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1.
Cell Rep ; 43(6): 114343, 2024 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-38865247

RESUMO

Activation of prepronociceptin (PNOC)-expressing neurons in the arcuate nucleus (ARC) promotes high-fat-diet (HFD)-induced hyperphagia. In turn, PNOCARC neurons can inhibit the anorexic response of proopiomelanocortin (POMC) neurons. Here, we validate the necessity of PNOCARC activity for HFD-induced inhibition of POMC neurons in mice and find that PNOCARC-neuron-dependent inhibition of POMC neurons is mediated by gamma-aminobutyric acid (GABA) release. When monitoring individual PNOCARC neuron activity via Ca2+ imaging, we find a subpopulation of PNOCARC neurons that is inhibited upon gastrointestinal calorie sensing and disinhibited upon HFD feeding. Combining retrograde rabies tracing and circuit mapping, we find that PNOC neurons from the bed nucleus of the stria terminalis (PNOCBNST) provide inhibitory input to PNOCARC neurons, and this inhibitory input is blunted upon HFD feeding. This work sheds light on how an increase in caloric content of the diet can rewire a neuronal circuit, paving the way to overconsumption and obesity development.


Assuntos
Dieta Hiperlipídica , Hiperfagia , Núcleos Septais , Animais , Hiperfagia/metabolismo , Camundongos , Núcleos Septais/metabolismo , Neurônios/metabolismo , Masculino , Ácido gama-Aminobutírico/metabolismo , Pró-Opiomelanocortina/metabolismo , Neurônios GABAérgicos/metabolismo , Núcleo Arqueado do Hipotálamo/metabolismo , Camundongos Endogâmicos C57BL , Precursores de Proteínas , Receptores Opioides
2.
Cell Metab ; 35(5): 786-806.e13, 2023 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-37075752

RESUMO

Autophagy represents a key regulator of aging and metabolism in sensing energy deprivation. We find that fasting in mice activates autophagy in the liver paralleled by activation of hypothalamic AgRP neurons. Optogenetic and chemogenetic activation of AgRP neurons induces autophagy, alters phosphorylation of autophagy regulators, and promotes ketogenesis. AgRP neuron-dependent induction of liver autophagy relies on NPY release in the paraventricular nucleus of the hypothalamus (PVH) via presynaptic inhibition of NPY1R-expressing neurons to activate PVHCRH neurons. Conversely, inhibiting AgRP neurons during energy deprivation abrogates induction of hepatic autophagy and rewiring of metabolism. AgRP neuron activation increases circulating corticosterone concentrations, and reduction of hepatic glucocorticoid receptor expression attenuates AgRP neuron-dependent activation of hepatic autophagy. Collectively, our study reveals a fundamental regulatory principle of liver autophagy in control of metabolic adaptation during nutrient deprivation.


Assuntos
Hipotálamo , Neurônios , Camundongos , Animais , Proteína Relacionada com Agouti/metabolismo , Neurônios/metabolismo , Hipotálamo/metabolismo , Fígado/metabolismo , Nutrientes
3.
JCI Insight ; 7(21)2022 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-36345942

RESUMO

Dopamine acts on neurons in the arcuate nucleus (ARC) of the hypothalamus, which controls homeostatic feeding responses. Here we demonstrate a differential enrichment of dopamine receptor 1 (Drd1) expression in food intake-promoting agouti related peptide (AgRP)/neuropeptide Y (NPY) neurons and a large proportion of Drd2-expressing anorexigenic proopiomelanocortin (POMC) neurons. Owing to the nature of these receptors, this translates into a predominant activation of AgRP/NPY neurons upon dopamine stimulation and a larger proportion of dopamine-inhibited POMC neurons. Employing intersectional targeting of Drd2-expressing POMC neurons, we reveal that dopamine-mediated POMC neuron inhibition is Drd2 dependent and that POMCDrd2+ neurons exhibit differential expression of neuropeptide signaling mediators compared with the global POMC neuron population, which manifests in enhanced somatostatin responsiveness of POMCDrd2+ neurons. Selective chemogenetic activation of POMCDrd2+ neurons uncovered their ability to acutely suppress feeding and to preserve body temperature in fasted mice. Collectively, the present study provides the molecular and functional characterization of POMCDrd2+ neurons and aids our understanding of dopamine-dependent control of homeostatic energy-regulatory neurocircuits.


