RESUMEN

SH2B1 mutations are associated with obesity, type 2 diabetes, and metabolic dysfunction-associated steatotic liver disease (MASLD) in humans. Global deletion of Sh2b1 results in severe obesity, type 2 diabetes, and MASLD in mice. Neuron-specific restoration of SH2B1 rescues the obesity phenotype of Sh2b1-null mice, indicating that the brain is a main SH2B1 target. However, SH2B1 neurocircuits remain elusive. SH2B1-expressing neurons in the paraventricular hypothalamus (PVHSH2B1) and a PVHSH2B1→dorsal raphe nucleus (DRN) neurocircuit are identified here. PVHSH2B1 axons monosynaptically innervate DRN neurons. Optogenetic stimulation of PVHSH2B1 axonal fibers in the DRN suppresses food intake. Chronic inhibition of PVHSH2B1 neurons causes obesity. In male and female mice, either embryonic-onset or adult-onset deletion of Sh2b1 in PVH neurons causes energy imbalance, obesity, insulin resistance, glucose intolerance, and MASLD. Ablation of Sh2b1 in the DRN-projecting PVHSH2B1 subpopulation also causes energy imbalance, obesity, and metabolic disorders. Conversely, SH2B1 overexpression in either total or DRN-projecting PVHSH2B1 neurons protects against diet-induced obesity. SH2B1 binds to TrkB and enhances brain-derived neurotrophic factor (BDNF) signaling. Ablation of Sh2b1 in PVHSH2B1 neurons induces BDNF resistance in the PVH, contributing to obesity. In conclusion, these results unveil a previously unrecognized PVHSH2B1→DRN neurocircuit through which SH2B1 defends against obesity by enhancing BDNF/TrkB signaling.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales , Obesidad , Núcleo Hipotalámico Paraventricular , Animales , Obesidad/metabolismo , Obesidad/genética , Ratones , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Masculino , Femenino , Núcleo Hipotalámico Paraventricular/metabolismo , Modelos Animales de Enfermedad , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/genética , Metabolismo Energético/genética , Metabolismo Energético/fisiología , Núcleo Dorsal del Rafe/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Factor Neurotrófico Derivado del Encéfalo/genética , Neuronas/metabolismo

RESUMEN

Metabolic dysfunction-associated steatohepatitis (MASH), formerly known as nonalcoholic steatohepatitis (NASH), is an advanced stage of metabolic fatty liver disease. The pathogenic mechanisms of MASH center on hepatocyte injury and the ensuing immune response within the liver microenvironment. Recent work has implicated TREM2+ macrophages in various disease conditions, and substantial induction of TREM2+ NASH-associated macrophages (NAMs) serves as a hallmark of metabolic liver disease. Despite this, the mechanisms through which NAMs contribute to MASH pathogenesis remain poorly understood. Here, we identify membrane-spanning 4-domains a7 (MS4A7) as a NAM-specific pathogenic factor that exacerbates MASH progression in mice. Hepatic MS4A7 expression was strongly induced in mouse and human MASH and associated with the severity of liver injury. Whole-body and myeloid-specific ablation of Ms4a7 alleviated diet-induced MASH pathologies in male mice. We demonstrate that exposure to lipid droplets (LDs), released upon injury of steatotic hepatocytes, triggered NAM induction and exacerbated MASH-associated liver injury in an MS4A7-dependent manner. Mechanistically, MS4A7 drove NLRP3 inflammasome activation via direct physical interaction and shaped disease-associated cell states within the liver microenvironment. This work reveals the LD-MS4A7-NLRP3 inflammasome axis as a pathogenic driver of MASH progression and provides insights into the role of TREM2+ macrophages in disease pathogenesis.

