Feeding circuit development and early-life influences on future feeding behaviour.

A wide range of maternal exposures - undernutrition, obesity, diabetes, stress and infection - are associated with an increased risk of metabolic disease in offspring. Developmental influences can cause persistent structural changes in hypothalamic circuits regulating food intake in the service of energy balance. The physiological relevance of these alterations has been called into question because maternal impacts on daily caloric intake do not persist to adulthood. Recent behavioural and epidemiological studies in humans provide evidence that the relative contribution of appetitive traits related to satiety, reward and the emotional aspects of food intake regulation changes across the lifespan. This Opinion article outlines a neurodevelopmental framework to explore the possibility that crosstalk between developing circuits regulating different modalities of food intake shapes future behavioural responses to environmental challenges.

MC4R-dependent suppression of appetite by bone-derived lipocalin 2.

Bone has recently emerged as a pleiotropic endocrine organ that secretes at least two hormones, FGF23 and osteocalcin, which regulate kidney function and glucose homeostasis, respectively. These findings have raised the question of whether other bone-derived hormones exist and what their potential functions are. Here we identify, through molecular and genetic analyses in mice, lipocalin 2 (LCN2) as an osteoblast-enriched, secreted protein. Loss- and gain-of-function experiments in mice demonstrate that osteoblast-derived LCN2 maintains glucose homeostasis by inducing insulin secretion and improves glucose tolerance and insulin sensitivity. In addition, osteoblast-derived LCN2 inhibits food intake. LCN2 crosses the blood-brain barrier, binds to the melanocortin 4 receptor (MC4R) in the paraventricular and ventromedial neurons of the hypothalamus and activates an MC4R-dependent anorexigenic (appetite-suppressing) pathway. These results identify LCN2 as a bone-derived hormone with metabolic regulatory effects, which suppresses appetite in a MC4R-dependent manner, and show that the control of appetite is an endocrine function of bone.

Corrigendum: MC4R-dependent suppression of appetite by bone-derived lipocalin 2.

This corrects the article DOI: 10.1038/nature21697.

Rethinking the Approach to Preclinical Models of Anorexia Nervosa.

PURPOSE OF REVIEW: The goal of this review is to describe how emerging technological developments in pre-clinical animal research can be harnessed to accelerate research in anorexia nervosa (AN). RECENT FINDINGS: The activity-based anorexia (ABA) paradigm, the best characterized animal model of AN, combines restricted feeding, excessive exercise, and weight loss. A growing body of evidence supports the idea that pathophysiological weight loss in this model is due to cognitive inflexibility, a clinical feature of AN. Targeted manipulations that recapitulate brain changes reported in AN - hyperdopaminergia or hyperactivity of cortical inputs to the nucleus accumbens - exacerbate weight loss in the ABA paradigm, providing the first evidence of causality. The power of preclinical research lies in the ability to assess the consequences of targeted manipulations of neuronal circuits that have been implicated in clinical research. Additional paradigms are needed to capture other features of AN that are not seen in ABA.

New roles for dopamine D2 and D3 receptors in pancreatic beta cell insulin secretion.

Although long-studied in the central nervous system, there is increasing evidence that dopamine (DA) has important roles in the periphery including in metabolic regulation. Insulin-secreting pancreatic ß-cells express the machinery for DA synthesis and catabolism, as well as all five DA receptors. In these cells, DA functions as a negative regulator of glucose-stimulated insulin secretion (GSIS), which is mediated by DA D2-like receptors including D2 (D2R) and D3 (D3R) receptors. However, the fundamental mechanisms of DA synthesis, storage, release, and signaling in pancreatic ß-cells and their functional relevance in vivo remain poorly understood. Here, we assessed the roles of the DA precursor L-DOPA in ß-cell DA synthesis and release in conjunction with the signaling mechanisms underlying DA's inhibition of GSIS. Our results show that the uptake of L-DOPA is essential for establishing intracellular DA stores in ß-cells. Glucose stimulation significantly enhances L-DOPA uptake, leading to increased DA release and GSIS reduction in an autocrine/paracrine manner. Furthermore, D2R and D3R act in combination to mediate dopaminergic inhibition of GSIS. Transgenic knockout mice in which ß-cell D2R or D3R expression is eliminated exhibit diminished DA secretion during glucose stimulation, suggesting a new mechanism where D2-like receptors modify DA release to modulate GSIS. Lastly, ß-cell-selective D2R knockout mice exhibit marked postprandial hyperinsulinemia in vivo. These results reveal that peripheral D2R and D3R receptors play important roles in metabolism through their inhibitory effects on GSIS. This opens the possibility that blockade of peripheral D2-like receptors by drugs including antipsychotic medications may significantly contribute to the metabolic disturbances observed clinically.

