test if maternal cold exposure influences fetal BAT development via placental remodeling. RESULTS: Maternal weight gain during pregnancy, the average number of fetuses per pregnancy, and placental weight did not differ between the groups at 28 °C and 18 °C. However, the average fetal weight at E18.5 was 6% lower in the 18 °C-group compared to the 28 °C-group. In fetal BATs, cold exposure during pregnancy induced increased expression of genes involved in de novo lipogenesis and lipid metabolism while decreasing the expression of genes associated with muscle cell differentiation, thus suggesting that maternal cold exposure may promote fetal brown adipogenesis by suppressing the myogenic lineage in bidirectional progenitors. In placental tissues, maternal cold exposure was associated with upregulation of genes involved in complement activation and downregulation of genes related to muscle contraction and actin-myosin filament sliding. These changes may coordinate placental adaptation to maternal cold exposure, potentially by protecting against cold stress-induced inflammatory damage and modulating the vascular and extravascular contractile system in the placenta. CONCLUSIONS: These findings provide evidence that environmental cold temperature sensed by the mother can modulate the transcriptome of placental and fetal BAT tissues. The ramifications of the observed gene expression changes warrant future investigation.

Sympathetic innervation of the mouse kidney and liver arising from prevertebral ganglia.

The sympathetic nervous system (SNS) plays a crucial role in the regulation of renal and hepatic functions. Although sympathetic nerves to the kidney and liver have been identified in many species, specific details are lacking in the mouse. In the absence of detailed information of sympathetic prevertebral innervation of specific organs, selective manipulation of a specific function will remain challenging. Despite providing major postganglionic inputs to abdominal organs, limited data are available about the mouse celiac-superior mesenteric complex. We used tyrosine hydroxylase (TH) and dopamine ß-hydroxylase (DbH) reporter mice to visualize abdominal prevertebral ganglia. We found that both the TH and DbH reporter mice are useful models for identification of ganglia and nerve bundles. We further tested if the celiac-superior mesenteric complex provides differential inputs to the mouse kidney and liver. The retrograde viral tracer, pseudorabies virus (PRV)-152 was injected into the cortex of the left kidney or the main lobe of the liver to identify kidney-projecting and liver-projecting neurons in the celiac-superior mesenteric complex. iDISCO immunostaining and tissue clearing were used to visualize unprecedented anatomical detail of kidney-related and liver-related postganglionic neurons in the celiac-superior mesenteric complex and aorticorenal and suprarenal ganglia compared with TH-positive neurons. Kidney-projecting neurons were restricted to the suprarenal and aorticorenal ganglia, whereas only sparse labeling was observed in the celiac-superior mesenteric complex. In contrast, liver-projecting postganglionic neurons were observed in the celiac-superior mesenteric complex and aorticorenal and suprarenal ganglia, suggesting spatial separation between the sympathetic innervation of the mouse kidney and liver.

Homeostatic sensing of dietary protein restriction: A case for FGF21.

Restriction of dietary protein intake increases food intake and energy expenditure, reduces growth, and alters amino acid, lipid, and glucose metabolism. While these responses suggest that animals 'sense' variations in amino acid consumption, the basic physiological mechanism mediating the adaptive response to protein restriction has been largely undescribed. In this review we make the case that the liver-derived metabolic hormone FGF21 is the key signal which communicates and coordinates the homeostatic response to dietary protein restriction. Support for this model centers on the evidence that FGF21 is induced by the restriction of dietary protein or amino acid intake and is required for adaptive changes in metabolism and behavior. FGF21 occupies a unique endocrine niche, being induced when energy intake is adequate but protein and carbohydrate are imbalanced. Collectively, the evidence thus suggests that FGF21 is the first known endocrine signal of dietary protein restriction.

Galanin-Expressing GABA Neurons in the Lateral Hypothalamus Modulate Food Reward and Noncompulsive Locomotion.

