Non-Neuronal Cells in the Hypothalamic Adaptation to Metabolic Signals

Although the brain is composed of numerous cell types, neurons have received the vast majority of attention in the attempt to understand how this organ functions. Neurons are indeed fundamental but, in order for them to function correctly, they rely on the surrounding “non-neuronal” cells. These different cell types, which include glia, epithelial cells, pericytes, and endothelia, supply essential substances to neurons, in addition to protecting them from dangerous substances and situations. Moreover, it is now clear that non-neuronal cells can also actively participate in determining neuronal signaling outcomes. Due to the increasing problem of obesity in industrialized countries, investigation of the central control of energy balance has greatly increased in attempts to identify new therapeutic targets. This has led to interesting advances in our understanding of how appetite and systemic metabolism are modulated by non-neuronal cells. For example, not only are nutrients and hormones transported into the brain by non-neuronal cells, but these cells can also metabolize these metabolic factors, thus modifying the signals reaching the neurons. The hypothalamus is the main integrating center of incoming metabolic and hormonal signals and interprets this information in order to control appetite and systemic metabolism. Hence, the factors transported and released from surrounding non-neuronal cells will undoubtedly influence metabolic homeostasis. This review focuses on what is known to date regarding the involvement of different cell types in the transport and metabolism of nutrients and hormones in the hypothalamus. The possible involvement of non-neuronal cells, in particular glial cells, in physiopathological outcomes of poor dietary habits and excess weight gain are also discussed.The authors are funded by grants from the Spanish Ministry of Science and Innovation (BFU2014-51836-C2-2 to JAC and BFU2014-51836-C2-1 to LG-S), Spanish Ministry of Education, Culture and Sports (university training grant FPU13/00909 to AF-R), Fondo de Investigación Sanitaria (PI-1302195, PI-1600485, and CIBEROBN to JA and CIBERFES to LG-S) and Fondos FEDER.Peer reviewedPeer Reviewe

Estradiol uses different mechanisms in astrocytes from the hippocampus of male and female rats to protect against damage Induced by palmitic acid

An excess of saturated fatty acids can be toxic for tissues, including the brain, and this has been associated with the progression of neurodegenerative diseases. Since palmitic acid (PA) is a free fatty acid that is abundant in the diet and circulation and can be harmful, we have investigated the effects of this fatty acid on lipotoxicity in hippocampal astrocytes and the mechanism involved. Moreover, as males and females have different susceptibilities to some neurodegenerative diseases, we accessed the responses of astrocytes from both sexes, as well as the possible involvement of estrogens in the protection against fatty acid toxicity. PA increased endoplasmic reticulum stress leading to cell death in astrocytes from both males and females. Estradiol (E2) increased the levels of protective factors, such as Hsp70 and the anti-inflammatory cytokine interleukin-10, in astrocytes from both sexes. In male astrocytes, E2 decreased pJNK, TNFα, and caspase-3 activation. In contrast, in female astrocytes E2 did not affect the activation of JNK or TNFα levels, but decreased apoptotic cell death. Hence, although E2 exerted protective effects against the detrimental effects of PA, the mechanisms involved appear to be different between male and female astrocytes. This sexually dimorphic difference in the protective mechanisms induced by E2 could be involved in the different susceptibilities of males and females to some neurodegenerative processesThis work was funded by grants from Ministerio de Ciencia e Innovación (BFU2014-51836-C2-2-R to JC and BFU2014-51836-C2-1-R to LG-S) and Fondos de Investigación Sanitaria (Grant PI16/00485 to JA), co-funded by European FEDER Program, and Centro de Investigación Biomédica en Red Fisiopatología de Obesidad y Nutrición (CIBEROBN) of the Instituto de Salud Carlos III, and Fundación de Endocrinología y Nutrició

Neonatal overnutrition increases testicular size and expression of luteinizing hormone ß-subunit in peripubertal male rats