Developmental influences on circuits programming susceptibility to obesity.

Suboptimal maternal nutrition exerts lasting impacts on obesity risk in offspring, but the direction of the effect is determined by the timing of exposure. While maternal undernutrition in early pregnancy is associated with increased body mass index, in later pregnancy it can be protective. The importance of the timing of maternal undernutrition is also observed in rodents, however, many of the processes that occur in the last trimester of human gestation are delayed to the postnatal period. Neonatal leptin administration exerts lasting impacts on susceptibility to obesity in rodents. Although leptin can influence the formation of hypothalamic circuits involved in homeostatic control of feeding during the postnatal period, these effects are too late to account for its ability to reverse adverse metabolic programming due to early gestational exposure to maternal undernutrition. This review presents an alternative framework for understanding the effects of neonatal leptin through influences on developing thermoregulatory circuits.

Developmental switch of leptin signaling in arcuate nucleus neurons.

Leptin is well known for its role in the regulation of energy homeostasis in adults, a mechanism that at least partially results from the inhibition of the activity of NPY/AgRP/GABA neurons (NAG) in the arcuate nucleus of the hypothalamus (ARH). During early postnatal development in the rodent, leptin promotes axonal outgrowth from ARH neurons, and preautonomic NAG neurons are particularly responsive to leptin's trophic effects. To begin to understand how leptin could simultaneously promote axonal outgrowth from and inhibit the activity of NAG neurons, we characterized the electrochemical effects of leptin on NAG neurons in mice during early development. Here, we show that NAG neurons do indeed express a functional leptin receptor throughout the early postnatal period in the mouse; however, at postnatal days 13-15, leptin causes membrane depolarization in NAG neurons, rather than the expected hyperpolarization. Leptin action on NAG neurons transitions from stimulatory to inhibitory in the periweaning period, in parallel with the acquisition of functional ATP-sensitive potassium channels. These findings are consistent with the idea that leptin provides an orexigenic drive through the NAG system to help rapidly growing pups meet their energy requirements.

Functional organization of neuronal and humoral signals regulating feeding behavior.

Energy homeostasis--ensuring that energy availability matches energy requirements--is essential for survival. One way that energy balance is achieved is through coordinated action of neural and neuroendocrine feeding circuits, which promote energy intake when energy supply is limited. Feeding behavior engages multiple somatic and visceral tissues distributed throughout the body--contraction of skeletal and smooth muscles in the head and along the upper digestive tract required to consume and digest food, as well as stimulation of endocrine and exocrine secretions from a wide range of organs. Accordingly, neurons that contribute to feeding behaviors are localized to central, peripheral, and enteric nervous systems. To promote energy balance, feeding circuits must be able to identify and respond to energy requirements, as well as the amount of energy available from internal and external sources, and then direct appropriate coordinated responses throughout the body.

Rearing mice at 22°C programs increased capacity to respond to chronic exposure to cold but not high fat diet.

OBJECTIVE: Rodent models raised at environmental temperatures of 21-22 °C are increasingly switched to thermoneutral housing conditions in adulthood to better capture human physiology. We quantified the developmental effects of rearing mice at an ambient temperature of 22 °C vs. 30 °C on metabolic responses to cold and high fat diet (HFD) in adulthood. METHODS: Mice were reared from birth to 8 weeks of age at 22 °C or 30 °C, when they were acclimated to single housing at the same temperature for 2-3 weeks in indirect calorimetry cages. Energy expenditure attributable to basal metabolic rate, physical activity, thermic effect of food, and adaptive cold- or diet-induced thermogenesis was calculated. Responses to cooling were evaluated by decreasing the ambient temperature from 22 °C to 14 °C, while responses to HFD feeding were assessed at 30 °C. Influences of rearing temperature on thermogenic responses that emerge over hours, days and weeks were assessed by maintaining mice in the indirect calorimetry cages throughout the study. RESULTS: At an ambient temperature of 22 °C, total energy expenditure (TEE) was 12-16% higher in mice reared at 22 °C as compared to 30 °C. Rearing temperature had no effect on responses in the first hours or week of the 14 °C challenge. Differences emerged in the third week, when TEE increased an additional 10% in mice reared at 22 °C, but mice reared at 30 °C could not sustain this level of cold-induced thermogenesis. Rearing temperature only affected responses to HFD during the first week, due to differences in the timing but not the strength of metabolic adaptations. CONCLUSION: Rearing at 22 °C does not have a lasting effect on metabolic adaptations to HFD at thermoneutrality, but it programs an enhanced capacity to respond to chronic cold challenges in adulthood. These findings highlight the need to consider rearing temperature when using mice to model cold-induced thermogenesis.