The lateral hypothalamus (LHA) integrates reward and appetitive behavior and is composed of many overlapping neuronal populations. Recent studies associated LHA GABAergic neurons (LHA GABA ), which densely innervate the ventral tegmental area (VTA), with modulation of food reward and consumption; yet, LHA GABA projections to the VTA exclusively modulated food consumption, not reward. We identified a subpopulation of LHA GABA neurons that coexpress the neuropeptide galanin (LHA Gal ). These LHA Gal neurons also modulate food reward, but lack direct VTA innervation. We hypothesized that LHA Gal neurons may represent a subpopulation of LHA GABA neurons that mediates food reward independent of direct VTA innervation. We used chemogenetic activation of LHA Gal or LHA GABA neurons in mice to compare their role in feeding behavior. We further analyzed locomotor behavior to understand how differential VTA connectivity and transmitter release in these LHA neurons influences this behavior. LHA Gal or LHA GABA neuronal activation both increased operant food-seeking behavior, but only activation of LHA GABA neurons increased overall chow consumption. Additionally, LHA Gal or LHA GABA neuronal activation similarly induced locomotor activity, but with striking differences in modality. Activation of LHA GABA neurons induced compulsive-like locomotor behavior; while LHA Gal neurons induced locomotor activity without compulsivity. Thus, LHA Gal neurons define a subpopulation of LHA GABA neurons without direct VTA innervation that mediate noncompulsive food-seeking behavior. We speculate that the striking difference in compulsive-like locomotor behavior is also based on differential VTA innervation. The downstream neural network responsible for this behavior and a potential role for galanin as neuromodulator remains to be identified.SIGNIFICANCE STATEMENT The lateral hypothalamus (LHA) regulates motivated feeding behavior via GABAergic LHA neurons. The molecular identity of LHA GABA neurons is heterogeneous and largely undefined. Here we introduce LHA Gal neurons as a subset of LHA GABA neurons that lack direct innervation of the ventral tegmental area (VTA). LHA Gal neurons are sufficient to drive motivated feeding and locomotor activity similar to LHA GABA neurons, but without inducing compulsive-like behaviors, which we propose to require direct VTA innervation. Our study integrates galanin-expressing LHA neurons into our current understanding of the neuronal circuits and molecular mechanisms of the LHA that contribute to motivated feeding behaviors.

Blaming the Brain for Obesity: Integration of Hedonic and Homeostatic Mechanisms.

The brain plays a key role in the controls of energy intake and expenditure, and many genes associated with obesity are expressed in the central nervous system. Technological and conceptual advances in both basic and clinical neurosciences have expanded the traditional view of homeostatic regulation of body weight by mainly the hypothalamus to include hedonic controls of appetite by cortical and subcortical brain areas processing external sensory information, reward, cognition, and executive functions. Hedonic controls interact with homeostatic controls to regulate body weight in a flexible and adaptive manner that takes environmental conditions into account. This new conceptual framework has several important implications for the treatment of obesity. Because much of this interactive neural processing is outside awareness, cognitive restraint in a world of plenty is made difficult and prevention and treatment of obesity should be more rationally directed to the complex and often redundant mechanisms underlying this interaction.

The Hypothalamic Preoptic Area and Body Weight Control.

The preoptic area (POA) of the hypothalamus is involved in many physiological and behavioral processes thanks to its interconnections to many brain areas and ability to respond to diverse humoral factors. One main function of the POA is to manage body temperature homeostasis, e.g. in response to ambient temperature change, which is achieved in part by controlling brown adipose tissue thermogenesis. The POA is also importantly involved in modulating food intake in response to temperature change, thus making it relevant for body weight homeostasis and obesity research. POA function in body weight control is highly unexplored, and a better understanding of POA circuits and their integration into classic hypothalamic circuits that regulate energy homeostasis is expected to provide new opportunities for the scientific basis and treatment of obesity and comorbidities.

Glutamatergic Preoptic Area Neurons That Express Leptin Receptors Drive Temperature-Dependent Body Weight Homeostasis.

UNLABELLED: The preoptic area (POA) regulates body temperature, but is not considered a site for body weight control. A subpopulation of POA neurons express leptin receptors (LepRb(POA) neurons) and modulate reproductive function. However, LepRb(POA) neurons project to sympathetic premotor neurons that control brown adipose tissue (BAT) thermogenesis, suggesting an additional role in energy homeostasis and body weight regulation. We determined the role of LepRb(POA) neurons in energy homeostasis using cre-dependent viral vectors to selectively activate these neurons and analyzed functional outcomes in mice. We show that LepRb(POA) neurons mediate homeostatic adaptations to ambient temperature changes, and their pharmacogenetic activation drives robust suppression of energy expenditure and food intake, which lowers body temperature and body weight. Surprisingly, our data show that hypothermia-inducing LepRb(POA) neurons are glutamatergic, while GABAergic POA neurons, originally thought to mediate warm-induced inhibition of sympathetic premotor neurons, have no effect on energy expenditure. Our data suggest a new view into the neurochemical and functional properties of BAT-related POA circuits and highlight their additional role in modulating food intake and body weight. SIGNIFICANCE STATEMENT: Brown adipose tissue (BAT)-induced thermogenesis is a promising therapeutic target to treat obesity and metabolic diseases. The preoptic area (POA) controls body temperature by modulating BAT activity, but its role in body weight homeostasis has not been addressed. LepRb(POA) neurons are BAT-related neurons and we show that they are sufficient to inhibit energy expenditure. We further show that LepRb(POA) neurons modulate food intake and body weight, which is mediated by temperature-dependent homeostatic responses. We further found that LepRb(POA) neurons are stimulatory glutamatergic neurons, contrary to prevalent models, providing a new view on thermoregulatory neural circuits. In summary, our study significantly expands our current understanding of central circuits and mechanisms that modulate energy homeostasis.

