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Fructose 1,6-bisphosphatase 2 (Fbp2) is a regulatory enzyme of gluco- and glyconeogenesis which, in the course of evolution, acquired non-catalytic functions. Fbp2 promotes cell survival during calcium stress, regulates glycolysis via inhibition of Hif-1α activity, and is indispensable for the formation of long-term potentiation in hippocampus. In hippocampal astrocytes, the amount of Fbp2 protein is reduced by signals delivered in neuronal extracellular vesicles (NEVs) through an unknown mechanism. The physiological role of Fbp2 (determined by its subcellular localization/interactions) depends on its oligomeric state and thus, we asked whether the cargo of NEVs is sufficient to change also the ratio of Fbp2 dimer/tetramer and, consequently, influence astrocyte basal metabolism. We found that the NEVs cargo reduced the Fbp2 mRNA level, stimulated the enzyme degradation and affected the cellular titers of different oligomeric forms of Fbp2. This was accompanied with increased glucose uptake and lactate release by astrocytes. Our results revealed that neuronal signals delivered to astrocytes in NEVs provide the necessary balance between enzymatic and non-enzymatic functions of Fbp2, influencing not only its amount but also subcellular localization. This may allow for the metabolic adjustments and ensure protection of mitochondrial membrane potential during the neuronal activity-related increase in astrocytic [Ca 2+ ]., (© 2024. The Author(s).)

Low-density lipoprotein receptor-related protein-1 (LRP1) is an endocytic/signaling cell-surface receptor that regulates diverse cellular functions, including cell survival, differentiation, and proliferation. LRP1 has been previously implicated in the pathogenesis of neurodegenerative disorders, but there are inconsistencies in its functions. Therefore, whether and how LRP1 maintains brain homeostasis remains to be clarified. Here, we report that astrocytic LRP1 promotes astrocyte-to-neuron mitochondria transfer by reducing lactate production and ADP-ribosylation factor 1 (ARF1) lactylation. In astrocytes, LRP1 suppressed glucose uptake, glycolysis, and lactate production, leading to reduced lactylation of ARF1. Suppression of astrocytic LRP1 reduced mitochondria transfer into damaged neurons and worsened ischemia-reperfusion injury in a mouse model of ischemic stroke. Furthermore, we examined lactate levels in human patients with stroke. Cerebrospinal fluid (CSF) lactate was elevated in stroke patients and inversely correlated with astrocytic mitochondria. These findings reveal a protective role of LRP1 in brain ischemic stroke by enabling mitochondria-mediated astrocyte-neuron crosstalk., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Alcohol abuse induces various neurological disorders including motor learning deficits, possibly by affecting neuronal and astrocytic activity. Physical exercise is one effective approach to remediate synaptic loss and motor deficits as shown by our previous works. In this study, we unrevealed the role of exercise training in the recovery of cortical neuronal and astrocytic functions. Using a chronic alcohol injection mouse model, we found the hyperreactivity of astrocytes along with dendritic spine loss plus lower neuronal activity in the primary motor cortex. Persistent treadmill exercise training, on the other hand, improved neural spine formation and inhibited reactive astrocytes, alleviating motor learning deficits induced by alcohol exposure. These data collectively support the potency of endurance exercise in the rehabilitation of motor functions under alcohol abuse., (© 2024. The Author(s).)

Direct neuronal reprogramming is a promising approach to replace neurons lost due to disease via the conversion of endogenous glia reacting to brain injury into neurons. However, it is essential to demonstrate that the newly generated neurons originate from glial cells and/or show that they are not pre-existing endogenous neurons. Here, we use controls for both requirements while comparing two viral vector systems (Mo-MLVs and AAVs) for the expression of the same neurogenic factor, the phosphorylation-resistant form of Neurogenin2. Our results show that Mo-MLVs targeting proliferating glial cells after traumatic brain injury reliably convert astrocytes into neurons, as assessed by genetic fate mapping of astrocytes. Conversely, expressing the same neurogenic factor in a flexed AAV system results in artefactual labelling of endogenous neurons fatemapped by birthdating in development that are negative for the genetic fate mapping marker induced in astrocytes. These results are further corroborated by chronic live in vivo imaging. Taken together, the phosphorylation-resistant form of Neurogenin2 is more efficient in reprogramming reactive glia into neurons than its wildtype counterpart in vivo using retroviral vectors (Mo-MLVs) targeting proliferating glia. Conversely, AAV-mediated expression generates artefacts and is not sufficient to achieve fate conversion.

The central control of energy homeostasis is a regulatory axis that involves the sensing of nutrients, signaling molecules, adipokines, and neuropeptides by neurons in the metabolic centers of the hypothalamus. However, non-neuronal glial cells are also abundant in the hypothalamus and recent findings have underscored the importance of the metabolic crosstalk and horizontal lipid flux between glia and neurons to the downstream regulation of systemic metabolism. New transgenic models and high-resolution analyses of glial phenotype and function have revealed that glia sit at the nexus between lipid metabolism and neural function, and may markedly impact the brain's response to dietary lipids or the supply of brain-derived lipids. Glia comprise the main cellular compartment involved in lipid synthesis, lipoprotein production, and lipid processing in the brain. In brief, tanycytes provide an interface between peripheral lipids and neurons, astrocytes produce lipoproteins that transport lipids to neurons and other glia, oligodendrocytes use brain-derived and dietary lipids to myelinate axons and influence neuronal function, while microglia can remove unwanted lipids in the brain and contribute to lipid re-utilization through cholesterol efflux. Here, we review recent findings regarding glial-lipid transport and highlight the specific molecular factors necessary for lipid processing in the brain, and how dysregulation of glial-neuronal metabolic crosstalk contributes to imbalanced energy homeostasis. Furthering our understanding of glial lipid metabolism will guide the design of future studies that target horizontal lipid processing in the brain to ameliorate the risk of developing obesity and metabolic disease., Competing Interests: Declaration of competing interest The Authors have no conflict of interest., (Copyright © 2022 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Background: Mitochondrial (mt) heteroplasmy can cause adverse biological consequences when deleterious mtDNA mutations accumulate disrupting "normal" mt-driven processes and cellular functions. To investigate the heteroplasmy of such mtDNA changes, we developed a moderate throughput mt isolation procedure to quantify the mt single-nucleotide variant (SNV) landscape in individual mouse neurons and astrocytes. In this study, we amplified mt-genomes from 1645 single mitochondria isolated from mouse single astrocytes and neurons to (1) determine the distribution and proportion of mt-SNVs as well as mutation pattern in specific target regions across the mt-genome, (2) assess differences in mtDNA SNVs between neurons and astrocytes, and (3) study co-segregation of variants in the mouse mtDNA., Results: (1) The data show that specific sites of the mt-genome are permissive to SNV presentation while others appear to be under stringent purifying selection. Nested hierarchical analysis at the levels of mitochondrion, cell, and mouse reveals distinct patterns of inter- and intra-cellular variation for mt-SNVs at different sites. (2) Further, differences in the SNV incidence were observed between mouse neurons and astrocytes for two mt-SNV 9027:G > A and 9419:C > T showing variation in the mutational propensity between these cell types. Purifying selection was observed in neurons as shown by the Ka/Ks statistic, suggesting that neurons are under stronger evolutionary constraint as compared to astrocytes. (3) Intriguingly, these data show strong linkage between the SNV sites at nucleotide positions 9027 and 9461., Conclusions: This study suggests that segregation as well as clonal expansion of mt-SNVs is specific to individual genomic loci, which is important foundational data in understanding of heteroplasmy and disease thresholds for mutation of pathogenic variants., (© 2024. The Author(s).)

