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Acyl-CoA binding protein (ACBP), also known as diazepam binding inhibitor (DBI), has recently emerged as a hypothalamic and brainstem gliopeptide regulating energy balance. Previous work has shown that the ACBP-derived octadecaneuropeptide exerts strong anorectic action via proopiomelanocortin (POMC) neuron activation and the melanocortin-4 receptor. Importantly, targeted ACBP loss-of-function in astrocytes promotes hyperphagia and diet-induced obesity while its overexpression in arcuate astrocytes reduces feeding and body weight. Despite this knowledge, the role of astroglial ACBP in adaptive feeding and metabolic responses to acute metabolic challenges has not been investigated. Using different paradigms, we found that ACBP deletion in glial fibrillary acidic protein (GFAP)-positive astrocytes does not affect weight loss when obese male mice are transitioned from a high fat diet to a chow diet, nor metabolic parameters in mice fed with a normal chow diet (e.g., energy expenditure, body temperature) during fasting, cold exposure and at thermoneutrality. In contrast, astroglial ACBP deletion impairs meal pattern and feeding responses during refeeding after a fast and during cold exposure, thereby showing that ACBP is required to stimulate feeding in states of increased energy demand. These findings challenge the general view that astroglial ACBP exerts anorectic effects and suggest that regulation of feeding by ACBP is dependent on metabolic status.

Aims/hypothesis Hypoglycaemia is a major barrier to good glucose control in type 1 diabetes. Frequent hypoglycaemic episodes impair awareness of subsequent hypoglycaemic bouts. Neural changes underpinning awareness of hypoglycaemia are poorly defined and molecular mechanisms by which glial cells contribute to hypoglycaemia sensing and glucose counterregulation require further investigation. The aim of the current study was to examine whether, and by what mechanism, human primary astrocyte (HPA) function was altered by acute and recurrent low glucose (RLG). Methods To test whether glia, specifically astrocytes, could detect changes in glucose, we utilised HPA and U373 astrocytoma cells and exposed them to RLG in vitro. This allowed measurement, with high specificity and sensitivity, of RLG-associated changes in cellular metabolism. We examined changes in protein phosphorylation/expression using western blotting. Metabolic function was assessed using a Seahorse extracellular flux analyser. Immunofluorescent imaging was used to examine cell morphology and enzymatic assays were used to measure lactate release, glycogen content, intracellular ATP and nucleotide ratios. Results AMP-activated protein kinase (AMPK) was activated over a pathophysiologically relevant glucose concentration range. RLG produced an increased dependency on fatty acid oxidation for basal mitochondrial metabolism and exhibited hallmarks of mitochondrial stress, including increased proton leak and reduced coupling efficiency. Relative to glucose availability, lactate release increased during low glucose but this was not modified by RLG. Basal glucose uptake was not modified by RLG and glycogen levels were similar in control and RLG-treated cells. Mitochondrial adaptations to RLG were partially recovered by maintaining euglycaemic levels of glucose following RLG exposure. Conclusions/interpretation Taken together, these data indicate that HPA mitochondria are altered following RLG, with a metabolic switch towards increased fatty acid oxidation, suggesting glial adaptations to RLG involve altered mitochondrial metabolism that could contribute to defective glucose counterregulation to hypoglycaemia in diabetes. Electronic supplementary material The online version of this article (10.1007/s00125-018-4744-6) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

Aim To test the hypothesis that, given the role of AMP-activated protein kinase (AMPK) in regulating intracellular ATP levels, AMPK may alter ATP release from astrocytes, the main sources of extracellular ATP (eATP) within the brain. Materials and Methods Measurements of ATP release were made from human U373 astrocytoma cells, primary mouse hypothalamic (HTAS) and cortical astrocytes (CRTAS) and wild-type and AMPK α1/α2 null mouse embryonic fibroblasts (MEFs). Cells were treated with drugs known to modulate AMPK activity: A-769662, AICAR and metformin, for up to 3 hours. Intracellular calcium was measured using Fluo4 and Fura-2 calcium-sensitive fluorescent dyes. Results In U373 cells, A-769662 (100 μM) increased AMPK phosphorylation, whereas AICAR and metformin (1 mM) induced a modest increase or had no effect, respectively. Only A-769662 increased eATP levels, and this was partially blocked by AMPK inhibitor Compound C. A-769662-induced increases in eATP were preserved in AMPK α1/α2 null MEF cells. A-769662 increased intracellular calcium in U373, HTAS and CRTAS cells and chelation of intracellular calcium using BAPTA-AM reduced A-769662-induced eATP levels. A-769662 also increased ATP release from a number of other central and peripheral endocrine cell types. Conclusions AMPK is required to maintain basal eATP levels but is not required for A-769662-induced increases in eATP. A-769662 (>50 μM) enhanced intracellular calcium levels leading to ATP release in an AMPK and purinergic receptor independent pathway.

Inflammation and metabolism are intrinsically linked with inflammatory stimuli inducing metabolic changes in cells and, in turn, metabolic capacity determining cellular inflammatory responses. Although well characterized in peripheral immune cells there is comparatively less known about these "immunometabolic" responses in astrocytes. In this study, we tested the hypothesis that the astrocytic inflammatory response driven by nuclear factor-kappa B (NF-κB) signaling is dependent on glycolytic metabolism. Using mouse primary cortical astrocyte cultures, we assessed changes in cellular metabolism after exposure to lipopolysaccharide (LPS), with cytokine ELISAs and immunoblotting being used to measure inflammatory responses. Results indicate temporally distinct metabolic adaptations to pro-inflammatory stimulation in astrocytes: 3 hr LPS treatment increased glycolysis but did not alter mitochondrial metabolism, while following 24 hr of LPS treatment we observed increased oxidative phosphorylation, and decreased glycolytic capacity and glucose uptake, partly due to reduced glucose transporter 1 expression. Inhibition of NF-κB signaling with the IKK-beta inhibitor TPCA-1 prevented the LPS induced changes to glycolysis and oxidative phosphorylation. Furthermore, TPCA-1 treatment altered both glycolysis and oxidative phosphorylation independently from inflammatory stimulation, indicating a role for NF-κB signaling in regulation of basal metabolism in astrocytes. Inhibition of glycolysis with 2-deoxyglucose significantly attenuated LPS-induced cytokine release and NF-κB phosphorylation, indicating that intact glycolysis is required for the full inflammatory response to LPS. Together our data indicate that astrocytes display immunometabolic responses to acute LPS stimulation which may represent a potential therapeutic target for neuroinflammatory disorders.