Asunto(s)

Inflamasomas , Enfermedad del Hígado Graso no Alcohólico , Animales , Humanos , Masculino , Ratones , Inflamasomas/metabolismo , Hígado/metabolismo , Macrófagos/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Receptores Inmunológicos/metabolismo

RESUMEN

The adipose-derived hormone leptin acts via its receptor (LepRb) in the brain to control energy balance. A potentially unidentified population of GABAergic hypothalamic LepRb neurons plays key roles in the restraint of food intake and body weight by leptin. To identify markers for candidate populations of LepRb neurons in an unbiased manner, we performed single-nucleus RNA-Seq of enriched mouse hypothalamic LepRb cells, identifying several previously unrecognized populations of hypothalamic LepRb neurons. Many of these populations displayed strong conservation across species, including GABAergic Glp1r-expressing LepRb (LepRbGlp1r) neurons, which expressed more Lepr than other LepRb cell populations. Ablating Lepr from LepRbGlp1r cells provoked hyperphagic obesity without impairing energy expenditure. Similarly, improvements in energy balance caused by Lepr reactivation in GABA neurons of otherwise Lepr-null mice required Lepr expression in GABAergic Glp1r-expressing neurons. Furthermore, restoration of Glp1r expression in LepRbGlp1r neurons in otherwise Glp1r-null mice enabled food intake suppression by the GLP1R agonist, liraglutide. Thus, the conserved GABAergic LepRbGlp1r neuron population plays crucial roles in the suppression of food intake by leptin and GLP1R agonists.

Asunto(s)

Leptina , Obesidad , Ratones , Animales , Leptina/genética , Leptina/metabolismo , Obesidad/genética , Obesidad/prevención & control , Obesidad/metabolismo , Hipotálamo/metabolismo , Ratones Noqueados , Neuronas GABAérgicas/metabolismo , Receptores de Leptina/genética , Receptores de Leptina/metabolismo , Ingestión de Alimentos/genética

RESUMEN

Early life stress (ELS) and major depressive disorder (MDD) share neural network abnormalities. However, it is unclear how ELS and MDD may separately and/or jointly relate to brain networks, and whether neural differences exist between depressed individuals with vs without ELS. Moreover, prior work evaluated static versus dynamic network properties, a critical gap considering brain networks show changes in coordinated activity over time. Seventy-one unmedicated females with and without childhood sexual abuse (CSA) histories and/or MDD completed a resting state scan and a stress task in which cortisol and affective ratings were collected. Recurring functional network co-activation patterns (CAPs) were examined and time in CAP (number of times each CAP is expressed) and transition frequencies (transitioning between different CAPs) were computed. The effects of MDD and CSA on CAP metrics were examined and CAP metrics were correlated with depression and stress-related variables. Results showed that MDD, but not CSA, related to CAP metrics. Specifically, individuals with MDD (N = 35) relative to HCs (N = 36), spent more time in a posterior default mode (DMN)-frontoparietal network (FPN) CAP and transitioned more frequently between posterior DMN-FPN and prototypical DMN CAPs. Across groups, more time spent in a posterior DMN-FPN CAP and greater DMN-FPN and prototypical DMN CAP transition frequencies were linked to higher rumination. Imbalances between the DMN and the FPN appear central to MDD and might contribute to MDD-related cognitive dysfunction, including rumination. Unexpectedly, CSA did not modulate such dysfunctions, a finding that needs to be replicated by future studies with larger sample sizes.

Asunto(s)

Trastorno Depresivo Mayor , Delitos Sexuales , Femenino , Niño , Humanos , Vías Nerviosas , Imagen por Resonancia Magnética/métodos , Encéfalo/diagnóstico por imagen , Mapeo Encefálico/métodos

RESUMEN

Variants in the gene encoding ankyrin repeat and SOCS box-containing 4 (ASB4) are linked to human obesity. Here, we characterized the pathways underlying the metabolic functions of ASB4. Hypothalamic Asb4 expression was suppressed by fasting in wild-type mice but not in mice deficient in AgRP, which encodes Agouti-related protein (AgRP), an appetite-stimulating hormone, suggesting that ASB4 is a negative target of AgRP. Many ASB4 neurons in the brain were adjacent to AgRP terminals, and feeding induced by AgRP neuronal activation was disrupted in Asb4-deficient mice. Acute knockdown of Asb4 in the brain caused marked hyperphagia due to increased meal size, and Asb4 deficiency led to increased meal size and food intake at the onset of refeeding, when very large meals were consumed. Asb4-deficient mice were resistant to the meal-terminating effects of exogenously administered calcitonin and showed decreased neuronal expression of Calcr, which encodes the calcitonin receptor. Pro-opiomelanocortin (POMC) neurons in the arcuate nucleus in mice are involved in glucose homeostasis, and Asb4 deficiency specifically in POMC neurons resulted in glucose intolerance that was independent of obesity. Furthermore, individuals with type 2 diabetes showed reduced ASB4 abundance in the infundibular nuclei, the human equivalent of the arcuate nucleus. Together, our results indicate that ASB4 acts in the brain to improve glucose homeostasis and to induce satiety after substantial meals, particularly those after food deprivation.