Astroglial networks closely interact with neuronal populations, but their functional contribution to neuronal representation of sensory information remains unexplored. The superior colliculus (SC) integrates multi-sensory information by generating distinct spatial patterns of neuronal functional responses to specific sensory stimulation. Here, we report that astrocytes from the mouse SC form extensive networks in the retinorecipient layer compared to visual cortex. This strong astroglial connectivity relies on high expression of gap-junction proteins. Genetic disruption of this connectivity functionally impairs SC retinotopic and orientation preference responses. These alterations are region specific, absent in primary visual cortex, and associated at the circuit level with a specific impairment of collicular neurons synaptic transmission. This has implications for SC-related visually induced innate behavior, as disrupting astroglial networks impairs light-evoked temporary arrest. Our results indicate that astroglial networks shape synaptic circuit activity underlying SC functional visual responses and play a crucial role in integrating visual cues to drive sensory-motor behavior., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

In animal models of epilepsy, cranial surgery is often required to implant electrodes for electroencephalography (EEG) recording. However, electrode implants can lead to the activation of glial cells and interfere with physiological neuronal activity. In this study, we evaluated the impact of epidural electrode implants in the pilocarpine mouse model of temporal lobe epilepsy. Brain neuroinflammation was assessed 1 and 3 weeks after surgery by cytokines quantification, immunohistochemistry, and western blotting. Moreover, we investigated the effect of pilocarpine, administered two weeks after surgery, on mice mortality rate. The reported results indicate that implanted mice suffer from neuroinflammation, characterized by an early release of pro-inflammatory cytokines, microglia activation, and subsequent astrogliosis, which persists after three weeks. Notably, mice subjected to electrode implants displayed a higher mortality rate following pilocarpine injection 2 weeks after the surgery. Moreover, the analysis of EEGs recorded from implanted mice revealed a high number of single spikes, indicating a possible increased susceptibility to seizures. In conclusion, epidural electrode implant in mice promotes neuroinflammation that could lower the seizure thresholds to pilocarpine and increase the death rate. An improved protocol considering the persistent neuroinflammation induced by electrode implants will address refinement and reduction, two of the 3Rs principles for the ethical use of animals in scientific research.

Background: Since the introduction of combination antiretroviral therapy (cART) the brain has become an important human immunodeficiency virus (HIV) reservoir due to the relatively low penetration of many drugs utilized in cART into the central nervous system (CNS). Given the inherent limitations of directly assessing acute HIV infection in the brains of people living with HIV (PLWH), animal models, such as humanized mouse models, offer the most effective means of studying the effects of different viral strains and their impact on HIV infection in the CNS. To evaluate CNS pathology during HIV-1 infection in the humanized bone marrow/liver/thymus (BLT) mouse model, a histological analysis was conducted on five CNS regions, including the frontal cortex, hippocampus, striatum, cerebellum, and spinal cord, to delineate the neuronal (MAP2ab, NeuN) and neuroinflammatory (GFAP, Iba-1) changes induced by two viral strains after 2 weeks and 8 weeks post-infection., Results: Findings reveal HIV-infected human cells in the brain of HIV-infected BLT mice, demonstrating HIV neuroinvasion. Further, both viral strains, HIV-1 JR-CSF and HIV-1 CH040 , induced neuronal injury and astrogliosis across all CNS regions following HIV infection at both time points, as demonstrated by decreases in MAP2ab and increases in GFAP fluorescence signal, respectively. Importantly, infection with HIV-1 JR-CSF had more prominent effects on neuronal health in specific CNS regions compared to HIV-1 CH040 infection, with decreasing number of NeuN + neurons, specifically in the frontal cortex. On the other hand, infection with HIV-1 CH040 demonstrated more prominent effects on neuroinflammation, assessed by an increase in GFAP signal and/or an increase in number of Iba-1 + microglia, across CNS regions., Conclusion: These findings demonstrate that CNS pathology is widespread during acute HIV infection. However, neuronal loss and the magnitude of neuroinflammation in the CNS is strain dependent indicating that strains of HIV cause differential CNS pathologies., (© 2024. The Author(s).)

Central neurodegenerative disorders (e.g. Alzheimer's disease (AD) and Parkinson's disease (PD)) are tightly associated with extensive neuron loss. Current therapeutic interventions merely mitigate the symptoms of these diseases, falling short of addressing the fundamental issue of neuron loss. Cell reprogramming, involving the transition of a cell from one gene expression profile to another, has made significant strides in the conversion between diverse somatic cell types. This advancement has been facilitated by gene editing techniques or the synergistic application of small molecules, enabling the conversion of glial cells into functional neurons. Despite this progress, the potential for in situ reprogramming of astrocytes in treating neurodegenerative disorders faces challenges such as immune rejection and genotoxicity. A novel avenue emerges through chemical reprogramming of astrocytes utilizing small molecules, circumventing genotoxic effects and unlocking substantial clinical utility. Recent studies have successfully demonstrated the in situ conversion of astrocytes into neurons using small molecules. Nonetheless, these findings have sparked debates, encompassing queries regarding the origin of newborn neurons, pivotal molecular targets, and alterations in metabolic pathways. This review succinctly delineates the background of astrocytes reprogramming, meticulously surveys the principal classes of small molecule combinations employed thus far, and examines the complex signaling pathways they activate. Finally, this article delves into the potential vistas awaiting exploration in the realm of astrocytes chemical reprogramming, heralding a promising future for advancing our understanding and treatment of neurodegenerative diseases., Competing Interests: Declaration of Competing interest All authors declare that there are no conflicts of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cholesterol is crucial for the proper functioning of eukaryotic cells, especially neurons, which rely on cholesterol to maintain their complex structure and facilitate synaptic transmission. However, brain cells are isolated from peripheral cholesterol by the blood-brain barrier and mature neurons primarily uptake the cholesterol synthesized by astrocytes for proper function. This study aimed to investigate the effect of aging on cholesterol trafficking in astrocytes and its delivery to neurons. We found that aged astrocytes accumulated high levels of cholesterol in the lysosomal compartment, and this cholesterol buildup can be attributed to the simultaneous occurrence of two events: decreased levels of the ABCA1 transporter, which impairs ApoE-cholesterol export from astrocytes, and reduced expression of NPC1, which hinders cholesterol release from lysosomes. We show that these two events are accompanied by increased microR-33 in aged astrocytes, which targets ABCA1 and NPC1. In addition, we demonstrate that the microR-33 increase is triggered by oxidative stress, one of the hallmarks of aging. By coculture experiments, we show that cholesterol accumulation in astrocytes impairs the cholesterol delivery from astrocytes to neurons. Remarkably, we found that this altered transport of cholesterol could be alleviated through treatment with endocannabinoids as well as cannabidiol or CBD. Finally, according to data demonstrating that aged astrocytes develop an A1 phenotype, we found that cholesterol buildup is also observed in reactive C3+ astrocytes. Given that reduced neuronal cholesterol affects synaptic plasticity, the ability of cannabinoids to restore cholesterol transport from aged astrocytes to neurons holds significant implications in aging and inflammation., (© 2024 Wiley Periodicals LLC.)

