Participación del receptor de lactato hipotalámico HCAR1 en la regulación de la ingesta de alimentos mediada por neuronas POMC.
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Date
2026
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Universidad de Concepción
Abstract
La obesidad es una epidemia global resultante del desbalance energético crónico, cuya regulación depende críticamente de la capacidad del hipotálamo para detectar nutrientes circulantes. El núcleo arqueado (ARC) en el hipotálamo alberga las neuronas pro-opiomelanocortina (POMC), que promueven la saciedad y aumentan el gasto energético. Si bien, se ha demostrado que el lactato producido y liberado por tanicitos en respuesta a glucosa, incrementa la excitabilidad de las neuronas POMC, en un mecanismo que involucra su transporte, se desconoce su rol como molécula señalizadora intercelular. A pesar de que el lactato activa estas neuronas, carecen del receptor de lactato HCAR1, que sí se expresa en astrocitos hipotalámicos. Esta tesis investigó si el lactato producido por tanicitos promueve la activación de HCAR1 en astrocitos hipotalámicos, incrementando la actividad de neuronas POMC. Confirmamos la expresión de HCAR1 en una subpoblación de astrocitos del ARC y células endoteliales empleando inmunofluorescencia e hibridación in situ, e identificamos novedosamente su expresión en neuronas NPY, a través de citometría de flujo. Por medio de imágenes de calcio in vitro e in situ, demostramos que L-Lactato, D-Lactato y el agonista farmacológico 3Cl-HBA activan HCAR1 en astrocitos, induciendo incrementos robustos en la concentración de calcio intracelular a través de una vía no canónica. Esta respuesta de calcio desencadenó la liberación de glutamato, proceso conocido como gliotransmisión, mediada parcialmente por la apertura de hemicanales de conexina 43, proceso que fue suprimido por el silenciamiento génico de HCAR1 (siRNA/shRNA) o el bloqueo de hemicanales. En experimentos in situ de patch-clamp en neuronas POMC, la estimulación con lactato incrementó la frecuencia de corrientes lentas (SICs), asociadas a gliotransmisión. Estos eventos fueron bloqueados por el antagonista de receptores NMDA, D-AP5, mientras que el silenciamiento específico de HCAR1 en astrocitos, redujo significativamente la frecuencia de SICs in situ. Además, demostramos un acoplamiento funcional en el que la perfusión de glucosa en un tanicito único indujo ondas de calcio en astrocitos adyacentes, de manera dependiente de HCAR1 astroglial. En conclusión, estos hallazgos establecen un nuevo circuito de integración metabólica en el ARC, donde el lactato liberado por tanicitos no solo activa directamente a las neuronas POMC, sino que también actúa por un mecanismo indirecto que involucra la gliotransmisión glutamatérgica extrasináptica inducida por la activación de HCAR1 en astrocitos del ARC. Este mecanismo revela un rol activo y regulatorio de la glía en la homeostasis energética y posiciona al receptor HCAR1 como un blanco potencial para el tratamiento de desórdenes metabólicos.