Amygdala AVPR1A mediates susceptibility to chronic social isolation in females.

Females are more sensitive to social exclusion, which could contribute to their heightened susceptibility to anxiety disorders. Chronic social isolation stress (CSIS) for at least 7 weeks after puberty induces anxiety-related behavioral adaptations in female mice. Here, we show that Arginine vasopressin receptor 1a ( Avpr1a )-expressing neurons in the central nucleus of the amygdala (CeA) mediate these sex-specific effects, in part, via projections to the caudate putamen. Loss of function studies demonstrate that AVPR1A signaling in the CeA is required for effects of CSIS on anxiety-related behaviors in females but has no effect in males or group housed females. This sex-specificity is mediated by AVP produced by a subpopulation of neurons in the posterodorsal medial nucleus of the amygdala that project to the CeA. Estrogen receptor alpha signaling in these neurons also contributes to preferential sensitivity of females to CSIS. These data support new therapeutic applications for AVPR1A antagonists in women.

The long-lasting shadow of litter size in rodents: litter size is an underreported variable that strongly determines adult physiology.

BACKGROUND/PURPOSE: Litter size is a biological variable that strongly influences adult physiology in rodents. Despite evidence from previous decades and recent studies highlighting its major impact on metabolism, information about litter size is currently underreported in the scientific literature. Here, we urge that this important biological variable should be explicitly stated in research articles. RESULTS/CONCLUSION: Below, we briefly describe the scientific evidence supporting the impact of litter size on adult physiology and outline a series of recommendations and guidelines to be implemented by investigators, funding agencies, editors in scientific journals, and animal suppliers to fill this important gap.

A Framework for Developing Translationally Relevant Animal Models of Stress-Induced Changes in Eating Behavior.

Stress often affects eating behaviors, leading to increased eating in some individuals and decreased eating in others. Identifying physiological and psychological factors that determine the direction of eating responses to stress has been a major goal of epidemiological and clinical studies. However, challenges of standardizing the stress exposure in humans hinder efforts to uncover the underlying mechanisms. The issue of what determines the direction of stress-induced feeding responses has not been directly addressed in animal models, but assays that combine stress with a feeding-related task are commonly used as readouts of other behaviors, such as anxiety. Sex, estrous cyclicity, circadian cyclicity, caloric restriction, palatable diets, elevated body weight, and properties of the stressors similarly influence feeding behavior in humans and rodent models. Yet, most rodent studies do not use conditions that are most relevant for studying feeding behavior in humans. This review proposes a conceptual framework for incorporating these influences to develop reproducible and translationally relevant assays to study effects of stress on food intake. Such paradigms have the potential to uncover links between emotional eating and obesity as well as to the etiology of eating disorders.

Assessing the effects of stress on feeding behaviors in laboratory mice.

Stress often affects eating behaviors, increasing caloric intake in some individuals and decreasing it in others. The determinants of feeding responses to stress are unknown, in part because this issue is rarely studied in rodents. We focused our efforts on the novelty-suppressed feeding (NSF) assay, which uses latency to eat as readout of anxiety-like behavior, but rarely assesses feeding per se. We explored how key variables in experimental paradigms - estrous and diurnal cyclicity, age and duration of social isolation, prandial state, diet palatability, and elevated body weight - influence stress-induced anxiety-like behavior and food intake in male and female C57BL/6J mice. Latency to eat in the novel environment is increased in both sexes across most of the conditions tested, while effects on caloric intake are variable. In the common NSF assay (i.e., lean mice in the light cycle), sex-specific effects of the length of social isolation, and not estrous cyclicity, are the main source of variability. Under conditions that are more physiologically relevant for humans (i.e., overweight mice in the active phase), the novel stress now elicits robust hyperphagia in both sexes . This novel model of stress eating can be used to identify underlying neuroendocrine and neuronal substrates. Moreover, these studies can serve as a framework to integrate cross-disciplinary studies of anxiety and feeding related behaviors in rodents.