Background: Astrocytes are the most abundant cell type of the central nervous system and are fundamentally involved in homeostasis, neuroprotection, and synaptic plasticity. This regulatory function of astrocytes on their neighboring cells in the healthy brain is subject of current research. In the ischemic brain we assume disease specific differences in astrocytic acting. The renin-angiotensin-aldosterone system regulates arterial blood pressure through endothelial cells and perivascular musculature. Moreover, astrocytes express angiotensin II type 1 and 2 receptors. However, their role in astrocytic function has not yet been fully elucidated. We hypothesized that the angiotensin II receptors impact astrocyte function as revealed in an in vitro system mimicking cerebral ischemia. Astrocytes derived from neonatal wistar rats were exposed to telmisartan (angiotensin II type 1 receptor-blocker) or PD123319 (angiotensin II type 2 receptor-blocker) under normal conditions (control) or deprivation from oxygen and glucose. Conditioned medium (CM) of astrocytes was harvested to elucidate astrocyte-mediated indirect effects on microglia and cortical neurons., Result: The blockade of angiotensin II type 1 receptor by telmisartan increased the survival of astrocytes during ischemic conditions in vitro without affecting their proliferation rate or disturbing their expression of S100A10, a marker of activation. The inhibition of the angiotensin II type 2 receptor pathway by PD123319 resulted in both increased expression of S100A10 and proliferation rate. The CM of telmisartan-treated astrocytes reduced the expression of pro-inflammatory mediators with simultaneous increase of anti-inflammatory markers in microglia. Increased neuronal activity was observed after treatment of neurons with CM of telmisartan- as well as PD123319-stimulated astrocytes., Conclusion: Data show that angiotensin II receptors have functional relevance for astrocytes that differs in healthy and ischemic conditions and effects surrounding microglia and neuronal activity via secretory signals. Above that, this work emphasizes the strong interference of the different cells in the CNS and that targeting astrocytes might serve as a therapeutic strategy to influence the acting of glia-neuronal network in de- and regenerative context., (© 2024. The Author(s).)

Astrocytes are in constant communication with neurons during the establishment and maturation of functional networks in the developing brain. Astrocytes release extracellular vesicles (EVs) containing microRNA (miRNA) cargo that regulates transcript stability in recipient cells. Astrocyte released factors are thought to be involved in neurodevelopmental disorders. Healthy astrocytes partially rescue Rett Syndrome (RTT) neuron function. EVs isolated from stem cell progeny also correct aspects of RTT. EVs cross the blood-brain barrier (BBB) and their cargo is found in peripheral blood which may allow non-invasive detection of EV cargo as biomarkers produced by healthy astrocytes. Here we characterize miRNA cargo and sequence motifs in healthy human astrocyte derived EVs (ADEVs). First, human induced Pluripotent Stem Cells (iPSC) were differentiated into Neural Progenitor Cells (NPCs) and subsequently into astrocytes using a rapid differentiation protocol. iPSC derived astrocytes expressed specific markers, displayed intracellular calcium transients and secreted ADEVs. miRNAs were identified by RNA-Seq on astrocytes and ADEVs and target gene pathway analysis detected brain and immune related terms. The miRNA profile was consistent with astrocyte identity, and included approximately 80 miRNAs found in astrocytes that were relatively depleted in ADEVs suggestive of passive loading. About 120 miRNAs were relatively enriched in ADEVs and motif analysis discovered binding sites for RNA binding proteins FUS, SRSF7 and CELF5. miR-483-5p was the most significantly enriched in ADEVs. This miRNA regulates MECP2 expression in neurons and has been found differentially expressed in blood samples from RTT patients. Our results identify potential miRNA biomarkers selectively sorted into ADEVs and implicate RNA binding protein sequence dependent mechanisms for miRNA cargo loading., Competing Interests: Declaration of competing interest The authors declare they have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Introduction: Preclinical research supports the benefits of pharmaceutical cannabis-based extracts for treating different medical conditions (e.g., epilepsy); however, their neuroprotective potential has not been widely investigated. Materials and Methods: Using primary cultures of cerebellar granule cells, we evaluated the neuroprotective activity of Epifractan (EPI), a cannabis-based medicinal extract containing a high level of cannabidiol (CBD), components like terpenoids and flavonoids, trace levels of Δ9-tetrahydrocannabinol, and the acid form of CBD. We determined the ability of EPI to counteract the rotenone-induced neurotoxicity by analyzing cell viability and morphology of neurons and astrocytes by immunocytochemical assays. The effect of EPI was compared with XALEX, a plant-derived and highly purified CBD formulation (XAL), and pure CBD crystals (CBD). Results: The results revealed that EPI induced a significant reduction in the rotenone-induced neurotoxicity in a wide range of concentrations without causing neurotoxicity per se . EPI showed a similar effect to XAL suggesting that no additive or synergistic interactions between individual substances present in EPI occurred. In contrast, CBD did show a different profile to EPI and XAL because a neurotoxic effect per se was observed at higher concentrations assayed. Medium-chain triglyceride oil used in EPI formulation could explain this difference. Conclusion: Our data support a neuroprotective effect of EPI that may provide neuroprotection in different neurodegenerative processes. The results highlight the role of CBD as the active component of EPI but also support the need for an appropriate formulation to dilute pharmaceutical cannabis-based products that could be critical to avoid neurotoxicity at very high doses.

In recent decades, there has been a dramatic rise in the rates of children being born after in utero exposure to drugs of abuse, particularly opioids. Opioids have been shown to have detrimental effects on neurons and glia in the central nervous system (CNS), but the impact of prenatal opioid exposure (POE) on still-developing synaptic circuitry is largely unknown. Astrocytes exert a powerful influence on synaptic development, secreting factors to either promote or inhibit synapse formation and neuronal maturation in the developing CNS. Here, we investigated the effects of the partial µ-opioid receptor agonist buprenorphine on astrocyte synaptogenic signaling and morphological development in cortical cell culture. Acute buprenorphine treatment had no effect on the excitatory synapse number in astrocyte-free neuron cultures. In conditions where neurons shared culture media with astrocytes, buprenorphine attenuated the synaptogenic capabilities of astrocyte-secreted factors. Neurons cultured from drug-naïve mice showed no change in synapses when treated with factors secreted by astrocytes from POE mice. However, this same treatment was synaptogenic when applied to neurons from POE mice, indicating a complex neuroadaptive response in the event of impaired astrocyte signaling. In addition to promoting morphological and connectivity changes in neurons, POE exerted a strong influence on astrocyte development, disrupting their structural maturation and promoting the accumulation of lipid droplets (LDs), suggestive of a maladaptive stress response in the developing CNS.

The transmembrane receptor for advanced glycation end products (RAGE) is a signaling receptor for many damage- and pathogen-associated molecules. Activation of RAGE is associated with inflammation and an increase in reactive oxygen species (ROS) production. Although several sources of ROS have been previously suggested, how RAGE induces ROS production is still unclear, considering the multiple targets of pathogen-associated molecules. Here, using acute brain slices and primary co-culture of cortical neurons and astrocytes, we investigated the effects of a range of synthetic peptides corresponding to the fragments of the RAGE V-domain on redox signaling. We found that the synthetic fragment (60-76) of the RAGE V-domain induces activation of ROS production in astrocytes and neurons from the primary co-culture and acute brain slices. This effect occurred through activation of RAGE and could be blocked by a RAGE inhibitor. Activation of RAGE by the synthetic fragment stimulates ROS production in NADPH oxidase (NOX). This RAGE-induced NOX activation produced only minor decreases in glutathione levels and increased the rate of lipid peroxidation, although it also reduced basal and β-amyloid induced cell death in neurons and astrocytes. Thus, specific activation of RAGE induces redox signaling through NOX, which can be a part of a cell protective mechanism., (© 2024 Federation of European Biochemical Societies.)