Proper nutrition is important for growth and development. Maturation of the reproductive axis and the timing of pubertal onset can be delayed when insufficient nutrition is available, or possibly advanced with nutritional abundance. The childhood obesity epidemic has been linked to a secular trend in advanced puberty in some populations. The increase in circulating leptin that occurs in association with obesity has been suggested to act as a signal that an adequate nutritional status exists for puberty to occur, allowing activation of central mechanisms. However, obesity-associated hyperleptinemia is linked to decreased leptin sensitivity, at least in adults. Here, we analyzed whether neonatal overnutrition modifies the response to an increase in leptin in peripubertal male rats, as previously demonstrated in females. Wistar rats were raised in litters of 4 (neonatal overnutrition) or 12 pups (controls) per dam. Leptin was administered sc (3 μg/g body weight) at postnatal day 35 and the rats killed 45 min or 2 h later. Postnatal overfeeding resulted in increased body weight and circulating leptin levels; however, we found no overweight-related changes in the mRNA levels of neuropeptides involved in metabolism or reproduction. In contrast, pituitary expression of luteinizing hormone (LH) beta-subunit was increased in overweight rats, as was testicular weight. There were no basal differences between L4 and L12 males or in their response to leptin administration in pSTAT3 levels in the hypothalamus at either 45 min or 2 h. In contrast, pJAK2 was found to be higher at 45 min in L4 compared to L12 males regardless of leptin treatment, while at 2 h it was higher in L4 leptin-treated males compared to L12 leptin-treated males, as well as L4 vehicle-treated rats. There were no changes in response to leptin administration in the expression of the neuropeptides analyzed. However, serum LH levels rose only in L4 males in response to leptin, but with no change in testosterone levels. In conclusion, the advancement in pubertal onset in males with neonatal overnutrition does not appear to be related to overt modifications in the central response to exogenous leptin during the peripubertal periodThe authors are funded by Fondos de Investigación Sanitaria (PI1600485 to JA), Ministerio de Ciencia e Innovación (BFU2014-51836-C2-2-R and BFU2017-82565-C2-1-R to JC) and fondos FEDER, Centro de Investigación Biomédica en Red Fisiopatología de Obesidad y Nutrición (CIBEROBN), Instituto de Salud Carlos III (JA), and Fundación Endocrinología y Nutrició

Age and sex dependent effects of early overnutrition on metabolic parameters and the role of neonatal androgens

Background: Males and females respond differently to diverse metabolic situations. Being raised in a small litter is reported to cause overnutrition that increases weight gain and predisposes an individual to metabolic disturbances in adulthood; however, existing data are inconsistent. Indeed, significant weight gain and/or metabolic disturbances, such as hyperinsulinemia and hyperleptinemia, are sometimes not encountered. We hypothesized that these inconsistencies could be due to the animal’s sex and/or age at which metabolic parameters are measured. Methods: To analyze the effects of neonatal overnutrition, male and female Wistar rats were raised in litters of 4 or 12 pups/dam and killed at postnatal days (PND) 10, 21, 30, 50, 85, or 150. In a second study to determine if neonatal sex steroid levels influence sex differences in metabolic parameters, female rats were treated with testosterone on PND1. Effects on weight, length, fat pads, adipokine production, and serum levels of glucose, metabolic hormones, and cytokines were analyzed in both studies. Results: By PND10, both males and females raised in small litters had increased body weight, body length, adiposity, and serum glucose, insulin, leptin, and adiponectin levels. Females had a greater increase in inguinal fat, and males had higher expression of leptin messenger RNA (mRNA) and serum insulin, as well as increased testosterone levels. Most of the litter size effects diminished or disappeared after weaning and reappeared during adulthood in males, with sex differences in body size and adiposity being apparent postpubertally. Treatment of females with testosterone on PND1 tended to masculinize some metabolic parameters in adulthood such as increased body weight and serum leptin levels. Conclusions: Our results indicate that (1) both sex and age determine the response to neonatal overnutrition; (2) differences in neonatal sex steroid levels may participate in the development of sex differences in metabolic parameters in adulthood and possibly in the response to neonatal overnutrition; and (3) the comparison of circulating hormone and cytokine levels, even in normal control animals, should take into consideration the early neonatal nutritional environmentThis work was supported by the Ministerio de Ciencia e Innovación Grants (BFU2011–27492 and BFU2014-51836-C2-2-R (to JAC); BFI2011-25021 (to MTS)) and Fondo de Investigación Sanitaria (PI13/02195 to JA) co-funded by European FEDER Program, Junta de Andalucia, Spain (Project FQM12-1943 to MTS) and Centro de Investigación Biomédica en Red Fisiopatología de Obesidad y Nutrición (JA and MTS) of the Instituto de Salud Carlos III, and Fundación de Endocrinología y Nutrició