In times of heightened anxiety ­ say, during a global pandemic ­ many of us will reach for donuts or a particularly appetizing pizza for comfort. Others, however, will tend to shun food. What underlies these differences, and, in fact, the neural and hormonal pathways at play during stress eating (when people eat without being hungry due to emotional reasons), remain unclear. This is partly because scientists lack good animal models in which to study these behaviors. In particular, female rodents are usually excluded from studies under the assumption that their hormonal cycles will disrupt the results. Yet, women are overrepresented in studies on feeding habits. Modeling human behaviors using rodents is harder than it may appear. These animals are most active at night ­ yet most experiments are performed during the day. The same stressors also have different outcomes in males and females. François et al. therefore explored better ways to induce anxiety and evaluate feeding behavior in mice, hoping to reliably elicit stress eating. The starting point was a common type of experiments known as novelty-suppressed feeding. First, mice are kept alone in a cage for up to two weeks on a normal diet so that they are used to experimental conditions. Then they are deprived of food overnight, before being given free access to food in the morning in a new environment. This stressful experience normally causes mice to take longer to eat than in their home cage. In rodents, the delay is thought to reflect stress as it is reliably reversed by anti-anxiety compounds approved for human use. In the novelty-suppressed feeding assay, both male and female animals exhibit signs of anxiety, but how much females eat is variable. François et al. showed that this variability is not due to hormonal changes, but instead to how long female mice had been kept alone. Crucially, the test could be adapted so that mice would consistently exhibit behavior similar to human stress eating, whereby they eat more during the test without having fasted the night before. The changes included running the experiment at night, when the animals are normally most active, and using overweight mice (which captures the fact that, in humans, being overweight is associated with being prone to stress eating). Stress eating is an important clinical issue, hindering weigh loss in people with obesity. The new model developed by François et al. could be adopted by other laboratories, enabling better research into this behavior.

COVID-19 vaccines are effective in people with obesity: A position statement from The Obesity Society.

The position statement is issued by The Obesity Society in response to published literature, as well as inquiries made to the Society by patients, providers, Society members, policy makers, and others regarding the efficacy of vaccines in persons with obesity against SARS-CoV-2, the virus that causes COVID-19. The Obesity Society has critically evaluated data from published peer-reviewed literature and briefing documents from Emergency Use Authorization applications submitted by Pfizer-BioNTech, Moderna, and Johnson & Johnson. We conclude that these vaccines are highly efficacious, and their efficacy is not significantly different in people with and without obesity, based on scientific evidence available at the time of publication. The Obesity Society believes there is no definitive way to determine which of these three COVID-19 vaccines is "best" for any weight subpopulation (because of differences in the trial design and outcome measures in the phase 3 trials, elapsed time between doses, and regional differences in the presence of SARS-CoV-2 variants [e.g., South Africa B.1.351 in Johnson & Johnson trial]). All three trials have demonstrated high efficacy against COVID-19-associated hospitalization and death. Therefore, The Obesity Society encourages adults with obesity ≥18 years (≥16 years for Pfizer-BioNTech) to undergo vaccination with any one of the currently available vaccines authorized for emergency use by the US Food and Drug Administration as soon as they are able.

POMC neuronal heterogeneity in energy balance and beyond: an integrated view.

Hypothalamic AgRP and POMC neurons are conventionally viewed as the yin and yang of the body's energy status, since they act in an opposite manner to modulate appetite and systemic energy metabolism. However, although AgRP neurons' functions are comparatively well understood, a unifying theory of how POMC neuronal cells operate has remained elusive, probably due to their high level of heterogeneity, which suggests that their physiological roles might be more complex than initially thought. In this Perspective, we propose a conceptual framework that integrates POMC neuronal heterogeneity with appetite regulation, whole-body metabolic physiology and the development of obesity. We highlight emerging evidence indicating that POMC neurons respond to distinct combinations of interoceptive signals and food-related cues to fine-tune divergent metabolic pathways and behaviours necessary for survival. The new framework we propose reflects the high degree of developmental plasticity of this neuronal population and may enable progress towards understanding of both the aetiology and treatment of metabolic disorders.