Assuntos
Dopamina , Pró-Opiomelanocortina , Animais , Camundongos , Proteína Relacionada com Agouti/metabolismo , Temperatura Corporal , Dopamina/metabolismo , Neurônios/metabolismo , Neuropeptídeo Y/metabolismo , Pró-Opiomelanocortina/metabolismo
4.
Nat Neurosci ; 24(7): 913-929, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34002087

RESUMO

Pro-opiomelanocortin (POMC)-expressing neurons in the arcuate nucleus of the hypothalamus represent key regulators of metabolic homeostasis. Electrophysiological and single-cell sequencing experiments have revealed a remarkable degree of heterogeneity of these neurons. However, the exact molecular basis and functional consequences of this heterogeneity have not yet been addressed. Here, we have developed new mouse models in which intersectional Cre/Dre-dependent recombination allowed for successful labeling, translational profiling and functional characterization of distinct POMC neurons expressing the leptin receptor (Lepr) and glucagon like peptide 1 receptor (Glp1r). Our experiments reveal that POMCLepr+ and POMCGlp1r+ neurons represent largely nonoverlapping subpopulations with distinct basic electrophysiological properties. They exhibit a specific anatomical distribution within the arcuate nucleus and differentially express receptors for energy-state communicating hormones and neurotransmitters. Finally, we identify a differential ability of these subpopulations to suppress feeding. Collectively, we reveal a notably distinct functional microarchitecture of critical metabolism-regulatory neurons.


Assuntos
Comportamento Alimentar/fisiologia , Hipotálamo/fisiologia , Neurônios/fisiologia , Pró-Opiomelanocortina/metabolismo , Animais , Metabolismo Energético/fisiologia , Homeostase/fisiologia , Hipotálamo/citologia , Camundongos , Camundongos Transgênicos , Neurônios/citologia
5.
Proc Natl Acad Sci U S A ; 117(25): 14473-14481, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32513737

RESUMO

Hypothalamic tanycytes are chemosensitive glial cells that contact the cerebrospinal fluid in the third ventricle and send processes into the hypothalamic parenchyma. To test whether they can activate neurons of the arcuate nucleus, we targeted expression of a Ca2+-permeable channelrhodopsin (CatCh) specifically to tanycytes. Activation of tanycytes ex vivo depolarized orexigenic (neuropeptide Y/agouti-related protein; NPY/AgRP) and anorexigenic (proopiomelanocortin; POMC) neurons via an ATP-dependent mechanism. In vivo, activation of tanycytes triggered acute hyperphagia only in the fed state during the inactive phase of the light-dark cycle.


Assuntos
Núcleo Arqueado do Hipotálamo/fisiopatologia , Células Ependimogliais/fisiologia , Hiperfagia/fisiopatologia , Neurônios/fisiologia , Proteína Relacionada com Agouti/metabolismo , Animais , Apetite/fisiologia , Núcleo Arqueado do Hipotálamo/citologia , Núcleo Arqueado do Hipotálamo/diagnóstico por imagem , Channelrhodopsins/genética , Channelrhodopsins/metabolismo , Metabolismo Energético/fisiologia , Comportamento Alimentar/fisiologia , Feminino , Genes Reporter , Injeções Intraventriculares , Masculino , Camundongos , Camundongos Transgênicos , Modelos Animais , Rede Nervosa/fisiologia , Neuropeptídeo Y/metabolismo , Imagem Óptica , Optogenética , Técnicas de Patch-Clamp , Pró-Opiomelanocortina/metabolismo , Técnicas Estereotáxicas
6.
Neuron ; 106(6): 1009-1025.e10, 2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32302532