Obesity is a global epidemic resulting from chronic energy imbalance, the regulation of which critically depends on the hypothalamus's ability to sense circulating nutrients. The arcuate nucleus (ARC) of the hypothalamus houses pro-opiomelanocortin (POMC) neurons, which promote satiety and increase energy expenditure. Although it has been demonstrated that lactate, produced and released by tanycytes in response to glucose, increases the excitability of POMC neurons through a transport-dependent mechanism, its role as an intercellular signaling molecule remains unknown. Despite lactate’s ability to activate these neurons, they lack the lactate receptor HCAR1, which is expressed in hypothalamic astrocytes. This thesis investigated whether tanycyte-derived lactate promotes HCAR1 activation in hypothalamic astrocytes, thereby increasing POMC neuron activity. Using immunofluorescence and in situ hybridization, we confirmed HCAR1 expression in a subpopulation of ARC astrocytes and endothelial cells; notably, we also identified its expression in NPY neurons using flow cytometry. Through in vitro and in situ calcium imaging, we demonstrated that L-lactate, D-lactate, and the pharmacological agonist 3Cl-HBA activate HCAR1 in astrocytes, inducing robust increases in intracellular calcium concentration via a non-canonical pathway. This calcium response triggered glutamate release, a process known as gliotransmission, partially mediated by the opening of connexin 43 hemichannels. This process was suppressed by HCAR1 gene silencing (siRNA/shRNA) or hemichannel blockade. In in situ patch-clamp experiments on POMC neurons, lactate stimulation increased the frequency of slow inward currents (SICs) associated with gliotransmission. These events were blocked by the NMDA receptor antagonist D-AP5, while astrocyte-specific HCAR1 silencing significantly reduced the frequency of SICs in situ. Furthermore, we demonstrated a functional coupling in which glucose perfusion into a single tanycyte induced calcium waves in adjacent astrocytes in an astrocyte-HCAR1-dependent manner. In conclusion, these findings establish a novel metabolic integration circuit in the ARC, where lactate released by tanycytes not only directly activates POMC neurons but also acts through an indirect mechanism involving extrasynaptic glutamatergic gliotransmission induced by HCAR1 activation in ARC astrocytes. This mechanism reveals an active regulatory role for glia in energy homeostasis and positions the HCAR1 receptor as a potential target for the treatment of metabolic disorders.
Obesity is a global epidemic resulting from chronic energy imbalance, the regulation of which critically depends on the hypothalamus's ability to sense circulating nutrients. The arcuate nucleus (ARC) of the hypothalamus houses pro-opiomelanocortin (POMC) neurons, which promote satiety and increase energy expenditure. Although it has been demonstrated that lactate, produced and released by tanycytes in response to glucose, increases the excitability of POMC neurons through a transport-dependent mechanism, its role as an intercellular signaling molecule remains unknown. Despite lactate’s ability to activate these neurons, they lack the lactate receptor HCAR1, which is expressed in hypothalamic astrocytes. This thesis investigated whether tanycyte-derived lactate promotes HCAR1 activation in hypothalamic astrocytes, thereby increasing POMC neuron activity. Using immunofluorescence and in situ hybridization, we confirmed HCAR1 expression in a subpopulation of ARC astrocytes and endothelial cells; notably, we also identified its expression in NPY neurons using flow cytometry. Through in vitro and in situ calcium imaging, we demonstrated that L-lactate, D-lactate, and the pharmacological agonist 3Cl-HBA activate HCAR1 in astrocytes, inducing robust increases in intracellular calcium concentration via a non-canonical pathway. This calcium response triggered glutamate release, a process known as gliotransmission, partially mediated by the opening of connexin 43 hemichannels. This process was suppressed by HCAR1 gene silencing (siRNA/shRNA) or hemichannel blockade. In in situ patch-clamp experiments on POMC neurons, lactate stimulation increased the frequency of slow inward currents (SICs) associated with gliotransmission. These events were blocked by the NMDA receptor antagonist D-AP5, while astrocyte-specific HCAR1 silencing significantly reduced the frequency of SICs in situ. Furthermore, we demonstrated a functional coupling in which glucose perfusion into a single tanycyte induced calcium waves in adjacent astrocytes in an astrocyte-HCAR1-dependent manner. In conclusion, these findings establish a novel metabolic integration circuit in the ARC, where lactate released by tanycytes not only directly activates POMC neurons but also acts through an indirect mechanism involving extrasynaptic glutamatergic gliotransmission induced by HCAR1 activation in ARC astrocytes. This mechanism reveals an active regulatory role for glia in energy homeostasis and positions the HCAR1 receptor as a potential target for the treatment of metabolic disorders.
Description
Tesis presentada para optar al grado de Doctor/a en Ciencias Biológicas con mención en Biología Celular y Molecular.
Keywords
Obesidad, Hipotálamo, Neuronas Metabolismo