Neural Control of Energy Expenditure.

The continuous rise in obesity is a major concern for future healthcare management. Many strategies to control body weight focus on a permanent modification of food intake with limited success in the long term. Metabolism or energy expenditure is the other side of the coin for the regulation of body weight, and strategies to enhance energy expenditure are a current focus for obesity treatment, especially since the (re)-discovery of the energy depleting brown adipose tissue in adult humans. Conversely, several human illnesses like neurodegenerative diseases, cancer, or autoimmune deficiency syndrome suffer from increased energy expenditure and severe weight loss. Thus, strategies to modulate energy expenditure to target weight gain or loss would improve life expectancies and quality of life in many human patients. The aim of this book chapter is to give an overview of our current understanding and recent progress in energy expenditure control with specific emphasis on central control mechanisms.

A guide to analysis of mouse energy metabolism.

We present a consolidated view of the complexity and challenges of designing studies for measurement of energy metabolism in mouse models, including a practical guide to the assessment of energy expenditure, energy intake and body composition and statistical analysis thereof. We hope this guide will facilitate comparisons across studies and minimize spurious interpretations of data. We recommend that division of energy expenditure data by either body weight or lean body weight and that presentation of group effects as histograms should be replaced by plotting individual data and analyzing both group and body-composition effects using analysis of covariance (ANCOVA).

GLP-1 receptor signaling is not required for reduced body weight after RYGB in rodents.

Exaggerated GLP-1 and PYY secretion is thought to be a major mechanism in the reduced food intake and body weight after Roux-en-Y gastric bypass surgery. Here, we use complementary pharmacological and genetic loss-of-function approaches to test the role of increased signaling by these gut hormones in high-fat diet-induced obese rodents. Chronic brain infusion of a supramaximal dose of the selective GLP-1 receptor antagonist exendin-9-39 into the lateral cerebral ventricle significantly increased food intake and body weight in both RYGB and sham-operated rats, suggesting that, while contributing to the physiological control of food intake and body weight, central GLP-1 receptor signaling tone is not the critical mechanism uniquely responsible for the body weight-lowering effects of RYGB. Central infusion of the selective Y2R-antagonist BIIE0246 had no effect in either group, suggesting that it is not critical for the effects of RYGB on body weight under the conditions tested. In a recently established mouse model of RYGB that closely mimics surgery and weight loss dynamics in humans, obese GLP-1R-deficient mice lost the same amount of body weight and fat mass and maintained similarly lower body weight compared with wild-type mice. Together, the results surprisingly provide no support for important individual roles of either gut hormone in the specific mechanisms by which RYGB rats settle at a lower body weight. It is likely that the beneficial effects of bariatric surgeries are expressed through complex mechanisms that require combination approaches for their identification.

Leucine acts in the brain to suppress food intake but does not function as a physiological signal of low dietary protein.

Intracerebroventricular injections of leucine are sufficient to suppress food intake, but it remains unclear whether brain leucine signaling represents a physiological signal of protein balance. We tested whether variations in dietary and circulating levels of leucine, or all three branched-chain amino acids (BCAAs), contribute to the detection of reduced dietary protein. Of the essential amino acids (EAAs) tested, only intracerebroventricular injection of leucine (10 µg) was sufficient to suppress food intake. Isocaloric low- (9% protein energy; LP) or normal- (18% protein energy) protein diets induced a divergence in food intake, with an increased consumption of LP beginning on day 2 and persisting throughout the study (P < 0.05). Circulating BCAA levels were reduced the day after LP diet exposure, but levels subsequently increased and normalized by day 4, despite persistent hyperphagia. Brain BCAA levels as measured by microdialysis on day 2 of diet exposure were reduced in LP rats, but this effect was most prominent postprandially. Despite these diet-induced changes in BCAA levels, reducing dietary leucine or total BCAAs independently from total protein was neither necessary nor sufficient to induce hyperphagia, while chronic infusion of EAAs into the brain of LP rats failed to consistently block LP-induced hyperphagia. Collectively, these data suggest that circulating BCAAs are transiently reduced by dietary protein restriction, but variations in dietary or brain BCAAs alone do not explain the hyperphagia induced by a low-protein diet.

