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Astrocytes respond and contribute to neuroinflammation by adopting inflammatory reactive states. Although recent efforts have characterized the gene expression signatures associated with these reactive states, the cell biology underlying inflammatory reactive astrocyte phenotypes remains under-explored. Here, we used CRISPR-based screening in human iPSC-derived astrocytes to identify mTOR activation a driver of cytokine-induced endolysosomal system remodeling, manifesting as alkalinization of endolysosomal compartments, decreased autophagic flux, and increased exocytosis of certain endolysosomal cargos. Through endolysosomal proteomics, we identified and focused on one such cargo-IL-32, a disease-associated pro-inflammatory cytokine not present in rodents, whose secretion mechanism is not well understood. We found that IL-32 was partially secreted in extracellular vesicles likely to be exosomes. Furthermore, we found that IL-32 was involved in the polarization of inflammatory reactive astrocyte states and was upregulated in astrocytes in multiple sclerosis lesions. We believe that our results advance our understanding of cell biological pathways underlying inflammatory reactive astrocyte phenotypes and identify potential therapeutic targets.

Central nervous system (CNS) lesions become surrounded by neuroprotective borders of newly proliferated reactive astrocytes; however, fundamental features of these cells are poorly understood. Here we show that following spinal cord injury or stroke, 90% and 10% of border-forming astrocytes derive, respectively, from proliferating local astrocytes and oligodendrocyte progenitor cells in adult mice of both sexes. Temporal transcriptome analysis, single-nucleus RNA sequencing and immunohistochemistry show that after focal CNS injury, local mature astrocytes dedifferentiate, proliferate and become transcriptionally reprogrammed to permanently altered new states, with persisting downregulation of molecules associated with astrocyte-neuron interactions and upregulation of molecules associated with wound healing, microbial defense and interactions with stromal and immune cells. These wound repair astrocytes share morphologic and transcriptional features with perimeningeal limitans astrocytes and are the predominant source of neuroprotective borders that re-establish CNS integrity around lesions by separating neural parenchyma from stromal and immune cells as occurs throughout the healthy CNS.

After traumatic brain injury, the brain extracellular matrix undergoes structural rearrangement due to changes in matrix composition, activation of proteases, and deposition of chondroitin sulfate proteoglycans by reactive astrocytes to produce the glial scar. These changes lead to a softening of the tissue, where the stiffness of the contusion core and peripheral pericontusional regions becomes softer than that of healthy tissue. Pioneering mechanotransduction studies have shown that soft substrates upregulate intermediate filament proteins in reactive astrocytes; however, many other aspects of astrocyte biology remain unclear. Here, we developed a platform for the culture of cortical astrocytes using polyacrylamide (PA) gels of varying stiffness (measured in Pascal; Pa) to mimic injury-related regions in order to investigate the effects of tissue stiffness on astrocyte reactivity and morphology. Our results show that substrate stiffness influences astrocyte phenotype; soft 300 Pa substrates led to increased GFAP immunoreactivity, proliferation, and complexity of processes. Intermediate 800 Pa substrates increased Aggrecan+, Brevican+, and Neurocan+ astrocytes. The stiffest 1 kPa substrates led to astrocytes with basal morphologies, similar to a physiological state. These results advance our understanding of astrocyte mechanotransduction processes and provide evidence of how substrates with engineered stiffness can mimic the injury microenvironment.

BackgroundThe neuronal and gliaI populations within the brain are tightly interwoven, making isolation and study of large populations of a single cell type from brain tissue a major technical challenge. Concurrently, cell-type specific extracellular vesicles (EVs) hold enormous diagnostic and therapeutic potential in neurodegenerative disorders including Alzheimer's disease (AD).New methodPostmortem AD cortical samples were thawed and gently dissociated. Following filtration, myelin and red blood cell removal, cell pellets were immunolabeled with fluorescent antibodies and analyzed by flow cytometry. The cell pellet supernatant was applied to a triple sucrose cushion for brain EV isolation.ResultsNeuronal, astrocyte and microglial cell populations were identified. Cell integrity was demonstrated using calcein AM, which is retained by cells with esterase activity and an intact membrane. For some experiments cell pellets were fixed, permeabilized, and immunolabeled for cell-specific markers. Characterization of brain small EV fractions showed the expected size, depletion of EV negative markers, and enrichment in positive and cell-type specific markers.Comparison with existing methods and conclusionsWe optimized and integrated established protocols, aiming to maximize information obtained from each human autopsy brain sample. The uniqueness of our method lies in its capability to isolate cells and EVs from a single cryopreserved brain sample. Our results not only demonstrate the feasibility of isolating specific brain cell subpopulations for RNA-seq but also validate these subpopulations at the protein level. The accelerated study of EVs from human samples is crucial for a better understanding of their contribution to neuron/glial crosstalk and disease progression.