Ghrelin regulates glucose and glutamate transporters in hypothalamic astrocytes

Hypothalamic astrocytes can respond to metabolic signals, such as leptin and insulin, to modulate adjacent neuronal circuits and systemic metabolism. Ghrelin regulates appetite, adiposity and glucose metabolism, but little is known regarding the response of astrocytes to this orexigenic hormone. We have used both in vivo and in vitro approaches to demonstrate that acylated ghrelin (acyl-ghrelin) rapidly stimulates glutamate transporter expression and glutamate uptake by astrocytes. Moreover, acyl-ghrelin rapidly reduces glucose transporter (GLUT) 2 levels and glucose uptake by these glial cells. Glutamine synthetase and lactate dehydrogenase decrease, while glycogen phosphorylase and lactate transporters increase in response to acyl-ghrelin, suggesting a change in glutamate and glucose metabolism, as well as glycogen storage by astrocytes. These effects are partially mediated through ghrelin receptor 1A (GHSR-1A) as astrocytes do not respond equally to desacyl-ghrelin, an isoform that does not activate GHSR-1A. Moreover, primary astrocyte cultures from GHSR-1A knock-out mice do not change glutamate transporter or GLUT2 levels in response to acyl-ghrelin. Our results indicate that acyl-ghrelin may mediate part of its metabolic actions through modulation of hypothalamic astrocytes and that this effect could involve astrocyte mediated changes in local glucose and glutamate metabolism that alter the signals/nutrients reaching neighboring neurons.This work was funded by grants from Fondo de Investigación Sanitaria (PI100747; PI1302195), Ministerio de Ciencia e Innovación (BFU2011–27492; BFU2014-51836-C2-2-R) and Fondos FEDER, Centro de Investigación Biomédica en Red Fisiopatología de Obesidad y Nutrición (CIBEROBN), Instituto de Salud Carlos III, and Fundación de Endocrinología y Nutrición. SD was funded by the Swedish Research Council (Vetenskapsrådet grant 2012– 1758), Läkarutbildningsavtalet Göteborg grant at Sahlgrenska Hospital (ALFGBG-138741, The European Union Seventh Framework Programme under Grant Agreement 607310, Nudge-it

Sex and age as determining factors in the peripheral and central responses to neonatal overnutrition: Possible implications of hypothalamic astrocytes in the sexual dimorphic response to metabolic challenges