Graphical abstract, Highlights • Glia direct respond to changes in nutrients and hormones regulating energy balance. • Microglia and astrocytes are implicated in the pathophysiology of obesity and diabetes. • Obesity causes gliosis: reversible changes in glia structure and function. • In metabolic disease inflammation-driven changes in glial likely regulates neurons. • Glial regulation of neural circuits controlling energy balance requires more study., Chronic low-grade inflammation is a feature of the pathophysiology of obesity and diabetes in the CNS as well as peripheral tissues. Glial cells are critical mediators of the response to inflammation in the brain. Key features of glia include their metabolic flexibility, sensitivity to changes in the CNS microenvironment, and ability to rapidly adapt their function accordingly. They are specialised cells which cooperate to promote and preserve neuronal health, playing important roles in regulating the activity of neuronal networks across the brain during different life stages. Increasing evidence points to a role of glia, most notably astrocytes and microglia, in the systemic regulation of energy and glucose homeostasis in the course of normal physiological control and during disease. Inflammation is an energetically expensive process that requires adaptive changes in cellular metabolism and, in turn, metabolic intermediates can also have immunomodulatory actions. Such “immunometabolic” changes in peripheral immune cells have been implicated in contributing to disease pathology in obesity and diabetes. This review will discuss the evidence for a role of immunometabolic changes in glial cells in the systemic regulation of energy and glucose homeostasis, and how this changes in the context of obesity and diabetes.

Hypoglycemia is a major barrier to good glucose control in type 1 diabetes and frequent exposure to hypoglycemia can impair awareness to subsequent bouts of hypoglycemia. The neural changes that occur to reduce a person’s awareness of hypoglycemia are poorly defined. Moreover, the molecular mechanisms by which glial cells contribute to hypoglycemia sensing and glucose counterregulation require further investigation. To test whether glia, specifically astrocytes, could detect changes in glucose, we utilized human primary astrocytes (HPA) and U373 astrocytoma cells and exposed them to recurrent low glucose (RLG) in vitro. This allowed measurement, with high specificity and sensitivity, of changes in cellular metabolism following RLG. We report that the AMP-activated protein kinase (AMPK) is activated over a pathophysiologically-relevant glucose concentration range. We observed an increased dependency on fatty acid oxidation for basal mitochondrial metabolism and hallmarks of mitochondrial stress including increased proton leak and reduced coupling efficiency. Relative to glucose availability, lactate release increased during low glucose but this was not modified by RLG, nor were glucose uptake or glycogen levels. Taken together, these data indicate that astrocyte mitochondria are dysfunctional following recurrent low glucose exposure, which could have implications for hypoglycemia glucose counterregulation and/or hypoglycemia awareness.

A class of glial cell, astrocytes, is highly abundant in the central nervous system (CNS). In addition to maintaining tissue homeostasis, astrocytes regulate neuronal communication and synaptic plasticity. There is an ever-increasing appreciation that astrocytes are involved in the regulation of physiology and behaviour in normal and pathological states, including within neuroendocrine systems. Indeed, astrocytes are direct targets of hormone action in the CNS, via receptors expressed on their surface, and are also a source of regulatory neuropeptides, neurotransmitters and gliotransmitters. Furthermore, as part of the neurovascular unit, astrocytes can regulate hormone entry into the CNS. This review is intended to provide an overview of how astrocytes are impacted by and contribute to the regulation of a diverse range of neuroendocrine systems: energy homeostasis and metabolism, reproduction, fluid homeostasis, the stress response and circadian rhythms.

To test the hypothesis that, given the role of AMP-activated protein kinase (AMPK) in regulating intracellular ATP levels, AMPK may alter ATP release from astrocytes, the main sources of extracellular ATP (eATP) within the brain.Measurements of ATP release were made from human U373 astrocytoma cells, primary mouse hypothalamic (HTAS) and cortical astrocytes (CRTAS) and wild-type and AMPK α1/α2 null mouse embryonic fibroblasts (MEFs). Cells were treated with drugs known to modulate AMPK activity: A-769662, AICAR and metformin, for up to 3 hours. Intracellular calcium was measured using Fluo4 and Fura-2 calcium-sensitive fluorescent dyes.In U373 cells, A-769662 (100 μM) increased AMPK phosphorylation, whereas AICAR and metformin (1 mM) induced a modest increase or had no effect, respectively. Only A-769662 increased eATP levels, and this was partially blocked by AMPK inhibitor Compound C. A-769662-induced increases in eATP were preserved in AMPK α1/α2 null MEF cells. A-769662 increased intracellular calcium in U373, HTAS and CRTAS cells and chelation of intracellular calcium using BAPTA-AM reduced A-769662-induced eATP levels. A-769662 also increased ATP release from a number of other central and peripheral endocrine cell types.AMPK is required to maintain basal eATP levels but is not required for A-769662-induced increases in eATP. A-769662 (50 μM) enhanced intracellular calcium levels leading to ATP release in an AMPK and purinergic receptor independent pathway.