Asunto(s)

Diabetes Mellitus Tipo 2 , Neuropéptidos , Proteína Relacionada con Agouti/genética , Proteína Relacionada con Agouti/metabolismo , Proteína Relacionada con Agouti/farmacología , Animales , Calcitonina/metabolismo , Calcitonina/farmacología , Diabetes Mellitus Tipo 2/metabolismo , Glucosa/metabolismo , Homeostasis , Hipotálamo/metabolismo , Ratones , Neuronas/metabolismo , Neuropéptidos/metabolismo , Obesidad/genética , Obesidad/metabolismo , Proopiomelanocortina/genética , Proopiomelanocortina/metabolismo , Proopiomelanocortina/farmacología

RESUMEN

Increasing evidence indicates that the melanocortin and mesolimbic dopamine (DA) systems interact to regulate feeding and body weight. Because melanocortin-3 receptors (MC3R) are highly expressed in the ventral tegmental area (VTA), we tested whether VTA neurons expressing these receptors (VTA MC3R neurons) control feeding and body weight in vivo. We also tested whether there were sex differences in the ability of VTA MC3R neurons to control feeding, as MC3R -/- mice show sex-dependent alterations in reward feeding and DA levels, and there are clear sex differences in multiple DA-dependent behaviors and disorders. Designer receptors exclusively activated by designer drugs (DREADD) were used to acutely activate and inhibit VTA MC3R neurons and changes in food intake and body weight were measured. Acutely altering the activity of VTA MC3R neurons decreased feeding in an activity- and sex-dependent manner, with acute activation decreasing feeding, but only in females, and acute inhibition decreasing feeding, but only in males. These differences did not appear to be due to sex differences in the number of VTA MC3R neurons, the ability of hM3Dq to activate VTA MC3R neurons, or the proportion of VTA MC3R neurons expressing tyrosine hydroxylase (TH). These studies demonstrate an important role for VTA MC3R neurons in the control of feeding and reveal important sex differences in behavior, whereby opposing changes in neuronal activity in male and female mice cause similar changes in behavior.

Asunto(s)

Actividad Motora/fisiología , Neuronas/fisiología , Receptor de Melanocortina Tipo 3/fisiología , Área Tegmental Ventral/fisiología , Animales , Peso Corporal , Drogas de Diseño/farmacología , Dopamina/metabolismo , Conducta Alimentaria , Femenino , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Receptor de Melanocortina Tipo 3/genética , Recompensa , Caracteres Sexuales , Área Tegmental Ventral/citología

RESUMEN

Calcitonin receptor (Calcr)-expressing neurons of the nucleus tractus solitarius (NTS; CalcrNTS cells) contribute to the long-term control of food intake and body weight. Here, we show that Prlh-expressing NTS (PrlhNTS) neurons represent a subset of CalcrNTS cells and that Prlh expression in these cells restrains body weight gain in the face of high fat diet challenge in mice. To understand the relationship of PrlhNTS cells to hypothalamic feeding circuits, we determined the ability of PrlhNTS-mediated signals to overcome enforced activation of AgRP neurons. We found that PrlhNTS neuron activation and Prlh overexpression in PrlhNTS cells abrogates AgRP neuron-driven hyperphagia and ameliorates the obesity of mice deficient in melanocortin signaling or leptin. Thus, enhancing Prlh-mediated neurotransmission from the NTS dampens hypothalamically-driven hyperphagia and obesity, demonstrating that NTS-mediated signals can override the effects of orexigenic hypothalamic signals on long-term energy balance.