Tesis doctoral inédita, leída en la Universidad Autónoma de Madrid, Facultad de Medicina, Departamento de Pediatría. Fecha de lectura: 20 de mayo de 2016The responses to metabolic challenges are suggested to be different in males and females. Not only is there a difference in weight gain and adipose tissue accumulation and distribution in response to increased energy intake, but the secondary complications in response to being overweight or obese also differ between the sexes. However, less is known about the differential sex response to early nutritional changes that can affect long-term metabolism. In addition, recent studies have indicated that hypothalamic astrocytes are involved in the neuroendocrine control of metabolism, as well as the development of secondary complications in response to obesity. However, whether hypothalamic astrocytes from male and female rats respond differently to metabolic challenges remains to be thoroughly analyzed. The aim of this thesis was to determine if neonatal overnutrition as a consequence of the reduction in the number of pups per dam during nursing produces similar effects on growth and metabolism in male and female rats throughout development. Moreover, although sex steroids are clearly involved in the post-pubertal differences between males and females in metabolism and adipose tissue distribution and function, less is known regarding the effect of changes in the neonatal sex steroid environment on metabolism. Hence, we employed a model of neonatal androgenization of females to determine how this affected both short-term and long-term metabolism. Metabolic profile, inflammatory responses and adipose tissue distribution and function, as well as possible alterations in hypothalamic astrocytes were analyzed in these studies. To further explore the sex differences in hypothalamic astrocytes, the response of these glial cells to fatty acids and the possible protective effect of estrogens against fatty acid-induced inflammation were analyzed in vitro employing primary hypothalamic astrocyte cultures. The results of these studies indicate that the response to nutritional disturbances during the neonatal period differ between the sexes and are age dependent. In addition to changes in insulin and leptin levels, differences in the sex steroid environment during the neonatal stage could participate in the distinct long-term metabolic responses to neonatal overnutrition and the differential effects in males and females. We also demonstrate that hypothalamic astrocytes of male and female rats respond differently to neonatal overnutrition, as well as to fatty acid and hormonal treatments in vitro. Thus, hypothalamic astrocytes are not only involved in the physiological and physiopathological neuroendocrine control of metabolism, but they may also participate in developmental responses to early nutritional changes and sexually dimorphic metabolic responses.Diversos estudios sugieren que la respuesta a los desafíos metabólicos es diferente entre machos y hembras. Como consecuencia de un aumento en el consumo de calorías, se producen diferencias en la ganancia de peso, así como en el aumento y distribución de tejido adiposo, pero, además, la respuesta al sobrepeso y obesidad es diferente entre los sexos. Sin embargo, aunque se sabe que las alteraciones nutricionales tempranas pueden tener efectos a largo plazo en el metabolismo, las diferentes respuestas entre los sexos son insuficientemente conocidas hasta la fecha. Junto a ello, estudios recientes indican que los astrocitos hipotalámicos están implicados en el control neuroendocrino del metabolismo, así como en el desarrollo de las complicaciones secundarias asociadas a la obesidad. No obstante, se desconoce en gran medida si los astrocitos hipotalámicos de machos y hembras responden de manera diferente a los desafíos metabólicos. El objetivo de esta Tesis fue determinar si la sobrenutrición neonatal como resultado de una reducción en el número de crías por madre durante el período de lactancia, produce efectos similares en el crecimiento y metabolismo de ratas macho y hembra a lo largo del desarrollo. Es bien sabido que los esteroides sexuales desempeñan una función crítica en las diferencias post-puberales entre machos y hembras en su metabolismo y distribución y función del tejido adiposo. No obstante, los efectos metabólicos de las variaciones en los esteroides sexuales durante la etapa neonatal deben ser estudiados en mayor profundidad. Por esta razón, empleamos también un modelo de androgenización neonatal en hembras para determinar sus efectos a corto y largo plazo en el metabolismo. Igualmente, la respuesta hipotalámica y, concretamente de los astrocitos hipotalámicos a la sobrenutrición neonatal, fue estudiada en ambos sexos. Por tanto, en el estudio de sobrenutrición neonatal en machos y hembras, se analizó el perfil metabólico, las respuestas inflamatorias y distribución y función del tejido adiposo, así como posibles alteraciones en los astrocitos hipotalámicos. Para ahondar en el estudio de las posibles diferencias entre los sexos específicamente en los astrocitos hipotalámicos, se estudió la respuesta in vitro de estas células gliales a determinados ácidos grasos, así como el posible efecto protector de los estrógenos a la inflamación producida por los ácidos grasos. Los resultados de estos estudios indican la diferente respuesta entre los sexos a alteraciones nutricionales en el periodo neonatal, siendo esta respuesta diferente también, en función de la edad o etapa del desarrollo en la que se encuentran. Asimismo, las modificaciones en los esteroides sexuales en el neonato, podrían contribuir a las diferencias metabólicas observadas a largo plazo a consecuencia de la sobrenutrición neonatal, así como en los diferentes efectos vistos en machos y hembras. Aún más, los astrocitos hipotalámicos procedentes de ratas macho y hembra, responden de manera diferente a la sobrenutrición neonatal, así como a los tratamientos in vitro con ácidos grasos y estrógenos. Por consiguiente, los astrocitos hipotalámicos no solamente estarían implicados en la fisiología y fisiopatología del control neuroendocrino del metabolismo, sino que también podrían estar participando en las respuestas a los cambios nutricionales tempranos y en las respuestas sexualmente dimórficas del sistema metabólico

Neonatal Overnutrition Increases Testicular Size and Expression of Luteinizing Hormone β-Subunit in Peripubertal Male Rats