Innate lymphocytes, including microglial cells, astrocytes, and oligodendrocytes, play a crucial role in initiating neuroinflammatory reactions inside the central nervous system (CNS). The prime focus of this paper is on the involvement and interplay of neurons and glial cells in neurological disorders such as Alzheimer's Disease (AD), Autism Spectrum Disorder (ASD), epilepsy, and multiple sclerosis (MS). In this review, we explore the specific contributions of microglia and astrocytes and analyzes multiple pathways implicated in neuroinflammation and disturbances in excitatory and inhibitory processes. Firstly, we elucidate the mechanisms through which toxic protein accumulation in AD results in synaptic dysfunction and deregulation of the immune system and examines the roles of microglia, astrocytes, and hereditary factors in the pathogenesis of the disease. Secondly, we focus on ASD and the involvement of glial cells in the development of the nervous system and the formation of connections between neurons and investigates the genetic connections associated with these processes. Lastly, we also address the participation of glial cells in epilepsy and MS, providing insights into their pivotal functions in both conditions. We also tried to give an overview of seven different pathways like toll-like receptor signalling pathway, MyD88-dependent and independent pathway, etc and its relevance in the context with these neurological disorders. In this review, we also explore the role of activated glial cells in AD, ASD, epilepsy, and MS which lead to neuroinflammation. Even we focus on excitatory and inhibitory imbalance in all four neurological disorders as imbalance affect the proper functioning of neuronal circuits. Finally, this review concludes that there is necessity for additional investigation on glial cells and their involvement in neurological illnesses., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

Chronic neuroinflammation has been implicated in neurodegenerative disease pathogenesis. A key feature of neuroinflammation is neuronal loss and glial activation, including microglia and astrocytes. 4R-cembranoid (4R) is a natural compound that inhibits hippocampal pro-inflammatory cytokines and increases memory function in mice. We used the lipopolysaccharide (LPS) injection model to study the effect of 4R on neuronal density and microglia and astrocyte activation. C57BL/6J wild-type mice were injected with LPS (5 mg/kg) and 2 h later received either 4R (6 mg/kg) or vehicle. Mice were sacrificed after 72 h for analysis of brain pathology. Confocal images of brain sections immunostained for microglial, astrocyte, and neuronal markers were used to quantify cellular hippocampal phenotypes and neurons. Hippocampal lysates were used to measure the expression levels of neuronal nuclear protein (NeuN), inducible nitrous oxide synthase (iNOS), arginase-1, thrombospondin-1 (THBS1), glial cell-derived neurotrophic factor (GDNF), and orosomucoid-2 (ORM2) by western blot. iNOS and arginase-1 are widely used protein markers of pro- and anti-inflammatory microglia, respectively. GDNF promotes neuronal survival, and ORM2 and THBS1 are astrocytic proteins that regulate synaptic plasticity and inhibit microglial activation. 4R administration significantly reduced neuronal loss and the number of pro-inflammatory microglia 72 h after LPS injection. It also decreased the expression of the pro-inflammatory protein iNOS while increasing arginase-1 expression, supporting its anti-inflammatory role. The protein expression of THBS1, GDNF, and ORM2 was increased by 4R. Our data show that 4R preserves the integrity of hippocampal neurons against LPS-induced neuroinflammation in mice., (© 2024 The Authors. Journal of Neuroscience Research published by Wiley Periodicals LLC.)

Botulinum neurotoxin serotype A (BoNT/A) is widely used in therapeutics and cosmetics. The effects of multi-dosed BoNT/A treatment are well documented on the peripheral nervous system (PNS), but much less is known on the central nervous system (CNS). Here, the mechanism of multi-dosed BoNT/A leading to CNS neurodegeneration is explored by using the 3D human neuron-glia model. BoNT/A treatment reduces acetylcholine, triggers astrocytic transforming growth factor beta, and upregulates C1q, C3, and C5 expression, inducing microglial proinflammation. The disintegration of the neuronal microtubules is escorted by microglial nitric oxide, interleukin 1β, tumor necrosis factor α, and interleukin 8. The microglial proinflammation eventually causes synaptic impairment, phosphorylated tau (pTau) aggregation, and the loss of the BoNT/A-treated neurons. Taking a more holistic approach, the model will allow to assess therapeutics for the CNS neurodegeneration under the prolonged use of BoNT/A., (© 2024 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

The central nervous system (CNS) comprises diverse and morphologically complex cells. To understand the molecular basis of their physiology, it is crucial to assess proteins expressed within intact cells. Commonly used methods utilize cell dissociation and sorting to isolate specific cell types such as neurons and astrocytes, the major CNS cells. Proteins purified from isolated cells are identified by mass spectrometry-based proteomics. However, dissociation and cell-sorting methods lead to near total loss of cellular morphology, thereby losing proteins from key relevant subcompartments such as processes, end feet, dendrites and axons. Here we provide a systematic protocol for cell- and subcompartment-specific labeling and identification of proteins found within intact astrocytes and neurons in vivo. This protocol utilizes the proximity-dependent biotinylation system BioID2, selectively expressed in either astrocytes or neurons, to label proximal proteins in a cell-specific manner. BioID2 is targeted genetically to assess the subproteomes of subcellular compartments such as the plasma membrane and sites of cell-cell contacts. We describe in detail the expression methods (variable timing), stereotaxic surgeries for expression (1-2 d and then 3 weeks), in vivo protein labeling (7 d), protein isolation (2-3 d), protein identification methods (2-3 d) and data analysis (1 week). The protocol can be applied to any area of the CNS in mouse models of physiological processes and for disease-related research., (© 2023. Springer Nature Limited.)