Asunto(s)

Obesidad/metabolismo , Hormona Liberadora de Prolactina/metabolismo , Núcleo Solitario/metabolismo , Animales , Apetito , Dieta , Ingestión de Alimentos , Metabolismo Energético , Femenino , Humanos , Hipotálamo/metabolismo , Leptina/metabolismo , Masculino , Melanocortinas/metabolismo , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Obesidad/genética , Obesidad/fisiopatología , Obesidad/psicología , Hormona Liberadora de Prolactina/genética , Receptores de Calcitonina/genética , Receptores de Calcitonina/metabolismo

RESUMEN

The dorsal vagal complex (DVC) in the hindbrain, composed of the area postrema, nucleus of the solitary tract, and dorsal motor nucleus of the vagus, plays a critical role in modulating satiety. The incretins glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) act directly in the brain to modulate feeding, and receptors for both are expressed in the DVC. Given the impressive clinical responses to pharmacologic manipulation of incretin signaling, understanding the central mechanisms by which incretins alter metabolism and energy balance is of critical importance. Here, we review recent single-cell approaches used to detect molecular signatures of GLP-1 and GIP receptor-expressing cells in the DVC. In addition, we discuss how current advancements in single-cell transcriptomics, epigenetics, spatial transcriptomics, and circuit mapping techniques have the potential to further characterize incretin receptor circuits in the hindbrain.

Asunto(s)

Receptor del Péptido 1 Similar al Glucagón/metabolismo , Receptores de la Hormona Gastrointestinal/metabolismo , Rombencéfalo/metabolismo , Animales , Conducta Alimentaria/fisiología , Polipéptido Inhibidor Gástrico/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Humanos , Análisis de la Célula Individual

RESUMEN

Here, we present an in-depth protocol for extracting ribosome-bound mRNAs in low-abundance cells of hypothalamic nuclei. mRNAs are extracted from the micropunched tissue using refined translating ribosome affinity purification. Isolated RNAs can be used for sequencing or transcript quantification. This protocol enables the identification of actively translated mRNAs in varying physiological states and can be modified for use in any neuronal subpopulation labeled with a ribo-tag. We use leptin receptor-expressing neurons as an example to illustrate the protocol. For complete details on the use and execution of this protocol, please refer to Han et al. (2020).

Asunto(s)

Cromatografía de Afinidad/métodos , Hipotálamo/metabolismo , ARN Mensajero/aislamiento & purificación , Ribosomas/metabolismo , Animales , Proteínas Fluorescentes Verdes/genética , Ratones , Neuronas/metabolismo , ARN Mensajero/metabolismo

RESUMEN

The ventromedial hypothalamic nucleus (VMH) controls diverse behaviors and physiologic functions, suggesting the existence of multiple VMH neural subtypes with distinct functions. Combing translating ribosome affinity purification with RNA-sequencing (TRAP-seq) data with single-nucleus RNA-sequencing (snRNA-seq) data, we identified 24 mouse VMH neuron clusters. Further analysis, including snRNA-seq data from macaque tissue, defined a more tractable VMH parceling scheme consisting of six major genetically and anatomically differentiated VMH neuron classes with good cross-species conservation. In addition to two major ventrolateral classes, we identified three distinct classes of dorsomedial VMH neurons. Consistent with previously suggested unique roles for leptin receptor (Lepr)-expressing VMH neurons, Lepr expression marked a single dorsomedial class. We also identified a class of glutamatergic VMH neurons that resides in the tuberal region, anterolateral to the neuroanatomical core of the VMH. This atlas of conserved VMH neuron populations provides an unbiased starting point for the analysis of VMH circuitry and function.

Asunto(s)

Familia de Multigenes , Neuronas/fisiología , Transcriptoma , Núcleo Hipotalámico Ventromedial/fisiología , Animales , Análisis por Conglomerados , Bases de Datos Genéticas , Perfilación de la Expresión Génica , Genotipo , Ácido Glutámico/metabolismo , Macaca mulatta , Ratones Transgénicos , Neuronas/metabolismo , Fenotipo , RNA-Seq , Receptores de Leptina/genética , Receptores de Leptina/metabolismo , Especificidad de la Especie , Factor Esteroidogénico 1/genética , Factor Esteroidogénico 1/metabolismo , Núcleo Hipotalámico Ventromedial/citología , Núcleo Hipotalámico Ventromedial/metabolismo