OBJECTIVES: People with HIV (PWH) have high rates of depression and neurocognitive impairment (NCI) despite viral suppression on antiretroviral therapy (ART). Mounting evidence suggests that immunometabolic disruptions may contribute to these conditions in some PWH. We hypothesized that metabolic dysfunction in astrocytes is associated with depressive symptoms and cognitive function in PWH. METHODS: Human astrocytes were exposed to sera from PWH (n=40) with varying degrees of depressive symptomatology and cognitive function. MitoTrackerTM Deep Red FM (MT) was used to visualize mitochondrial activity and glial fibrillary acidic protein (GFAP) as an indicator of astrocyte reactivity using the high-throughput fluorescent microscopy and image analyses platform, CellInsight CX5 (CX5). The Seahorse platform was used to assess glycolytic and mitochondrial metabolism. RESULTS: More severe depression, as indexed by higher Becks Depression Inventory (BDI-II) scores, was associated with lower MT signal measures. Better cognitive function, as assessed by neuropsychiatric testing t-scores, was associated with increased MT signal measures. GFAP intensity negatively correlated with several cognitive t-scores. Age positively correlated with (higher) MT signal measures and GFAP intensity. Worse depressive symptoms (higher BDI-II scores) were associated with decreased oxygen consumption rate and spare respiratory capacity, concomitant with increased extracellular acidification rate in astrocytes. CONCLUSIONS: These findings show that factors in the sera of PWH alter mitochondrial activity in cultured human astrocytes, suggesting that mechanisms that alter mitochondrial and astrocyte homeostasis can be detected peripherally. Thus, in vitro cultures may provide a model to identify neuropathogenic mechanisms of depression or neurocognitive impairment in PWH and test personalized therapeutics for neurologic and psychiatric disorders.

The SARS-CoV-2 virus that led to COVID-19 is associated with significant and long-lasting neurologic symptoms in many patients, with an increased mortality risk for people with Alzheimer's disease (AD) and/or Down syndrome (DS). However, few studies have evaluated the neuropathological and inflammatory sequelae in postmortem brain tissue obtained from AD and people with DS with severe SARS-CoV-2 infections. We examined tau, beta-amyloid (Aβ), inflammatory markers and SARS-CoV-2 nucleoprotein in DS, AD, and healthy non-demented controls with COVID-19 and compared with non-infected brain tissue from each disease group (total n = 24). A nested ANOVA was used to determine regional effects of the COVID-19 infection on arborization of astrocytes (Sholl analysis) and percent-stained area of Iba-1 and TMEM 119. SARS-CoV-2 antibodies labeled neurons and glial cells in the frontal cortex of all subjects with COVID-19, and in the hippocampus of two of the three DS COVID-19 cases. SARS-CoV-2-related alterations were observed in peri-vascular astrocytes and microglial cells in the gray matter of the frontal cortex, hippocampus, and para-hippocampal gyrus. Bright field microscopy revealed scattered intracellular and diffuse extracellular Aβ deposits in the hippocampus of controls with confirmed SARS-CoV-2 infections. Overall, the present preliminary findings suggest that SARS-CoV-2 infections induce abnormal inflammatory responses in Down syndrome.