RESUMO

Calorie-rich diets induce hyperphagia and promote obesity, although the underlying mechanisms remain poorly defined. We find that short-term high-fat-diet (HFD) feeding of mice activates prepronociceptin (PNOC)-expressing neurons in the arcuate nucleus of the hypothalamus (ARC). PNOCARC neurons represent a previously unrecognized GABAergic population of ARC neurons distinct from well-defined feeding regulatory AgRP or POMC neurons. PNOCARC neurons arborize densely in the ARC and provide inhibitory synaptic input to nearby anorexigenic POMC neurons. Optogenetic activation of PNOCARC neurons in the ARC and their projections to the bed nucleus of the stria terminalis promotes feeding. Selective ablation of these cells promotes the activation of POMC neurons upon HFD exposure, reduces feeding, and protects from obesity, but it does not affect food intake or body weight under normal chow consumption. We characterize PNOCARC neurons as a novel ARC neuron population activated upon palatable food consumption to promote hyperphagia.


Assuntos
Núcleo Arqueado do Hipotálamo/fisiologia , Dieta Hiperlipídica , Comportamento Alimentar/fisiologia , Neurônios GABAérgicos/fisiologia , Hiperfagia , Obesidade , Aumento de Peso/fisiologia , Animais , Núcleo Arqueado do Hipotálamo/citologia , Núcleo Arqueado do Hipotálamo/metabolismo , Neurônios GABAérgicos/metabolismo , Camundongos , Inibição Neural/fisiologia , Neurônios/metabolismo , Neurônios/fisiologia , Optogenética , Pró-Opiomelanocortina/metabolismo , Precursores de Proteínas/metabolismo , Receptores Opioides/metabolismo , Núcleos Septais/fisiologia
7.
Proc Natl Acad Sci U S A ; 112(35): 11090-5, 2015 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-26286989

RESUMO

Neural activity is accompanied by a transient mismatch between local glucose and oxygen metabolism, a phenomenon of physiological and pathophysiological importance termed aerobic glycolysis. Previous studies have proposed glutamate and K(+) as the neuronal signals that trigger aerobic glycolysis in astrocytes. Here we used a panel of genetically encoded FRET sensors in vitro and in vivo to investigate the participation of NH4(+), a by-product of catabolism that is also released by active neurons. Astrocytes in mixed cortical cultures responded to physiological levels of NH4(+) with an acute rise in cytosolic lactate followed by lactate release into the extracellular space, as detected by a lactate-sniffer. An acute increase in astrocytic lactate was also observed in acute hippocampal slices exposed to NH4(+) and in the somatosensory cortex of anesthetized mice in response to i.v. NH4(+). Unexpectedly, NH4(+) had no effect on astrocytic glucose consumption. Parallel measurements showed simultaneous cytosolic pyruvate accumulation and NADH depletion, suggesting the involvement of mitochondria. An inhibitor-stop technique confirmed a strong inhibition of mitochondrial pyruvate uptake that can be explained by mitochondrial matrix acidification. These results show that physiological NH4(+) diverts the flux of pyruvate from mitochondria to lactate production and release. Considering that NH4(+) is produced stoichiometrically with glutamate during excitatory neurotransmission, we propose that NH4(+) behaves as an intercellular signal and that pyruvate shunting contributes to aerobic lactate production by astrocytes.