RESUMEN

The paraventricular nucleus of the hypothalamus (PVH) is a heterogeneous collection of neurons that play important roles in modulating feeding and energy expenditure. Abnormal development or ablation of the PVH results in hyperphagic obesity and defects in energy expenditure whereas selective activation of defined PVH neuronal populations can suppress feeding and may promote energy expenditure. Here, we characterize the contribution of calcitonin receptor-expressing PVH neurons (CalcRPVH) to energy balance control. We used Cre-dependent viral tools delivered stereotaxically to the PVH of CalcR2Acre mice to activate, silence, and trace CalcRPVH neurons and determine their contribution to body weight regulation. Immunohistochemistry of fluorescently-labeled CalcRPVH neurons demonstrates that CalcRPVH neurons are largely distinct from several PVH neuronal populations involved in energy homeostasis; these neurons project to regions of the hindbrain that are implicated in energy balance control, including the nucleus of the solitary tract and the parabrachial nucleus. Acute activation of CalcRPVH neurons suppresses feeding without appreciably augmenting energy expenditure, whereas their silencing leads to obesity that may be due in part due to loss of PVH melanocortin-4 receptor signaling. These data show that CalcRPVH neurons are an essential component of energy balance neurocircuitry and their function is important for body weight maintenance. A thorough understanding of the mechanisms by which CalcRPVH neurons modulate energy balance might identify novel therapeutic targets for the treatment and prevention of obesity.

Asunto(s)

Metabolismo Energético/fisiología , Núcleo Hipotalámico Paraventricular/fisiología , Receptores de Calcitonina/fisiología , Animales , Ingestión de Alimentos/fisiología , Metabolismo Energético/genética , Conducta Alimentaria/fisiología , Homeostasis/fisiología , Hipotálamo/metabolismo , Hipotálamo/fisiología , Masculino , Ratones , Ratones Transgénicos , Neuronas/metabolismo , Neuronas/fisiología , Núcleo Hipotalámico Paraventricular/metabolismo , Receptor de Melanocortina Tipo 4/genética , Receptor de Melanocortina Tipo 4/metabolismo , Receptor de Melanocortina Tipo 4/fisiología , Receptores de Calcitonina/genética , Receptores de Calcitonina/metabolismo

RESUMEN

The ventromedial hypothalamus (VMH) is a critical neural node that senses blood glucose and promotes glucose utilization or mobilization during hypoglycemia. The VMH neurons that control these distinct physiologic processes are largely unknown. Here, we show that melanocortin 3 receptor (Mc3R)-expressing VMH neurons (VMHMC3R) sense glucose changes both directly and indirectly via altered excitatory input. We identify presynaptic nodes that potentially regulate VMHMC3R neuronal activity, including inputs from proopiomelanocortin (POMC)-producing neurons in the arcuate nucleus. We find that VMHMC3R neuron activation blunts, and their silencing enhances glucose excursion following a glucose load. Overall, these findings demonstrate that VMHMC3R neurons are a glucose-responsive hypothalamic subpopulation that promotes glucose disposal upon activation; this highlights a potential site for targeting dysregulated glycemia.

Asunto(s)

Glucosa/metabolismo , Hiperglucemia/metabolismo , Hipotálamo/metabolismo , Neuronas/metabolismo , Receptor de Melanocortina Tipo 3/metabolismo , Animales , Hipotálamo/citología , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/fisiología , Proopiomelanocortina/metabolismo , Receptor de Melanocortina Tipo 3/genética , Potenciales Sinápticos

RESUMEN

While Cre-dependent viral systems permit the manipulation of many neuron types, some cell populations cannot be targeted by a single DNA recombinase. Although the combined use of Flp and Cre recombinases can overcome this limitation, insufficient recombinase activity can reduce the efficacy of existing Cre+Flp-dependent viral systems. We developed a sensitive dual recombinase-activated viral approach: tTA-driven Recombinase-Guided Intersectional Targeting (tTARGIT) adeno-associated viruses (AAVs). tTARGIT AAVs utilize a Flp-dependent tetracycline transactivator (tTA) 'Driver' AAV and a tetracycline response element-driven, Cre-dependent 'Payload' AAV to express the transgene of interest. We employed this system in Slc17a6FlpO;LeprCre mice to manipulate LepRb neurons of the ventromedial hypothalamus (VMH; LepRbVMH neurons) while omitting neighboring LepRb populations. We defined the circuitry of LepRbVMH neurons and roles for these cells in the control of food intake and energy expenditure. Thus, the tTARGIT system mediates robust recombinase-sensitive transgene expression, permitting the precise manipulation of previously intractable neural populations.