Single-cell RNA sequencing (scRNA-seq) has been widely applied in neuroscience research, enabling the investigation of cellular heterogeneity at the transcriptional level, the characterization of rare cell types, and the detailed analysis of the stochastic nature of gene expression. Isolation of single nerve cells in good health, especially from the adult rodent brain, is the most difficult and critical process for scRNA-seq. Here, we describe methods to optimize protease digestion of brain slices, which enable yield of millions of cells in good health from the adult brain., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Spinal cord injuries cause irreversible loss of sensory and motor functions. In mammals, intrinsic and extrinsic inhibitions of neuronal regeneration obstruct neural repair after spinal cord injury. Although astrocytes have been involved in a growing list of vital homeostatic functions in the nervous system, their roles after injury have fascinated and puzzled scientists for decades. Astrocytes undergo long-lasting morphological and functional changes after injury, referred to as reactive astrogliosis. Although reactive astrogliosis is required to contain spinal cord lesions and restore the blood-spinal cord barrier, reactive astrocytes have detrimental effects that inhibit neuronal repair and remyelination. Intriguingly, elevated regenerative capacity is preserved in some non-mammalian vertebrates, where astrocyte-like glial cells display exclusively pro-regenerative effects after injury. A detailed molecular and phenotypic catalog of the continuum of astrocyte reactivity states is an essential first step toward the development of glial cell manipulations for spinal cord repair., (© 2024. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Proper functioning of the central nervous system depends on various tightly regulated phenomena, among which astrocyte-neuron interactions are of critical importance. Various studies across the species have highlighted the diverse yet crucial roles of astrocytes in regulating the nervous system development and functions. In simpler organisms like worms or insects, astrocyte-like cells govern basic functions such as structural support to neurons or regulation of extracellular ions. As the species complexity increases, so does the functional and morphological complexity of astrocytes. For example, in fish and amphibians, these cells are involved in synaptic development and ion homeostasis, while in reptiles and birds, astrocytes regulate synaptic transmission and plasticity and are reported to be involved in complex behaviors. Other species like those belonging to mammals and, in particular, primates have a heterogeneous population of astrocytes, exhibiting region-specific functional properties. In primates, these cells are responsible for proper synaptic transmission, neurotransmitter release and metabolism, and higher cognitive functions like learning, memory, or information processing. This chapter highlights the well-established and somewhat conserved roles of astrocytes and astrocyte-neuron interactions across the evolution of both invertebrates and vertebrates., (© 2024. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Astrocytes are intricately involved in the activity of neural circuits; however, their basic physiology of interacting with nearby neurons is not well established. Using two-photon imaging of neurons and astrocytes during higher frequency stimulation of hippocampal CA3-CA1 Schaffer collateral (Scc) excitatory synapses, we could show that increasing levels of released glutamate accelerated local astrocytic Ca 2+ elevation. However, blockage of glutamate transporters did not abolish this astrocytic Ca 2+ response, suggesting that astrocytic Ca 2+ elevation is indirectly associated with an uptake of extracellular glutamate. However, during the astrocytic glutamate uptake, the Na + /Ca 2+ exchanger (NCX) reverse mode was activated, and mediated extracellular Ca 2+ entry, thereby triggering the internal release of Ca 2+ . In addition, extracellular Ca 2+ entry via membrane P2X receptors further facilitated astrocytic Ca 2+ elevation via ATP binding. These findings suggest a novel mechanism of activity induced Ca 2+ permeability increases of astrocytic membranes, which drives astrocytic responses during neuronal stimulation of CA3-CA1 Scc excitatory synapses., (© 2023 The Authors. GLIA published by Wiley Periodicals LLC.)

The conversion of non-neuronal cells to neurons is a promising potential strategy for the treatment of neurodegenerative diseases. Recent studies have reported that shRNA-, CasRx-, or ASO-mediated Ptbp1 suppression could reprogram resident astrocytes to neurons. However, some groups have disputed the interpretation of the data underlying the reported neuron conversion events. These controversies surrounding neuron conversion may be due to differences in the astrocyte fate-mapping systems. Here, we suppressed Ptbp1 using Cas13X and labelled astrocytes with an HA tag fused to Cas13X (Cas13X-NLS-HA). We found no astrocyte-to-neuron conversion in the mouse striatum via the HA-tagged labelling system compared with the GFAP-driven tdTomato labelling system (AAV-GFAP::tdTomato-WPRE) used in previous studies. Our findings indicate that Cas13X-mediated Ptbp1 knockdown failed to induce neuron conversion in vivo., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

This study proposes an innovative analog neuromorphic circuit design to mimic spontaneous Ca 2+ oscillations observed in astrocytes. Unlike traditional models, this approach does not rely on synaptic stimulation, suggesting that astrocytes may play a key role in generating neuronal activity. The circuit is built using transistor differential pairs to approximate the nonlinear sigmoidal biological functions, and its performance is validated through simulation and compared against mathematical models using phase diagram analysis. Results indicate a good fit between the circuit and the mathematical model. Finally, the circuit's ability to simulate the release of glutamate and ATP through spontaneous oscillations is demonstrated., (© 2023. The Author(s).)

Acute brain slices are a common and useful preparation in experimental neuroscience. A wide range of incubation chambers for brain slices exists but only a few are designed with very low volumes of the bath solution in mind. Such chambers are necessary when high-cost chemicals are to be added to the solution or when small amounts of substances released by the slice are to be collected for analysis. The principal challenge in designing a very low-volume incubation chamber is maintaining good oxygenation and flow without mechanically disturbing or damaging the slices. We designed and validated BubbleDrive, a 3D-printed incubation chamber with a minimum volume of 1.5 mL which can hold up to three coronal mouse slices from one hemisphere. It employs the carbogen gas bubbles to drive the flow circulation in a consistent and reproducible manner, and without disturbing the brain slices. The BubbleDrive design and construction were successfully validated by comparison to a conventional large-volume incubation chamber in several experimental designs involving measurements of extracellular diffusion parameters, the electrophysiology of neuronal and astrocytic networks, and the effectiveness of slice incubation with hyaluronidase enzyme., (© 2023. The Author(s).)

Emerging evidence emphasizes lactate's involvement in both physiological processes (energy metabolism, memory, etc.) and disease (traumatic brain injury, epilepsy, etc.). Furthermore, the usefulness of mathematical modeling in deciphering underlying dynamics of the brain to investigate lactate roles and mechanisms of action has been well established. Here, we analyze a novel mathematical model of brain lactate exchanges between four compartments: neurons, astrocytes, capillaries, and extracellular space. A system of four ordinary differential equations is proposed to explain interactions between these compartments. We first optimize and analyze the model's parameters under normal, resting state conditions, and then use it to simulate changes linked to elevated arterial lactate. Our results show that even though increased arterial lactate results in increased uptake by astrocytes and release to the extracellular space, it cannot strongly recover the initial drop in neuronal lactate concentration. Also, we show that the direction of lactate transport between the compartments is influenced by the maximum astrocyte production rate and the transport rate between astrocytes and extracellular space., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Brain activities are featured by spatially distributed neural clusters of coherent firings and a spontaneous slow switching of the clusters between the coherent and incoherent states. Evidences from recent in vivo experiments suggest that astrocytes, a type of glial cell regarded previously as providing only structural and metabolic supports to neurons, participate actively in brain functions by regulating the neural firing activities, yet the underlying mechanism remains unknown. Here, introducing astrocyte as a reservoir of the glutamate released from the neuron synapses, we propose the model of the complex neuron-astrocyte network, and investigate the roles of astrocytes in regulating the cluster synchronization behaviors of networked chaotic neurons. It is found that a specific set of neurons on the network are synchronized and form a cluster, while the remaining neurons are kept as desynchronized. Moreover, during the course of network evolution, the cluster is switching between the synchrony and asynchrony states in an intermittent fashion, henceforth the phenomenon of "breathing cluster." By the method of symmetry-based analysis, we conduct a theoretical investigation on the synchronizability of the cluster. It is revealed that the contents of the cluster are determined by the network symmetry, while the breathing of the cluster is attributed to the interplay between the neural network and the astrocyte. The phenomenon of breathing cluster is demonstrated in different network models, including networks with different sizes, nodal dynamics, and coupling functions. The findings shed light on the cellular mechanism of astrocytes in regulating neural activities and give insights into the state-switching of the neocortex., (© 2023 Author(s). Published under an exclusive license by AIP Publishing.)