AimAstrocytes respond to stressors by acquiring a reactive state characterized by changes in their morphology and function. Molecules underlying reactive astrogliosis, however, remain largely unknown. Given that several studies observed increase in the Amyloid Precursor Protein (APP) in reactive astrocytes, we here test whether APP plays a role in reactive astrogliosis.MethodsWe investigated whether APP instigates reactive astroglios by examining in vitro and in vivo the morphology and function of naive and APP-deficient astrocytes in response to APP and well-established stressors.ResultsOverexpression of APP in cultured astrocytes led to remodeling of the intermediate filament network, enhancement of cytokine production, and activation of cellular programs centered around the interferon (IFN) pathway, all signs of reactive astrogliosis. Conversely, APP deletion abrogated remodeling of the intermediate filament network and blunted expression of IFN-stimulated gene products in response to lipopolysaccharide. Following traumatic brain injury (TBI), mouse reactive astrocytes also exhibited an association between APP and IFN, while APP deletion curbed the increase in glial fibrillary acidic protein observed canonically in astrocytes in response to TBI.ConclusionsThe APP thus represents a candidate molecular inducer and regulator of reactive astrogliosis. This finding has implications for understanding pathophysiology of neurodegenerative and other diseases of the nervous system characterized by reactive astrogliosis and opens potential new therapeutic avenues targeting APP and its pathways to modulate reactive astrogliosis.

The prion-like spread of protein aggregates is a leading hypothesis for the propagation of neurofibrillary lesions in the brain, including the spread of tau inclusions associated with Alzheimers disease. The mechanisms of cellular uptake of tau seeds and subsequent nucleated polymerization of cytosolic tau are major questions in the field, and the potential for coupling between the entry and nucleation mechanisms has been little explored. We found that in primary astrocytes and neurons, endocytosis of tau seeds leads to their accumulation in lysosomes. This in turn leads to lysosomal swelling, deacidification, and recruitment of ESCRT proteins, but not Galectin-3, to the lysosomal membrane. These observations are consistent with nanoscale damage of the lysosomal membrane. Live cell imaging and STORM superresolution microscopy further show that the nucleation of cytosolic tau occurs primarily at the lysosome membrane under these conditions. These data suggest that tau seeds escape from lysosomes via nanoscale damage rather than wholesale rupture and that nucleation of cytosolic tau commences as soon as tau fibril ends emerge from the lysosomal membrane.

Sleep disruption is a frequent problem of advancing age, often accompanied by low-grade chronic central and peripheral inflammation. We examined whether chronic neuroinflammation in the preoptic and basal forebrain area (POA-BF), a critical sleep-wake regulatory structure, contributes to this disruption. We developed a targeted viral vector designed to overexpress tumor necrosis factor-alpha (TNFα), specifically in astrocytes (AAV5-GFAP-TNFα-mCherry), and injected it into the POA of young mice to induce heightened neuroinflammation within the POA-BF. Compared to the control (treated with AAV5-GFAP-mCherry), mice with astrocytic TNFα overproduction within the POA-BF exhibited signs of increased microglia activation, indicating a heightened local inflammatory milieu. These mice also exhibited aging-like changes in sleep-wake organization and physical performance, including (a) impaired sleep-wake functions characterized by disruptions in sleep and waking during light and dark phases, respectively, and a reduced ability to compensate for sleep loss; (b) dysfunctional VLPO sleep-active neurons, indicated by fewer neurons expressing c-fos after suvorexant-induced sleep; and (c) compromised physical performance as demonstrated by a decline in grip strength. These findings suggest that inflammation-induced dysfunction of sleep- and wake-regulatory mechanisms within the POA-BF may be a critical component of sleep-wake disturbances in aging.

Astrocytes, the most abundant non-neuronal cell type in the mammalian brain, are crucial circuit components that respond to and modulate neuronal activity through calcium (Ca2+) signalling1-7. Astrocyte Ca2+ activity is highly heterogeneous and occurs across multiple spatiotemporal scales-from fast, subcellular activity3,4 to slow, synchronized activity across connected astrocyte networks8-10-to influence many processes5,7,11. However, the inputs that drive astrocyte network dynamics remain unclear. Here we used ex vivo and in vivo two-photon astrocyte imaging while mimicking neuronal neurotransmitter inputs at multiple spatiotemporal scales. We find that brief, subcellular inputs of GABA and glutamate lead to widespread, long-lasting astrocyte Ca2+ responses beyond an individual stimulated cell. Further, we find that a key subset of Ca2+ activity-propagative activity-differentiates astrocyte network responses to these two main neurotransmitters, and may influence responses to future inputs. Together, our results demonstrate that local, transient neurotransmitter inputs are encoded by broad cortical astrocyte networks over a minutes-long time course, contributing to accumulating evidence that substantial astrocyte-neuron communication occurs across slow, network-level spatiotemporal scales12-14. These findings will enable future studies to investigate the link between specific astrocyte Ca2+ activity and specific functional outputs, which could build a consistent framework for astrocytic modulation of neuronal activity.