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The brain contains hundreds of types of neurons, which differ in size, shape and behavior. But neuroscientists often wish to study individual neuronal types in isolation. They are able to do this with the aid of a toolkit made up of two parts: viral vectors and genetically modified mice. Viral vectors are viruses that have been modified so that they are no longer harmful and can instead be used to introduce genetic material into cells on demand. To create a viral vector, the virus' own genetic material is replaced with a 'cargo' gene, such as the gene for a fluorescent protein. The virus is then introduced into a new host such as a mouse. Importantly, the virus only produces the protein encoded by its 'cargo' gene if it is inside a cell that also contains one of two specific enzymes. These enzymes are called Cre and Flp. This is where the second part of the toolkit comes in. Mice can be genetically engineered to produce either Cre or Flp exclusively in specific cell types. By introducing a viral vector into mice that produce either Cre or Flp only in one particular type of neuron, researchers can limit the activity of the cargo gene to that neuronal type. But sometimes even this approach is not selective enough. Researchers may wish to limit the activity of the cargo gene to a subpopulation of cells that produce Cre or Flp. Or they may wish to target only Cre- or Flp-producing cells in a small area of the brain, while leaving cells in neighboring areas unaffected. Sabatini et al. have now overcome this limitation by developing and testing a new set of viral vectors that are active only in neurons that produce both Cre and Flp. The vectors are called tTARGIT AAVs and allow researchers to target cells more precisely than was possible with the previous version of the toolkit. Sabatini et al. show tTARGIT AAVs in action by using them to identify a group of neurons that control how much energy mice use and how much food they eat. As well as applying the vectors to their own research on obesity, Sabatini et al. have also made them freely available for other researchers to use in their own projects.

Asunto(s)

Expresión Génica , Neuronas/fisiología , Transgenes , Animales , Dependovirus/genética , Femenino , Masculino , Ratones , Ratones Transgénicos

RESUMEN

The TGFß cytokine family member, GDF-15, reduces food intake and body weight and represents a potential treatment for obesity. Because the brainstem-restricted expression pattern of its receptor, GDNF Family Receptor α-like (GFRAL), presents an exciting opportunity to understand mechanisms of action for area postrema neurons in food intake; we generated GfralCre and conditional GfralCreERT mice to visualize and manipulate GFRAL neurons. We found infection or pathophysiologic states (rather than meal ingestion) stimulate GFRAL neurons. TRAP-Seq analysis of GFRAL neurons revealed their expression of a wide range of neurotransmitters and neuropeptides. Artificially activating GfralCre -expressing neurons inhibited feeding, decreased gastric emptying, and promoted a conditioned taste aversion (CTA). GFRAL neurons most strongly innervate the parabrachial nucleus (PBN), where they target CGRP-expressing (CGRPPBN) neurons. Silencing CGRPPBN neurons abrogated the aversive and anorexic effects of GDF-15. These findings suggest that GFRAL neurons link non-meal-associated pathophysiologic signals to suppress nutrient uptake and absorption.

Asunto(s)

Reacción de Prevención/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Conducta Alimentaria/efectos de los fármacos , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Factor 15 de Diferenciación de Crecimiento/farmacología , Neuronas/fisiología , Núcleos Parabraquiales/fisiología , Animales , Peso Corporal , Femenino , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/genética , Masculino , Ratones , Neuronas/efectos de los fármacos , Núcleos Parabraquiales/efectos de los fármacos , Ratas , Ratas Long-Evans

RESUMEN

The mesolimbic dopamine (DA) system is involved in the regulation of multiple behaviors, including feeding, and evidence demonstrates that the melanocortin system can act on the mesolimbic DA system to control feeding and other behaviors. The melanocortin-3 receptor (MC3R) is an important component of the melanocortin system, but its overall role is poorly understood. Because MC3Rs are highly expressed in the ventral tegmental area (VTA) and are likely to be the key interaction point between the melanocortin and mesolimbic DA systems, we set out to identify both the efferent projection patterns of VTA MC3R neurons and the location of the neurons providing afferent input to them. VTA MC3R neurons were broadly connected to neurons across the brain but were strongly connected to a discrete set of brain regions involved in the regulation of feeding, reward, and aversion. Surprisingly, experiments using monosynaptic rabies virus showed that proopiomelanocortin (POMC) and agouti-related protein (AgRP) neurons in the arcuate nucleus made few direct synapses onto VTA MC3R neurons or any of the other major neuronal subtypes in the VTA, despite being extensively labeled by general retrograde tracers injected into the VTA. These results greatly contribute to our understanding of the anatomical interactions between the melanocortin and mesolimbic systems and provide a foundation for future studies of VTA MC3R neurons and the circuits containing them in the control of feeding and other behaviors.