Assuntos
Compostos de Amônio/metabolismo , Astrócitos/metabolismo , Ácido Láctico/metabolismo , Mitocôndrias/metabolismo , Ácido Pirúvico/metabolismo , Animais , Camundongos
8.
J Neurosci ; 35(10): 4168-78, 2015 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-25762664

RESUMO

Excitatory synaptic transmission is accompanied by a local surge in interstitial lactate that occurs despite adequate oxygen availability, a puzzling phenomenon termed aerobic glycolysis. In addition to its role as an energy substrate, recent studies have shown that lactate modulates neuronal excitability acting through various targets, including NMDA receptors and G-protein-coupled receptors specific for lactate, but little is known about the cellular and molecular mechanisms responsible for the increase in interstitial lactate. Using a panel of genetically encoded fluorescence nanosensors for energy metabolites, we show here that mouse astrocytes in culture, in cortical slices, and in vivo maintain a steady-state reservoir of lactate. The reservoir was released to the extracellular space immediately after exposure of astrocytes to a physiological rise in extracellular K(+) or cell depolarization. Cell-attached patch-clamp analysis of cultured astrocytes revealed a 37 pS lactate-permeable ion channel activated by cell depolarization. The channel was modulated by lactate itself, resulting in a positive feedback loop for lactate release. A rapid fall in intracellular lactate levels was also observed in cortical astrocytes of anesthetized mice in response to local field stimulation. The existence of an astrocytic lactate reservoir and its quick mobilization via an ion channel in response to a neuronal cue provides fresh support to lactate roles in neuronal fueling and in gliotransmission.


Assuntos
Astrócitos/efeitos dos fármacos , Canais Iônicos/fisiologia , Ácido Láctico/metabolismo , Potássio/farmacologia , Animais , Animais Recém-Nascidos , Bário/farmacologia , Cádmio/farmacologia , Células Cultivadas , Córtex Cerebral/citologia , Feminino , Fluoresceínas/metabolismo , Glicogênio/metabolismo , Humanos , Técnicas In Vitro , Canais Iônicos/efeitos dos fármacos , Íons/farmacologia , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Ácido Pirúvico/farmacologia , Córtex Somatossensorial/citologia , Córtex Somatossensorial/fisiologia , Transfecção
9.
Neurophotonics ; 1(1): 011004, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26157964

RESUMO

Neurophotonics comes to light at a time in which advances in microscopy and improved calcium reporters are paving the way toward high-resolution functional mapping of the brain. This review relates to a parallel revolution in metabolism. We argue that metabolism needs to be approached both in vitro and in vivo, and that it does not just exist as a low-level platform but is also a relevant player in information processing. In recent years, genetically encoded fluorescent nanosensors have been introduced to measure glucose, glutamate, ATP, NADH, lactate, and pyruvate in mammalian cells. Reporting relative metabolite levels, absolute concentrations, and metabolic fluxes, these sensors are instrumental for the discovery of new molecular mechanisms. Sensors continue to be developed, which together with a continued improvement in protein expression strategies and new imaging technologies, herald an exciting era of high-resolution characterization of metabolism in the brain and other organs.

10.
J Neurosci ; 31(40): 14264-71, 2011 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-21976511

RESUMO

Excitatory synaptic transmission stimulates brain tissue glycolysis. This phenomenon is the signal detected in FDG-PET imaging and, through enhanced lactate production, is also thought to contribute to the fMRI signal. Using a method based on Förster resonance energy transfer in mouse astrocytes, we have recently observed that a small rise in extracellular K(+) can stimulate glycolysis by >300% within seconds. The K(+) response was blocked by ouabain, but intracellular engagement of the Na(+)/K(+) ATPase pump with Na(+) was ineffective, suggesting that the canonical feedback regulatory pathway involving the Na(+) pump and ATP depletion is only permissive and that a second mechanism is involved. Because of their predominant K(+) permeability and high expression of the electrogenic Na(+)/HCO(3)(-) cotransporter NBCe1, astrocytes respond to a rise in extracellular K(+) with plasma membrane depolarization and intracellular alkalinization. In the present article, we show that a fast glycolytic response can be elicited independently of K(+) by plasma membrane depolarization or by intracellular alkalinization. The glycolytic response to K(+) was absent in astrocytes from NBCe1 null mice (Slc4a4) and was blocked by functional or pharmacological inhibition of the NBCe1. Hippocampal neurons acquired K(+)-sensitive glycolysis upon heterologous NBCe1 expression. The phenomenon could also be reconstituted in HEK293 cells by coexpression of the NBCe1 and a constitutively open K(+) channel. We conclude that the NBCe1 is a key element in a feedforward mechanism linking excitatory synaptic transmission to fast modulation of glycolysis in astrocytes.