Glaucoma leads to vision loss due to retinal ganglion cell death. Astrocyte reactivity contributes to neurodegeneration. Our recent study found that lipoxin B4 (LXB4), produced by retinal astrocytes, has direct neuroprotective actions on retinal ganglion cells. In this study, we aimed to investigate how the autacoid LXB4 influences astrocyte reactivity in the retina under inflammatory cytokine-induced activation and during ocular hypertension. The protective activity of LXB4 was investigated in vivo using the mouse silicone-oil model of chronic ocular hypertension. By employing a range of analytical techniques, including bulk RNA-seq, RNAscope in-situ hybridization, qPCR, and lipidomic analyses, we discovered the formation of lipoxins and expression of the lipoxin pathway in rodents (including the retina and optic nerve), primates (optic nerve), and human brain astrocytes, indicating the presence of this neuroprotective pathway across various species. Findings in the mouse retina identified significant dysregulation of the lipoxin pathway in response to chronic ocular hypertension, leading to an increase in 5-lipoxygenase (5-LOX) activity and a decrease in 15-LOX activity. This dysregulation was coincident with a marked upregulation of astrocyte reactivity. Reactive human brain astrocytes also showed a significant increase in 5-LOX. Treatment with LXB4 amplified the lipoxin biosynthetic pathway by restoring and amplifying the generation of another member of the lipoxin family, LXA4, and mitigated astrocyte reactivity in mouse retinas and human brain astrocytes. In conclusion, the lipoxin pathway is functionally expressed in rodents, primates, and human astrocytes, and is a resident neuroprotective pathway that is downregulated in reactive astrocytes. Novel cellular targets for LXB4s neuroprotective action are inhibition of astrocyte reactivity and restoration of lipoxin generation. Amplifying the lipoxin pathway is a potential target to disrupt or prevent astrocyte reactivity in neurodegenerative diseases, including retinal ganglion cell death in glaucoma.

Astrocytes are a direct target of neuromodulators and can influence neuronal activity on broad spatial and temporal scales in response to a rise in cytosolic calcium. However, our knowledge about how astrocytes are recruited during different animal behaviors remains limited. To measure astrocyte activity calcium in vivo during normative behaviors, we utilize a high-resolution, long working distance multicore fiber optic imaging system that allows visualization of individual astrocyte calcium transients in the cerebral cortex of freely moving mice. We define the spatiotemporal dynamics of astrocyte calcium changes during diverse behaviors, ranging from sleep-wake cycles to the exploration of novel objects, showing that their activity is more variable and less synchronous than apparent in head-immobilized imaging conditions. In accordance with their molecular diversity, individual astrocytes often exhibit distinct thresholds and activity patterns during explorative behaviors, allowing temporal encoding across the astrocyte network. Astrocyte calcium events were induced by noradrenergic and cholinergic systems and modulated by internal state. The distinct activity patterns exhibited by astrocytes provides a means to vary their neuromodulatory influence in different behavioral contexts and internal states.

IntroductionAlzheimer's disease (AD) is a neurodegenerative disorder with multifactorial etiology, including genetic factors that play a significant role in disease risk and resilience. However, the role of genetic diversity in preclinical AD studies has received limited attention.MethodsWe crossed five Collaborative Cross strains with 5xFAD C57BL/6J female mice to generate F1 mice with and without the 5xFAD transgene. Amyloid plaque pathology, microglial and astrocytic responses, neurofilament light chain levels, and gene expression were assessed at various ages.ResultsGenetic diversity significantly impacts AD-related pathology. Hybrid strains showed resistance to amyloid plaque formation and neuronal damage. Transcriptome diversity was maintained across ages and sexes, with observable strain-specific variations in AD-related phenotypes. Comparative gene expression analysis indicated correlations between mouse strains and human AD.DiscussionIncreasing genetic diversity promotes resilience to AD-related pathogenesis, relative to an inbred C57BL/6J background, reinforcing the importance of genetic diversity in uncovering resilience in the development of AD.HighlightsGenetic diversity's impact on AD in mice was explored. Diverse F1 mouse strains were used for AD study, via the Collaborative Cross. Strain-specific variations in AD pathology, glia, and transcription were found. Strains resilient to plaque formation and plasma neurofilament light chain (NfL) increases were identified. Correlations with human AD transcriptomics were observed.