Asunto(s)

Neuronas Dopaminérgicas/metabolismo , Neuronas Aferentes/metabolismo , Neuronas Eferentes/metabolismo , Receptor de Melanocortina Tipo 3/biosíntesis , Área Tegmental Ventral/metabolismo , Animales , Neuronas Dopaminérgicas/química , Femenino , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas Aferentes/química , Neuronas Eferentes/química , Receptor de Melanocortina Tipo 3/análisis , Receptor de Melanocortina Tipo 3/genética , Área Tegmental Ventral/química

RESUMEN

Enhancer redundancy has been postulated to provide a buffer for gene expression against genetic and environmental perturbations. While work in Drosophila has identified functionally overlapping enhancers, work in mammalian models has been limited. Recently, we have identified two partially redundant enhancers, nPE1 and nPE2, that drive proopiomelanocortin gene expression in the hypothalamus. Here we demonstrate that deletion of nPE1 produces mild obesity while knockout of nPE2 has no discernible metabolic phenotypes. Additionally, we show that acute leptin administration has significant effects on nPE1 knockout mice, with food intake and body weight change significantly impacted by peripheral leptin treatment. nPE1 knockout mice became less responsive to leptin treatment over time as percent body weight change increased over 2 week exposure to peripheral leptin. Both Pomc and Agrp mRNA were not differentially affected by chronic leptin treatment however we did see a decrease in Pomc and Agrp mRNA in both nPE1 and nPE2 knockout calorie restricted mice as compared to calorie restricted PBS-treated WT mice. Collectively, these data suggest dynamic regulation of Pomc by nPE1 such that mice with nPE1 knockout become less responsive to the anorectic effects of leptin treatment over time. Our results also support our earlier findings in which nPE2 may only be critical in adult mice that lack nPE1, indicating that these neural enhancers work synergistically to influence metabolism.

Asunto(s)

Depresores del Apetito/farmacología , Peso Corporal/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Elementos de Facilitación Genéticos , Hipotálamo/efectos de los fármacos , Leptina/farmacología , Neuronas/efectos de los fármacos , Proopiomelanocortina/genética , Animales , Hipotálamo/metabolismo , Masculino , Ratones , Ratones Noqueados , Neuronas/metabolismo

RESUMEN

Epidemiological and genome-wide association studies (GWAS) have shown high correlation between childhood obesity and advance in puberty. Early age at menarche is associated with a series of morbidities, including breast cancer, cardiovascular diseases, type 2 diabetes, and obesity. The adipocyte hormone leptin signals the amount of fat stores to the neuroendocrine reproductive axis via direct actions in the brain. Using mouse genetics, we and others have identified the hypothalamic ventral premammillary nucleus (PMv) and the agouti-related protein (AgRP) neurons in the arcuate nucleus (Arc) as primary targets of leptin action in pubertal maturation. However, the molecular mechanisms underlying leptin's effects remain unknown. Here we assessed changes in the PMv and Arc transcriptional program during leptin-stimulated and typical pubertal development using overlapping analysis of bulk RNA sequecing, TRAP sequencing, and the published database. Our findings demonstrate that dynamic somatodendritic remodeling and extracellular space organization underlie leptin-induced and typical pubertal maturation in female mice.

RESUMEN

Polycystic ovary syndrome (PCOS) is the most common form of infertility in women. The causes of PCOS are not yet understood and both genetics and early-life exposure have been considered as candidates. With regard to the latter, circulating androgens are elevated in mid-late gestation in women with PCOS, potentially exposing offspring to elevated androgens in utero; daughters of women with PCOS are at increased risk for developing this disorder. Consistent with these clinical observations, prenatal androgenization (PNA) of several species recapitulates many phenotypes observed in PCOS. There is increasing evidence that symptoms associated with PCOS, including elevated luteinizing hormone (LH) (and presumably gonadotropin-releasing hormone [GnRH]) pulse frequency emerge during the pubertal transition. We utilized translating ribosome affinity purification coupled with ribonucleic acid (RNA) sequencing to examine GnRH neuron messenger RNAs from prepubertal (3 weeks) and adult female control and PNA mice. Prominent in GnRH neurons were transcripts associated with protein synthesis and cellular energetics, in particular oxidative phosphorylation. The GnRH neuron transcript profile was affected more by the transition from prepuberty to adulthood than by PNA treatment; however, PNA did change the developmental trajectory of GnRH neurons. This included families of transcripts related to both protein synthesis and oxidative phosphorylation, which were more prevalent in adults than in prepubertal mice but were blunted in PNA adults. These findings suggest that prenatal androgen exposure can program alterations in the translatome of GnRH neurons, providing a mechanism independent of changes in the genetic code for altered expression.