Hepatic interoception in health and disease.

The liver is a large organ with crucial functions in metabolism and immune defense, as well as blood homeostasis and detoxification, and it is clearly in bidirectional communication with the brain and rest of the body via both neural and humoral pathways. A host of neural sensory mechanisms have been proposed, but in contrast to the gut-brain axis, details for both the exact site and molecular signaling steps of their peripheral transduction mechanisms are generally lacking. Similarly, knowledge about function-specific sensory and motor components of both vagal and spinal access pathways to the hepatic parenchyma is missing. Lack of progress largely owes to controversies regarding selectivity of vagal access pathways and extent of hepatocyte innervation. In contrast, there is considerable evidence for glucose sensors in the wall of the hepatic portal vein and their importance for glucose handling by the liver and the brain and the systemic response to hypoglycemia. As liver diseases are on the rise globally, and there are intriguing associations between liver diseases and mental illnesses, it will be important to further dissect and identify both neural and humoral pathways that mediate hepatocyte-specific signals to relevant brain areas. The question of whether and how sensations from the liver contribute to interoceptive self-awareness has not yet been explored.

History and future of leptin: Discovery, regulation and signaling.

The cloning of leptin 30 years ago in 1994 was an important milestone in obesity research. Prior to the discovery of leptin, obesity was stigmatized as a condition caused by lack of character and self-control. Mutations in either leptin or its receptor were the first single gene mutations found to cause severe obesity, and it is now recognized that obesity is caused mostly by a dysregulation of central neuronal circuits. Since the discovery of the leptin-deficient obese mouse (ob/ob) the cloning of leptin (ob aka lep) and leptin receptor (db aka lepr) genes, we have learned much about leptin and its action in the central nervous system. The first hope that leptin would cure obesity was quickly dampened because humans with obesity have increased leptin levels and develop leptin resistance. Nevertheless, leptin target sites in the brain represent an excellent blueprint to understand how neuronal circuits control energy homeostasis. Our expanding understanding of leptin function, interconnection of leptin signaling with other systems and impact on distinct physiological functions continues to guide and improve the development of safe and effective interventions to treat metabolic illnesses. This review highlights past concepts and current emerging concepts of the hormone leptin, leptin receptor signaling pathways and central targets to mediate distinct physiological functions.

FGF21 as a mediator of adaptive changes in food intake and macronutrient preference in response to protein restriction.

Free-feeding animals navigate complex nutritional landscapes in which food availability, cost, and nutritional value can vary markedly. Animals have thus developed neural mechanisms that enable the detection of nutrient restriction, and these mechanisms engage adaptive physiological and behavioral responses that limit or reverse this nutrient restriction. This review focuses specifically on dietary protein as an essential and independently defended nutrient. Adequate protein intake is required for life, and ample evidence exists to support an active defense of protein that involves behavioral changes in food intake, food preference, and food motivation, likely mediated by neural changes that increase the reward value of protein foods. Available evidence also suggests that the circulating hormone fibroblast growth factor 21 (FGF21) acts in the brain to coordinate these adaptive changes in food intake, making it a unique endocrine signal that drives changes in macronutrient preference in the context of protein restriction. This article is part of the Special Issue on "Food intake and feeding states".

FGF21 acts in the brain to drive macronutrient-specific changes in behavioral motivation and brain reward signaling.

Dietary protein restriction induces adaptive changes in food preference, increasing protein consumption over carbohydrates or fat. We investigated whether motivation and reward signaling underpin these preferences. In an operant task, protein-restricted male mice increased their responding for liquid protein rewards, but not carbohydrate, fat, or sweet rewards. The protein restriction-induced increase in operant responding for protein was absent in Fgf21-KO mice and mice with neuron-specific deletion of the FGF21 co-receptor beta-Klotho (KlbCam2ka) mice. Fiber photometry recording of VTA dopamine neurons revealed that oral delivery of maltodextrin triggered a larger activation of dopamine neurons as compared to casein in control-fed mice, while casein produced a larger response in protein-restricted mice. This restriction-induced shift in nutrient-specific VTA dopamine signaling was lost in Fgf21-KO mice. These data demonstrate that FGF21 acts in the brain to induce a protein-specific appetite by specifically enhancing the reward value of protein-containing foods and the motivation to consume them.