Assuntos
Astrócitos/metabolismo , Espaço Extracelular/metabolismo , Glicólise/fisiologia , Potássio/metabolismo , Simportadores de Sódio-Bicarbonato/fisiologia , Animais , Células Cultivadas , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Fatores de Tempo
11.
J Neurosci ; 31(12): 4709-13, 2011 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-21430169

RESUMO

Synaptic activity is followed within seconds by a local surge in lactate concentration, a phenomenon that underlies functional magnetic resonance imaging and whose causal mechanisms are unclear, partly because of the limited spatiotemporal resolution of standard measurement techniques. Using a novel Förster resonance energy transfer-based method that allows real-time measurement of the glycolytic rate in single cells, we have studied mouse astrocytes in search for the mechanisms responsible for the lactate surge. Consistent with previous measurements with isotopic 2-deoxyglucose, glutamate was observed to stimulate glycolysis in cultured astrocytes, but the response appeared only after a lag period of several minutes. Na(+) overloads elicited by engagement of the Na(+)-glutamate cotransporter with d-aspartate or application of the Na(+) ionophore gramicidin also failed to stimulate glycolysis in the short term. In marked contrast, K(+) stimulated astrocytic glycolysis by fourfold within seconds, an effect that was observed at low millimolar concentrations and was also present in organotypic hippocampal slices. After removal of the agonists, the stimulation by K(+) ended immediately but the stimulation by glutamate persisted unabated for >20 min. Both stimulations required an active Na(+)/K(+) ATPase pump. By showing that small rises in extracellular K(+) mediate short-term, reversible modulation of astrocytic glycolysis and that glutamate plays a long-term effect and leaves a metabolic trace, these results support the view that astrocytes contribute to the lactate surge that accompanies synaptic activity and underscore the role of these cells in neurometabolic and neurovascular coupling.


Assuntos
Astrócitos/fisiologia , Ácido Glutâmico/farmacologia , Glicólise/fisiologia , Potássio/farmacologia , Animais , Células Cultivadas , Transferência Ressonante de Energia de Fluorescência , Técnicas In Vitro , Indicadores e Reagentes , Cinética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , ATPase Trocadora de Sódio-Potássio/metabolismo , Estimulação Química
12.
Artigo em Inglês | MEDLINE | ID: mdl-20890447

RESUMO

The glycolytic rate is sensitive to physiological activity, hormones, stress, aging, and malignant transformation. Standard techniques to measure the glycolytic rate are based on radioactive isotopes, are not able to resolve single cells and have poor temporal resolution, limitations that hamper the study of energy metabolism in the brain and other organs. A new method is described in this article, which makes use of a recently developed FRET glucose nanosensor to measure the rate of glycolysis in single cells with high temporal resolution. Used in cultured astrocytes, the method showed for the first time that glycolysis can be activated within seconds by a combination of glutamate and K(+), supporting a role for astrocytes in neurometabolic and neurovascular coupling in the brain. It was also possible to make a direct comparison of metabolism in neurons and astrocytes lying in close proximity, paving the way to a high-resolution characterization of brain energy metabolism. Single-cell glycolytic rates were also measured in fibroblasts, adipocytes, myoblasts, and tumor cells, showing higher rates for undifferentiated cells and significant metabolic heterogeneity within cell types. This method should facilitate the investigation of tissue metabolism at the single-cell level and is readily adaptable for high-throughput analysis.

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