Asunto(s)

Neurogénesis/efectos de los fármacos , Neuronas/efectos de los fármacos , Efectos Tardíos de la Exposición Prenatal , Área Preóptica/efectos de los fármacos , Virilismo , Andrógenos/efectos adversos , Animales , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Hormona Liberadora de Gonadotropina/metabolismo , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Neurogénesis/genética , Neuronas/metabolismo , Neuronas/fisiología , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/genética , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Área Preóptica/citología , Área Preóptica/crecimiento & desarrollo , Área Preóptica/metabolismo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores Sexuales , Virilismo/inducido químicamente , Virilismo/genética , Virilismo/fisiopatología

RESUMEN

The hypothalamic suprachiasmatic (SCN) clock contains several neurochemically defined cell groups that contribute to the genesis of circadian rhythms. Using cell-specific and genetically targeted approaches we have confirmed an indispensable role for vasoactive intestinal polypeptide-expressing SCN (SCNVIP) neurons, including their molecular clock, in generating the mammalian locomotor activity (LMA) circadian rhythm. Optogenetic-assisted circuit mapping revealed functional, di-synaptic connectivity between SCNVIP neurons and dorsomedial hypothalamic neurons, providing a circuit substrate by which SCNVIP neurons may regulate LMA rhythms. In vivo photometry revealed that while SCNVIP neurons are acutely responsive to light, their activity is otherwise behavioral state invariant. Single-nuclei RNA-sequencing revealed that SCNVIP neurons comprise two transcriptionally distinct subtypes, including putative pacemaker and non-pacemaker populations. Altogether, our work establishes necessity of SCNVIP neurons for the LMA circadian rhythm, elucidates organization of circadian outflow from and modulatory input to SCNVIP cells, and demonstrates a subpopulation-level molecular heterogeneity that suggests distinct functions for specific SCNVIP subtypes.

Asunto(s)

Ritmo Circadiano/fisiología , Neuronas/metabolismo , Núcleo Supraquiasmático , Animales , Mapeo Encefálico , Relojes Circadianos/fisiología , Locomoción/fisiología , Ratones , Optogenética/métodos , Núcleo Supraquiasmático/citología , Núcleo Supraquiasmático/metabolismo

RESUMEN

The suprachiasmatic nucleus (SCN) acts as a master pacemaker driving circadian behavior and physiology. Although the SCN is small, it is composed of many cell types, making it difficult to study the roles of particular cells. Here we develop bioluminescent circadian reporter mice that are Cre dependent, allowing the circadian properties of genetically defined populations of cells to be studied in real time. Using a Color-Switch PER2::LUCIFERASE reporter that switches from red PER2::LUCIFERASE to green PER2::LUCIFERASE upon Cre recombination, we assess circadian rhythms in two of the major classes of peptidergic neurons in the SCN: AVP (arginine vasopressin) and VIP (vasoactive intestinal polypeptide). Surprisingly, we find that circadian function in AVP neurons, not VIP neurons, is essential for autonomous network synchrony of the SCN and stability of circadian rhythmicity.

Asunto(s)

Arginina Vasopresina/metabolismo , Ritmo Circadiano , Red Nerviosa/metabolismo , Proteínas Circadianas Period/genética , Neuronas del Núcleo Supraquiasmático/metabolismo , Péptido Intestinal Vasoactivo/metabolismo , Factores de Transcripción ARNTL/genética , Animales , Técnicas de Inactivación de Genes , Luciferasas , Ratones , Ratones Transgénicos , Proteínas Circadianas Period/metabolismo , Análisis de la Célula Individual , Núcleo Supraquiasmático/metabolismo , Imagen de Lapso de Tiempo