Glial cells account for between 50% and 90% of all human brain cells, and serve a variety of important developmental, structural, and metabolic functions. Recent experimental efforts suggest that astrocytes, a type of glial cell, are also directly involved in core cognitive processes such as learning and memory. While it is well established that astrocytes and neurons are connected to one another in feedback loops across many timescales and spatial scales, there is a gap in understanding the computational role of neuron-astrocyte interactions. To help bridge this gap, we draw on recent advances in AI and astrocyte imaging technology. In particular, we show that neuron-astrocyte networks can naturally perform the core computation of a Transformer, a particularly successful type of AI architecture. In doing so, we provide a concrete, normative, and experimentally testable account of neuron-astrocyte communication. Because Transformers are so successful across a wide variety of task domains, such as language, vision, and audition, our analysis may help explain the ubiquity, flexibility, and power of the brain's neuron-astrocyte networks.

Over the last decades, lactate has emerged as important energy substrate for the brain fueling of neurons. A growing body of evidence now indicates that it is also a signaling molecule modulating neuronal excitability and activity as well as brain functions. In this review, we will briefly summarize how different cell types produce and release lactate. We will further describe different signaling mechanisms allowing lactate to fine-tune neuronal excitability and activity, and will finally discuss how these mechanisms could cooperate to modulate neuroenergetics and higher order brain functions both in physiological and pathological conditions., Competing Interests: Declaration of Competing Interest The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

A holy grail of regenerative medicine is to replenish the cells that are lost due to disease. The adult mammalian central nervous system (CNS) has, however, largely lost such a regenerative ability. An emerging strategy for the generation of new neurons is through glia-to-neuron (GtN) conversion in vivo, mainly accomplished by the regulation of fate-determining factors. When inhibited, PTBP1, a factor involved in RNA biology, was reported to induce rapid and efficient GtN conversion in multiple regions of the adult CNS. Remarkably, PTBP1 inhibition was also claimed to greatly improve behaviors of mice with neurological diseases or aging. These phenomenal claims, if confirmed, would constitute a significant advancement in regenerative medicine. Unfortunately, neither GtN conversion nor therapeutic potential via PTBP1 inhibition was validated by the results of multiple subsequent replication studies with stringent methods. Here we review these controversial studies and conclude with recommendations for examining GtN conversion in vivo and future investigations of PTBP1.

Breast milk contains constituents, such as 1,3-dioleoyl-2-palmitoylglycerol (OPO), that are beneficial for infants. Herein, we hypothesized that 2-palmitoyl glycerol (2-PG), a derivative of OPO, is advantageous to infants' development. Gamma aminobutyric acid (GABA) is a major neurotransmitter involved in neural development. Although GABA is generally known to be produced in neurons, astrocytes can also produce it in immature brains. In this study, we used expression analysis techniques to show that 2-PG upregulates the mRNA and protein expression of glutamate decarboxylases (GAD1 and GAD2) in normal human fetal-derived astrocytes. Our data suggest that 2-PG promotes GABA synthesis in astrocytes, which may contribute to brain development because GABA is involved in neural development in the developing brain. This may help to elucidate the mechanism by which breast milk affects infant brain development., (© 2023 The Authors. FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Viral tracers that enable efficient retrograde labeling of projection neurons are powerful vehicles for structural and functional dissections of the neural circuit and for the treatment of brain diseases. Currently, some recombinant adeno-associated viruses (rAAVs) based on capsid engineering are widely used for retrograde tracing, but display undesirable brain area selectivity due to inefficient retrograde transduction in certain neural connections. Here we developed an easily editable toolkit to produce high titer AAV11 and demonstrated that it exhibits potent and stringent retrograde labeling of projection neurons in adult male wild-type or Cre transgenic mice. AAV11 can function as a powerful retrograde viral tracer complementary to AAV2-retro in multiple neural connections. In combination with fiber photometry, AAV11 can be used to monitor neuronal activities in the functional network by retrograde delivering calcium-sensitive indicator under the control of a neuron-specific promoter or the Cre-lox system. Furthermore, we showed that GfaABC 1 D promoter embedding AAV11 is superior to AAV8 and AAV5 in astrocytic tropism in vivo, combined with bidirectional multi-vector axoastrocytic labeling, AAV11 can be used to study neuron-astrocyte connection. Finally, we showed that AAV11 allows for analyzing circuit connectivity difference in the brains of the Alzheimer's disease and control mice. These properties make AAV11 a promising tool for mapping and manipulating neural circuits and for gene therapy of some neurological and neurodegenerative disorders., (© 2023. The Author(s).)

Disrupted brain metabolism is a critical component of several neurodegenerative diseases. Energy metabolism of both neurons and astrocytes is closely connected to neurotransmitter recycling via the glutamate/GABA-glutamine cycle. Neurons and astrocytes hereby work in close metabolic collaboration which is essential to sustain neurotransmission. Elucidating the mechanistic involvement of altered brain metabolism in disease progression has been aided by the advance of techniques to monitor cellular metabolism, in particular by mapping metabolism of substrates containing stable isotopes, a technique known as isotope tracing. Here we review key aspects of isotope tracing including advantages, drawbacks and applications to different cerebral preparations. In addition, we narrate how isotope tracing has facilitated the discovery of central metabolic features in neurodegeneration with a focus on the metabolic cooperation between neurons and astrocytes., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to declare., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Neuronal rhythmogenesis in the spinal cord is correlated with variations in extracellular K + levels ([K + ] e ). Astrocytes play important role in [K + ] e homeostasis and compute neuronal information. Yet it is unclear how neuronal oscillations are regulated by astrocytic K + homeostasis. Here we identify the astrocytic inward-rectifying K + channel Kir4.1 (a.k.a. Kcnj10) as a key molecular player for neuronal rhythmicity in the spinal central pattern generator (CPG). By combining two-photon calcium imaging with electrophysiology, immunohistochemistry and genetic tools, we report that astrocytes display Ca 2+ transients before and during oscillations of neighboring neurons. Inhibition of astrocytic Ca 2+ transients with BAPTA decreases the barium-sensitive Kir4.1 current responsible of K + clearance. Finally, we show in mice that Kir4.1 knockdown in astrocytes progressively prevents neuronal oscillations and alters the locomotor pattern resulting in lower motor performances in challenging tasks. These data identify astroglial Kir4.1 channels as key regulators of neuronal rhythmogenesis in the CPG driving locomotion., (© 2023 The Authors. GLIA published by Wiley Periodicals LLC.)

Resolving fundamental molecular and functional processes underlying human synaptic development is crucial for understanding normal brain function as well as dysfunction in disease. Based upon increasing evidence of species-divergent features of brain cell types, coupled with emerging studies of complex human disease genetics, we developed the first automated and quantitative high-content synaptic phenotyping platform using human neurons and astrocytes. To establish the robustness of our platform, we screened the effects of 376 small molecules on presynaptic density, neurite outgrowth, and cell viability, validating six small molecules that specifically enhanced human presynaptic density in vitro. Astrocytes were essential for mediating the effects of all six small molecules, underscoring the relevance of non-cell-autonomous factors in synapse assembly and their importance in synaptic screening applications. Bromodomain and extraterminal (BET) inhibitors emerged as the most prominent hit class and global transcriptional analyses using multiple BET inhibitors confirmed upregulation of synaptic gene expression. Through these analyses, we demonstrate the robustness of our automated screening platform for identifying potent synaptic modulators, which can be further leveraged for scaled analyses of human synaptic mechanisms and drug discovery efforts., Competing Interests: MB, GR, AN, JK, DT, SS, DL, AM, KH, KK, BC, KP, AC, LB No competing interests declared, LR Reviewing editor, eLife. LLR is a founder of Elevian, Rejuveron, and Vesalius Therapeutics, a member of their scientific advisory boards and a private equity shareholder. All are interested in formulating approaches intended to treat diseases of the nervous system and other tissues. He is also on the advisory board of Alkahest, a Grifols company, focused on the plasma proteome and brain aging. None of these companies provided any financial support for the work in this paper, (© 2023, Berryer et al.)