Proper nutrition is important for growth and development. Maturation of the reproductive axis and the timing of pubertal onset can be delayed when insufficient nutrition is available, or possibly advanced with nutritional abundance. The childhood obesity epidemic has been linked to a secular trend in advanced puberty in some populations. The increase in circulating leptin that occurs in association with obesity has been suggested to act as a signal that an adequate nutritional status exists for puberty to occur, allowing activation of central mechanisms. However, obesity-associated hyperleptinemia is linked to decreased leptin sensitivity, at least in adults. Here, we analyzed whether neonatal overnutrition modifies the response to an increase in leptin in peripubertal male rats, as previously demonstrated in females. Wistar rats were raised in litters of 4 (neonatal overnutrition) or 12 pups (controls) per dam. Leptin was administered sc (3 µg/g body weight) at postnatal day 35 and the rats killed 45 min or 2 h later. Postnatal overfeeding resulted in increased body weight and circulating leptin levels; however, we found no overweight-related changes in the mRNA levels of neuropeptides involved in metabolism or reproduction. In contrast, pituitary expression of luteinizing hormone (LH) beta-subunit was increased in overweight rats, as was testicular weight. There were no basal differences between L4 and L12 males or in their response to leptin administration in pSTAT3 levels in the hypothalamus at either 45 min or 2 h. In contrast, pJAK2 was found to be higher at 45 min in L4 compared to L12 males regardless of leptin treatment, while at 2 h it was higher in L4 leptin-treated males compared to L12 leptin-treated males, as well as L4 vehicle-treated rats. There were no changes in response to leptin administration in the expression of the neuropeptides analyzed. However, serum LH levels rose only in L4 males in response to leptin, but with no change in testosterone levels. In conclusion, the advancement in pubertal onset in males with neonatal overnutrition does not appear to be related to overt modifications in the central response to exogenous leptin during the peripubertal period

Estradiol Uses Different Mechanisms in Astrocytes from the Hippocampus of Male and Female Rats to Protect against Damage Induced by Palmitic Acid

An excess of saturated fatty acids can be toxic for tissues, including the brain, and this has been associated with the progression of neurodegenerative diseases. Since palmitic acid (PA) is a free fatty acid that is abundant in the diet and circulation and can be harmful, we have investigated the effects of this fatty acid on lipotoxicity in hippocampal astrocytes and the mechanism involved. Moreover, as males and females have different susceptibilities to some neurodegenerative diseases, we accessed the responses of astrocytes from both sexes, as well as the possible involvement of estrogens in the protection against fatty acid toxicity. PA increased endoplasmic reticulum stress leading to cell death in astrocytes from both males and females. Estradiol (E2) increased the levels of protective factors, such as Hsp70 and the anti-inflammatory cytokine interleukin-10, in astrocytes from both sexes. In male astrocytes, E2 decreased pJNK, TNFα, and caspase-3 activation. In contrast, in female astrocytes E2 did not affect the activation of JNK or TNFα levels, but decreased apoptotic cell death. Hence, although E2 exerted protective effects against the detrimental effects of PA, the mechanisms involved appear to be different between male and female astrocytes. This sexually dimorphic difference in the protective mechanisms induced by E2 could be involved in the different susceptibilities of males and females to some neurodegenerative processes

Glial cells and energy balance

The search for new strategies and drugs to abate the current obesity epidemic has led to the intensification of research aimed at understanding the neuroendocrine control of appetite and energy expenditure. This intensified investigation of metabolic control has also included the study of how glial cells participate in this process. Glia, the most abundant cell type in the central nervous system, perform a wide spectrum of functions and are vital for the correct functioning of neurons and neuronal circuits. Current evidence indicates that hypothalamic glia, in particular astrocytes, tanycytes and microglia, are involved in both physiological and pathophysiological mechanisms of appetite and metabolic control, at least in part by regulating the signals reaching metabolic neuronal circuits. Glia transport nutrients, hormones and neurotransmitters; they secrete growth factors, hormones, cytokines and gliotransmitters and are a source of neuroprogenitor cells. These functions are regulated, as glia also respond to numerous hormones and nutrients, with the lack of specific hormonal signaling in hypothalamic astrocytes disrupting metabolic homeostasis. Here, we review some of the more recent advances in the role of glial cells in metabolic control, with a special emphasis on the differences between glial cell responses in males and females.The authors are funded by grants from Ministerio de Ciencia e Innovación (BFU2014-51836-C2-1-R and BFU2014-51836-C2-2-R), Fondo de Investigación Sanitaria (PI1302195), and Fondos FEDER, Centro de Investigación Biomédica en Red Fisiopatología de Obesidad y Nutrición (CIBEROBN) Instituto de Salud Carlos III and Fundación de Endocrinología y Nutrición