IntroductionThe BIN1 coding variant rs138047593 (K358R) is linked to Late-Onset Alzheimer's Disease (LOAD) via targeted exome sequencing.MethodsTo elucidate the functional consequences of this rare coding variant on brain amyloidosis and neuroinflammation, we generated BIN1K358R knock-in mice using CRISPR/Cas9 technology. These mice were subsequently bred with 5xFAD transgenic mice, which serve as a model for Alzheimer's pathology.ResultsThe presence of the BIN1K358R variant leads to increased cerebral amyloid deposition, with a dampened response of astrocytes and oligodendrocytes, but not microglia, at both the cellular and transcriptional levels. This correlates with decreased neurofilament light chain in both plasma and brain tissue. Synaptic densities are significantly increased in both wild-type and 5xFAD backgrounds homozygous for the BIN1K358R variant.DiscussionThe BIN1 K358R variant modulates amyloid pathology in 5xFAD mice, attenuates the astrocytic and oligodendrocytic responses to amyloid plaques, decreases damage markers, and elevates synaptic densities.HighlightsBIN1 rs138047593 (K358R) coding variant is associated with increased risk of LOAD. BIN1 K358R variant increases amyloid plaque load in 12-month-old 5xFAD mice. BIN1 K358R variant dampens astrocytic and oligodendrocytic response to plaques. BIN1 K358R variant decreases neuronal damage in 5xFAD mice. BIN1 K358R upregulates synaptic densities and modulates synaptic transmission.

Canavan disease is a leukodystrophy caused by ASPA mutations that diminish oligodendroglial aspartoacylase activity, and is characterized by markedly elevated brain concentrations of the aspartoacylase substrate N-acetyl-l-aspartate (NAA) and by astroglial and intramyelinic vacuolation. Astroglia express NaDC3 (encoded by SLC13A3), a sodium-coupled transporter for NAA and other dicarboxylates. Astroglial conditional Slc13a3 deletion in aspartoacylase-deficient Canavan disease model mice ("CD mice") reversed brain NAA elevation and improved motor function. These results demonstrate that astroglial NaDC3 contributes to brain NAA elevation in CD mice, and suggest that suppressing astroglial NaDC3 activity would ameliorate human Canavan disease.

The ɛ4 allele of the apolipoprotein E gene (APOE), which translates to the APOE4 isoform, is the strongest genetic risk factor for late-onset Alzheimer disease (AD). Within the CNS, APOE is produced by a variety of cell types under different conditions, posing a challenge for studying its roles in AD pathogenesis. However, through powerful advances in research tools and the use of novel cell culture and animal models, researchers have recently begun to study the roles of APOE4 in AD in a cell type-specific manner and at a deeper and more mechanistic level than ever before. In particular, cutting-edge omics studies have enabled APOE4 to be studied at the single-cell level and have allowed the identification of critical APOE4 effects in AD-vulnerable cellular subtypes. Through these studies, it has become evident that APOE4 produced in various types of CNS cell - including astrocytes, neurons, microglia, oligodendrocytes and vascular cells - has diverse roles in AD pathogenesis. Here, we review these scientific advances and propose a cell type-specific APOE4 cascade model of AD. In this model, neuronal APOE4 emerges as a crucial pathological initiator and driver of AD pathogenesis, instigating glial responses and, ultimately, neurodegeneration. In addition, we provide perspectives on future directions for APOE4 research and related therapeutic developments in the context of AD.