Background: Three-dimensional (3D) human brain spheroids are instrumental to study central nervous system (CNS) development and (dys)function. Yet, in current brain spheroid models the limited variety of cell types hampers an integrated exploration of CNS (disease) mechanisms., Methods: Here we report a 5-month culture protocol that reproducibly generates H9 embryonic stem cell-derived human cortical spheroids (hCSs) with a large cell-type variety., Results: We established the presence of not only neuroectoderm-derived neural progenitor populations, mature excitatory and inhibitory neurons, astrocytes and oligodendrocyte (precursor) cells, but also mesoderm-derived microglia and endothelial cell populations in the hCSs via RNA-sequencing, qPCR, immunocytochemistry and transmission electron microscopy. Transcriptomic analysis revealed resemblance between the 5-months-old hCSs and dorsal frontal rather than inferior regions of human fetal brains of 19-26 weeks of gestational age. Pro-inflammatory stimulation of the generated hCSs induced a neuroinflammatory response, offering a proof-of-principle of the applicability of the spheroids., Conclusions: Our protocol provides a 3D human brain cell model containing a wide variety of innately developing neuroectoderm- as well as mesoderm-derived cell types, furnishing a versatile platform for comprehensive examination of intercellular CNS communication and neurological disease mechanisms., (© 2023. The Author(s).)

Aging drives cognitive decline, and mitochondrial dysfunction is a hallmark of age-induced neurodegeneration. Recently, we demonstrated that astrocytes secrete functional mitochondria (Mt), which help adjacent cells to resist damage and promote repair after neurological injuries. However, the relationship between age-dependent changes in astrocytic Mt function and cognitive decline remains poorly understood. Here, we established that aged astrocytes secret less functional Mt compared to young astrocytes. We found the aging factor C-C motif chemokine 11 (CCL11) is elevated in the hippocampus of aged mice, and that its level is reduced upon systemic administration of young Mt, in vivo. Aged mice receiving young Mt, but not aged Mt improved cognitive function and hippocampal integrity. Using a CCL11-induced aging-like model in vitro, we found that astrocytic Mt protect hippocampal neurons and enhance a regenerative environment through upregulating synaptogenesis-related gene expression and anti-oxidants that were suppressed by CCL11. Moreover, the inhibition of CCL11-specific receptor C-C chemokine receptor 3 (CCR3) boosted the expression of synaptogenesis-related genes in the cultured hippocampal neurons and restored the neurite outgrowth. This study suggests that young astrocytic Mt can preserve cognitive function in the CCL11-mediated aging brain by promoting neuronal survival and neuroplasticity in the hippocampus.

The brain is a complex organ even when viewed from a cell biological perspective. Neuronal networks are embedded in a dense milieu of diverse and specialised cell types, including several types of vascular, immune, and macroglial cells. To view each cell as a small cog in a highly complex machine is itself an oversimplification. Not only are they functionally coupled to enable the brain to operate, each cell type's functions are themselves influenced by each other, in development, maturity, and also in disease. Astrocytes are a type of macroglia that occupy a significant fraction of the human forebrain. They play a critical role in sustaining functional neuronal circuits across the lifespan through myriad homeostatic functions including the maintenance of redox balance, ionic gradients, neurotransmitter clearance, and bioenergetic support. It is becoming apparent that astrocytes' capacity to carry out these and other neurosupportive roles is not fixed, but is regulated by signals coming from the neurons themselves, both in the healthy brain but also in response to neuron-derived disease pathology. Here, we review mechanisms by which neurons control the properties of astrocytes long term in order to alter their homeostatic capacity both in development and maturity. Our working hypothesis is that these signals are designed to change and maintain the homeostatic capacity of local astrocytes to suit the needs of nearby neurons. Knowledge of the external signals that can control core aspects of a healthy astrocytic phenotype are being uncovered, raising the question as to whether this knowledge can be harnessed to promote astrocyte-mediated neurosupport in brain disorders., (© 2023 The Author(s).)

The duck is a representative and good model for studying the development and physiological mechanisms of the nervous system (NS) in waterfowl. Neurons are the basic structural and functional units of NS, but there is no detailed method for cultured duck neurons in vitro. An efficient and simple method for duck neuron culture is reported in this study. First, the sfigpecific markers (NSE and GFAP, respectively) were used to explore the timing of the development of neurons and astrocytes during the duck embryonic stage (E5-E18). The cytomorphology of tissues and cells was tracked with the microscope at different time points. The brain tissues from 10-day-old duck embryos were determined as the optimal sampling embryo age for neuron culture. Then, the brain tissue isolation method (papain digestion) and cell suspension inoculation density (7 × 10 5 cells/mL) were identified as the culture protocol to obtain target cells with high viability and high density. The purity of the cultured neurons was more than 95%. This experiment provides a supplement for the study of in vitro culture of waterfowl neurons and lays a good foundation for various subsequent studies., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Astrocytic morphological plasticity and its modulation of adjacent neuronal activity are largely determined by astrocytic volume regulation, in which glial fibrillary acidic protein (GFAP), aquaporin 4 (AQP4), and potassium channels including inwardly rectifying K + channel 4.1 (Kir4.1) are essential. However, associations of astrocyte-dominant Kir4.1 with other molecules in astrocytic volume regulation and the subsequent influence on neuronal activity remain unclear. Here, we report our study on these issues using primary cultures of rat pups' hypothalamic astrocytes and male adult rat brain slices. In astrocyte culture, hyposmotic challenge (HOC) significantly decreased GFAP monomer expression and astrocytic volume at 1.5 min and increased Kir4.1 expression and inwardly rectifying currents (IRCs) at 10 min. BaCl 2 (100 μmol/l) suppressed the HOC-increased IRCs, which was simulated by VU0134992 (2 μmol/l), a Kir4.1 blocker. Preincubation of the astrocyte culture with TGN-020 (10 μmol/l, a specific AQP4 blocker) made the HOC-increased Kir4.1 currents insignificant. In hypothalamic brain slices, HOC initially decreased and then increased the firing rate of vasopressin (VP) neurons in the supraoptic nucleus. In the presence of BaCl 2 or VU0134992, HOC-elicited rebound increase in VP neuronal activity was blocked. GFAP was molecularly associated with Kir4.1, which was increased by HOC at 20 min; this increase was blocked by BaCl 2 . These results suggest that HOC-evoked astrocytic retraction or decrease in the volume and length of its processes is associated with increased Kir4.1 activity. Kir4.1 involvement in HOC-elicited astrocytic retraction is associated with AQP4 activity and GFAP plasticity, which together determines the rebound excitation of VP neurons., (© 2022 Wiley Periodicals LLC.)