Axon Guidance Molecules Implicated in Early-Onset Obesity.

Maternal nutritional status and the early growth rates of offspring influence the development of the melanocortin system and later susceptibility to metabolic dysregulation, but it is difficult to assess causality. A recent study by van der Klaauw et al. (Cell 2019;176:729-742) provides direct evidence that disrupting systems regulating neuronal circuit formation leads to early-onset obesity in zebrafish, mouse, and humans.

Proinsulin misfolding is an early event in the progression to type 2 diabetes.

Biosynthesis of insulin - critical to metabolic homeostasis - begins with folding of the proinsulin precursor, including formation of three evolutionarily conserved intramolecular disulfide bonds. Remarkably, normal pancreatic islets contain a subset of proinsulin molecules bearing at least one free cysteine thiol. In human (or rodent) islets with a perturbed endoplasmic reticulum folding environment, non-native proinsulin enters intermolecular disulfide-linked complexes. In genetically obese mice with otherwise wild-type islets, disulfide-linked complexes of proinsulin are more abundant, and leptin receptor-deficient mice, the further increase of such complexes tracks with the onset of islet insulin deficiency and diabetes. Proinsulin-Cys(B19) and Cys(A20) are necessary and sufficient for the formation of proinsulin disulfide-linked complexes; indeed, proinsulin Cys(B19)-Cys(B19) covalent homodimers resist reductive dissociation, highlighting a structural basis for aberrant proinsulin complex formation. We conclude that increased proinsulin misfolding via disulfide-linked complexes is an early event associated with prediabetes that worsens with ß-cell dysfunction in type two diabetes.

Perineuronal Net Formation during the Critical Period for Neuronal Maturation in the Hypothalamic Arcuate Nucleus.

In leptin-deficient ob/ob mice, obesity and diabetes are associated with abnormal development of neurocircuits in the hypothalamic arcuate nucleus (ARC)1, a critical brain area for energy and glucose homeostasis2,3. As this developmental defect can be remedied by systemic leptin administration, but only if given before postnatal day 28, a critical period (CP) for leptin-dependent development of ARC neurocircuits has been proposed4. In other brain areas, CP closure coincides with the appearance of perineuronal nets (PNNs), extracellular matrix specializations that restrict the plasticity of neurons that they enmesh5. Here we report that in humans as well as rodents, subsets of neurons in the mediobasal aspect of the ARC are enmeshed by PNN-like structures. In mice, these neurons are densely-packed into a continuous ring that encircles the junction of the ARC and median eminence, which facilitates exposure of ARC neurons to the circulation. Most of the enmeshed neurons are both GABAergic and leptin receptor-positive, including a majority of Agrp neurons. Postnatal formation of the PNN-like structures coincides precisely with closure of the CP for Agrp neuron maturation and is dependent on input from circulating leptin, as postnatal ob/ob mice have reduced ARC PNN-like material that is restored by leptin administration during the CP. We conclude that neurons crucial to metabolic homeostasis are enmeshed by PNN-like structures and organized into a densely packed cluster situated circumferentially at the ARC-ME junction, where metabolically-relevant humoral signals are sensed.

Functional identity of hypothalamic melanocortin neurons depends on Tbx3.

Heterogeneous populations of hypothalamic neurons orchestrate energy balance via the release of specific signatures of neuropeptides. However, how specific intracellular machinery controls peptidergic identities and function of individual hypothalamic neurons remains largely unknown. The transcription factor T-box 3 (Tbx3) is expressed in hypothalamic neurons sensing and governing energy status, whereas human TBX3 haploinsufficiency has been linked with obesity. Here, we demonstrate that loss of Tbx3 function in hypothalamic neurons causes weight gain and other metabolic disturbances by disrupting both the peptidergic identity and plasticity of Pomc/Cart and Agrp/Npy neurons. These alterations are observed after loss of Tbx3 in both immature hypothalamic neurons and terminally differentiated mouse neurons. We further establish the importance of Tbx3 for body weight regulation in Drosophila melanogaster and show that TBX3 is implicated in the differentiation of human embryonic stem cells into hypothalamic Pomc neurons. Our data indicate that Tbx3 directs the terminal specification of neurons as functional components of the melanocortin system and is required for maintaining their peptidergic identity. In summary, we report the discovery of a key mechanistic process underlying the functional heterogeneity of hypothalamic neurons governing body weight and systemic metabolism.