Understanding metabolic heterogeneity is the key to uncovering the underlying mechanisms of metabolic-related diseases. Current metabolic imaging studies suffer from limitations including low resolution and specificity, and the model systems utilized often lack human relevance. Here, we present a single-cell metabolic imaging platform to enable direct imaging of lipid metabolism with high specificity in various human-derived 2D and 3D culture systems. Through the incorporation of an azide-tagged infrared probe, selective detection of newly synthesized lipids in cells and tissue became possible, while simultaneous fluorescence imaging enabled cell-type identification in complex tissues. In proof-of-concept experiments, newly synthesized lipids were directly visualized in human-relevant model systems among different cell types, mutation status, differentiation stages, and over time. We identified upregulated lipid metabolism in progranulin-knockdown human induced pluripotent stem cells and in their differentiated microglia cells. Furthermore, we observed that neurons in brain organoids exhibited a significantly lower lipid metabolism compared to astrocytes.

Alzheimer's disease (AD) is the leading cause of dementia in older adults, and the need for effective, sustainable therapeutic targets is imperative. The complement pathway has been proposed as a therapeutic target. C5aR1 inhibition reduces plaque load, gliosis, and memory deficits in animal models, however, the cellular bases underlying this neuroprotection were unclear. Here, we show that the C5aR1 antagonist PMX205 improves outcomes in the Arctic48 mouse model of AD. A combination of single cell and single nucleus RNA-seq analysis of hippocampi derived from males and females identified neurotoxic disease-associated microglia clusters in Arctic mice that are C5aR1-dependent, while microglial genes associated with synapse organization and transmission and learning were overrepresented in PMX205-treated mice. PMX205 also reduced neurotoxic astrocyte gene expression, but clusters associated with protective responses to injury were unchanged. C5aR1 inhibition promoted mRNA-predicted signaling pathways between brain cell types associated with cell growth and repair, while suppressing inflammatory pathways. Finally, although hippocampal plaque load was unaffected, PMX205 prevented deficits in short-term memory in female Arctic mice. In conclusion, C5aR1 inhibition prevents cognitive loss, limits detrimental glial polarization while permitting neuroprotective responses, as well as leaving most protective functions of complement intact, making C5aR1 antagonism an attractive therapeutic strategy for AD.

Breast cancer metastasis to the brain is a clinical challenge rising in prevalence. However, the underlying mechanisms, especially how cancer cells adapt a distant brain niche to facilitate colonization, remain poorly understood. A unique metabolic feature of the brain is the coupling between neurons and astrocytes through glutamate, glutamine, and lactate. Here we show that extracellular vesicles from breast cancer cells with a high potential to develop brain metastases carry high levels of miR-199b-5p, which shows higher levels in the blood of breast cancer patients with brain metastases comparing to those with metastatic cancer in other organs. miR-199b-5p targets solute carrier transporters (SLC1A2/EAAT2 in astrocytes and SLC38A2/SNAT2 and SLC16A7/MCT2 in neurons) to hijack the neuron-astrocyte metabolic coupling, leading to extracellular retention of these metabolites and promoting cancer cell growth. Our findings reveal a mechanism through which cancer cells of a non-brain origin reprogram neural metabolism to fuel brain metastases.

Astrocytes play a role in healthy cognitive function and Alzheimer's disease (AD). The transcriptional factor nuclear factor-κB (NF-κB) drives astrocyte diversity, but the mechanisms are not fully understood. By combining studies in human brains and animal models and selectively manipulating NF-κB function in astrocytes, we deepened the understanding of the role of astrocytic NF-κB in brain health and AD. In silico analysis of bulk and cell-specific transcriptomic data revealed the association of NF-κB and astrocytes in AD. Confocal studies validated the higher level of p50 NF-κB and phosphorylated-p65 NF-κB in glial fibrillary acidic protein (GFAP)+-astrocytes in AD versus non-AD subjects. In the healthy mouse brain, chronic activation of astrocytic NF-κB disturbed the proteomic milieu, causing a loss of mitochondrial-associated proteins and the rise of inflammatory-related proteins. Sustained NF-κB signaling also led to microglial reactivity, production of pro-inflammatory mediators, and buildup of senescence-related protein p16INK4A in neurons. However, in an AD mouse model, NF-κB inhibition accelerated β-amyloid and tau accumulation. Molecular biology studies revealed that astrocytic NF-κB activation drives the increase in GFAP and inflammatory proteins and aquaporin-4, a glymphatic system protein that assists in mitigating AD. Our investigation uncovered fundamental mechanisms by which NF-κB enables astrocytes' neuroprotective and neurotoxic responses in the brain.