Lipids comprise an extremely heterogeneous group of compounds that perform a wide variety of biological functions. Traditional view of lipids as important structural components of the cell and compounds playing a trophic role is currently being supplemented by information on the possible participation of lipids in signaling, not only intracellular, but also intercellular. The review article discusses current data on the role of lipids and their metabolites formed in glial cells (astrocytes, oligodendrocytes, microglia) in communication of these cells with neurons. In addition to metabolic transformations of lipids in each type of glial cells, special attention is paid to the lipid signal molecules (phosphatidic acid, arachidonic acid and its metabolites, cholesterol, etc.) and the possibility of their participation in realization of synaptic plasticity, as well as in other possible mechanisms associated with neuroplasticity. All these new data can significantly expand our knowledge about the regulatory functions of lipids in neuroglial relationships.

Glial cells, consisting of astrocytes, oligodendrocyte lineage cells, and microglia, account for >50% of the total number of cells in the mammalian brain. They play key roles in the modulation of various brain activities under physiological and pathological conditions. Although the typical morphological features and characteristic functions of these cells are well described, the organization of interconnections of the different glial cell populations and their impact on the healthy and diseased brain is not completely understood. Understanding these processes remains a profound challenge. Accumulating evidence suggests that glial cells can form highly complex interconnections with each other. The astroglial network has been well described. Oligodendrocytes and microglia may also contribute to the formation of glial networks under various circumstances. In this review, we discuss the structure and function of glial networks and their pathological relevance to central nervous system diseases. We also highlight opportunities for future research on the glial connectome., (© 2022. Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences.)

Structural changes of astrocytes and their perisynaptic processes occur in response to various physiological and pathophysiological stimuli. They are thought to profoundly affect synaptic signalling and neuron-astrocyte communication. Understanding the causal relationship between astrocyte morphology changes and their functional consequences requires experimental tools to selectively manipulate astrocyte morphology. Previous studies indicate that RhoA-related signalling can play a major role in controlling astrocyte morphology, but the direct effect of increased RhoA activity has not been documented in vitro and in vivo. Therefore, we established a viral approach to manipulate astrocytic RhoA activity. We tested if and how overexpression of wild-type RhoA, of a constitutively active RhoA mutant (RhoA-CA), and of a dominant-negative RhoA variant changes the morphology of cultured astrocytes. We found that astrocytic expression of RhoA-CA induced robust cytoskeletal changes and a withdrawal of processes in cultured astrocytes. In contrast, overexpression of other RhoA variants led to more variable changes of astrocyte morphology. These induced morphology changes were reproduced in astrocytes of the hippocampus in vivo. Importantly, astrocytic overexpression of RhoA-CA did not alter the branching pattern of larger GFAP-positive processes of astrocytes. This indicates that a prolonged increase of astrocytic RhoA activity leads to a distinct morphological phenotype in vitro and in vivo, which is characterized by an isolated reduction of fine peripheral astrocyte processes in vivo. At the same time, we identified a promising experimental approach for investigating the functional consequences of astrocyte morphology changes.

Neurons in the brain build circuits by synapsing with each other, and glial cells are involved in the formation and elimination of synapses. Glial cells include microglia, astrocytes, and oligodendrocytes, each with distinctive functions supported by different gene expression patterns and morphologies, but all have been shown to regulate the number of synapses in the neuronal circuits through a common function, synaptic phagocytosis. It has also been reported that specific glial cell types phagocytose specific synapses in different brain regions and at different times, and some of the molecular mechanisms involved in each phagocytotic process have been elucidated. For example, microglia, the most frequently reported glial cell type in relation to synaptic phagocytes, are known to recognize various "eat me signals" including complement and phagocytose synapses, contributing to the refinement of neuronal circuits during development. More recently, astrocytes and oligodendrocyte precursor cells have also been shown to be involved in synaptic phagocytosis. Interestingly, there are also reports of different types of glial cells phagocytosing the same types of synapses. And in some cases, it has been suggested that different glial cell types regulate each other's synaptic phagocytosis. In this review, we will discuss the significance of synaptic phagocytosis by multiple types of glial cells by presenting recent studies on synaptic phagocytosis by glial cells.

Neuromorphic engineering is an essential science field which incorporates the basic aspects of issues together such as: physics, mathematics, electronics, etc. The primary block in the Central Nervous System (CNS) is neurons that have functional roles such as: receiving, processing, and transmitting data in the brain. This paper presents Wilson Multiplierless Neuron (WMN) model which is a modified version of the original model. This model uses power-2 based functions, Look-Up Table (LUT) approach and shifters to apply a multiplierless digital realization leads to overhead costs reduction and increases in the final system frequency. The proposed model specifically follows the original neuron model in case of spiking patterns and also dynamical pathways. To validate the proposed model in digital hardware implementation, the FPGA board (Xilinx Virtex II XC2VP30) can be used. Hardware results show the increasing in the system frequency compared with the original model and other similar papers. Numerical results demonstrate that the proposed system speed-up is 210 MHz that is higher than the original one, 85 MHz. Additionally, the overall saving in FPGA resources for the proposed model is 96.86 % that is more than the original model, 95.13 %. From case study viewpoint for CNS consideration, a network consisting of Wilson neurons, synapses, and astrocytes have been considered to test the controlling effects on LTP and LTD processes for investigating the neuronal diseases (medical approaches) such as Epilepsy.

In ischemic stroke and post-traumatic brain injury (TBI), blood-brain barrier disruption leads to leaking plasma amino acids (AA) into cerebral parenchyma. Bleeding in hemorrhagic stroke and TBI also release plasma AA. Although excitotoxic AA were extensively studied, little is known about non-excitatory AA during hypoxic injury. Hypoxia-induced synaptic depression in hippocampal slices becomes irreversible with non-excitatory AA, alongside their intracellular accumulation and increased tissue electrical resistance. Four non-excitatory AA (l-alanine, glycine, l-glutamine, l-serine: AGQS) at plasmatic concentrations were applied to slices from mice expressing EGFP in pyramidal neurons or astrocytes during normoxia or hypoxia. Two-photon imaging, light transmittance (LT) changes, and electrophysiological field recordings followed by electron microscopy in hippocampal CA1 st. radiatum were used to monitor synaptic function concurrently with cellular swelling and injury. During normoxia, AGQS-induced increase in LT was due to astroglial but not neuronal swelling. LT raise during hypoxia and AGQS manifested astroglial and neuronal swelling accompanied by a permanent loss of synaptic transmission and irreversible dendritic beading, signifying acute damage. Neuronal injury was not triggered by spreading depolarization which did not occur in our experiments. Hypoxia without AGQS did not cause cell swelling, leaving dendrites intact. Inhibition of NMDA receptors prevented neuronal damage and irreversible loss of synaptic function. Deleterious effects of AGQS during hypoxia were prevented by alanine-serine-cysteine transporters (ASCT2) and volume-regulated anion channels (VRAC) blockers. Our findings suggest that astroglial swelling induced by accumulation of non-excitatory AA and release of excitotoxins through antiporters and VRAC may exacerbate the hypoxia-induced neuronal injury., (© 2022 Wiley Periodicals LLC.)

The role of glia as active participants in brain functions has become increasingly evident. In this issue of Cell Metabolism, Herrera Moro Chao et al. reveal that astrocytes in the hypothalamic paraventricular nucleus (PVN) bidirectionally control neuronal behavior in response to metabolic cues and that this control is disrupted in obesity., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)