Leptin receptor neurons in the mouse hypothalamus are colocalized with the neuropeptide galanin and mediate anorexigenic leptin action.

Leptin acts centrally via leptin receptor (LepRb)-expressing neurons to regulate food intake, energy expenditure, and other physiological functions. LepRb neurons are found throughout the brain, and several distinct populations contribute to energy homeostasis control. However, the function of most LepRb populations remains unknown, and their contribution to regulate energy homeostasis has not been studied. Galanin has been hypothesized to interact with the leptin signaling system, but literature investigating colocalization of LepRb and galanin has been inconsistent, which is likely due to technical difficulties to visualize both. We used reporter mice with green fluorescent protein expression from the galanin locus to recapitulate the colocalization of galanin and leptin-induced p-STAT3 as a marker for LepRb expression. Here, we report the existence of two populations of galanin-expressing LepRb neurons (Gal-LepRb neurons): in the hypothalamus overspanning the perifornical area and adjacent dorsomedial and lateral hypothalamus [collectively named extended perifornical area (exPFA)] and in the brainstem (nucleus of the solitary tract). Surprisingly, despite the known orexigenic galanin action, leptin induces galanin mRNA expression and stimulates LepRb neurons in the exPFA, thus conflicting with the expected anorexigenic leptin action. However, we confirmed that intra-exPFA leptin injections were indeed sufficient to mediate anorexic responses. Interestingly, LepRb and galanin-expressing neurons are distinct from orexin or melanin-concentrating hormone (MCH)-expressing neurons, but exPFA galanin neurons colocalized with the anorexigenic neuropeptides neurotensin and cocaine- and amphetamine-regulated transcript (CART). Based on galanin's known inhibitory function, we speculate that in exPFA Gal-LepRb neurons galanin acts inhibitory rather than orexigenic.

Sensory spinal interoceptive pathways and energy balance regulation.

Interoception plays an important role in homeostatic regulation of energy intake and metabolism. Major interoceptive pathways include gut-to-brain and adipose tissue-to brain signaling via vagal sensory nerves and hormones, such as leptin. However, signaling via spinal sensory neurons is rapidly emerging as an additional important signaling pathway. Here we provide an in-depth review of the known anatomy and functions of spinal sensory pathways and discuss potential mechanisms relevant for energy balance homeostasis in health and disease. Because sensory innervation by dorsal root ganglia (DRG) neurons goes far beyond vagally innervated viscera and includes adipose tissue, skeletal muscle, and skin, it is in a position to provide much more complete metabolic information to the brain. Molecular and anatomical identification of function specific DRG neurons will be important steps in designing pharmacological and neuromodulation approaches to affect energy balance regulation in disease states such as obesity, diabetes, and cancer.

Regulation of body weight: Lessons learned from bariatric surgery.

BACKGROUND: Bariatric or weight loss surgery is currently the most effective treatment for obesity and metabolic disease. Unlike dieting and pharmacology, its beneficial effects are sustained over decades in most patients, and mortality is among the lowest for major surgery. Because there are not nearly enough surgeons to implement bariatric surgery on a global scale, intensive research efforts have begun to identify its mechanisms of action on a molecular level in order to replace surgery with targeted behavioral or pharmacological treatments. To date, however, there is no consensus as to the critical mechanisms involved. SCOPE OF REVIEW: The purpose of this non-systematic review is to evaluate the existing evidence for specific molecular and inter-organ signaling pathways that play major roles in bariatric surgery-induced weight loss and metabolic benefits, with a focus on Roux-en-Y gastric bypass (RYGB) and vertical sleeve gastrectomy (VSG), in both humans and rodents. MAJOR CONCLUSIONS: Gut-brain communication and its brain targets of food intake control and energy balance regulation are complex and redundant. Although the relatively young science of bariatric surgery has generated a number of hypotheses, no clear and unique mechanism has yet emerged. It seems increasingly likely that the broad physiological and behavioral effects produced by bariatric surgery do not involve a single mechanism, but rather multiple signaling pathways. Besides a need to improve and better validate surgeries in animals, advanced techniques, including inducible, tissue-specific knockout models, and the use of humanized physiological traits will be necessary. State-of-the-art genetically-guided neural identification techniques should be used to more selectively manipulate function-specific pathways.