The Wnt/β-catenin pathway contains multiple high-confidence risk genes that are linked to neurodevelopmental disorders, including autism spectrum disorder. However, its ubiquitous roles across brain cell types and developmental stages have made it challenging to define its impact on neural circuit development and behavior. Here, we show that TCF7L2, which is a key transcriptional effector of the Wnt/β-catenin pathway, plays a cell-autonomous role in postnatal astrocyte maturation and impacts adult social behavior. TCF7L2 was the dominant Wnt effector that was expressed in both mouse and human astrocytes, with a peak during astrocyte maturation. The conditional knockout of Tcf7l2 in postnatal astrocytes led to an enlargement of astrocytes with defective tiling and gap junction coupling. These mice also exhibited an increase in the number of cortical excitatory and inhibitory synapses and a marked increase in social interaction by adulthood. These data reveal an astrocytic role for developmental Wnt/β-catenin signaling in restricting excitatory synapse numbers and regulating adult social behavior.

A cancerous tumor in the brain known as glioblastoma multiforme (GBM) originates from astrocytes of the central nervous system. Consequently, GBM poses significant challenges to the oncology community because of its aggressive characteristics and poor prognosis. GBM hallmarks include fast growth, invasiveness, and high rates of recurrence. This tumor is highly heterogeneous with different genetic and molecular features found within the tumor cells. There is an ongoing obstacle to conceptualizing effective management for this grappling disease. This is largely due to the tumor displaying intra-heterogeneity, in addition to a plethora of differences in the tumor’s microenvironment. The heterogeneity exhibited by this tumor not only makes it more resistant to treatment but also influences its ability to evolve. The Epidermal Growth Factor Receptor (EGFR) which falls under the Human Epidermal Growth Factor (ErbB) family is a transmembrane receptor that assists in understanding complex molecular pathways involved in GBM formation. EGFR mutations have been shown to affect signaling cascades including Ras/Raf/MEK/ERK, PI3K/Akt/mTOR, JAK/STAT, PLC/PKC among others transforming cellular machinery involved in cell survival, proliferation and invasion. Knowledge about EGFR’s aberrant mutations can be useful for developing novel therapeutic strategies aimed at EGFR inhibition in GBM therapy. This gives hope for patients with this challenging disease to have better outcomes.

This study suggests that glial cells, specifically astrocytes, in discrete functional domains of the hippocampus play different roles in the processing of aversive and rewarding experiences. DH, dorsal hippocampus; IH, intermediate hippocampus; VH, ventral hippocampus; MWM, Morris water maze; TPP, T-maze place preference; PA, passive avoidance. [Display omitted] • The hippocampus long-axis is functionally organized for reward and aversion. • Astrocytes promote the functional distinction of the hippocampus long-axis. • Different functions of astrocytes in discrete functional domains of the hippocampus. This study aims to investigate whether glial cells, in particular putative astrocytes, contribute to functional distinctions between the dorsal (DH), intermediate (IH), and ventral (VH) hippocampus. To evaluate this, we performed three different behavioral tasks (i.e., Morris water maze; MWM, Passive avoidance; PA, T-maze place preference; TPP) to determine whether the DH, IH, and VH are necessary for each task. Sensitivity of behavioral tasks was confirmed using lidocaine (2 %, 1 μl) reversible inactivation. Subsequently, we examined the effects of silencing astrocytes, using fluorocitrate (FC, 1 mM/1 μl), into the DH, IH, and VH on these tasks. The effects of drugs were examined separately. We observed that injection of FC into the DH resulted in a significant impairment in MWM performance. In contrast, while FC injections into the IH or VH did not prevent platform localization during the acquisition phase, rats showed difficulty recalling the target zone during the retrieval phase. In the PA test, FC injection into the VH impaired task learning and memory. During the acquisition phase, FC injection into the DH or IH did not differ from the control in the number of shocks; however, during retrieval, there was a significant decrease in the latency before entering the dark chamber. The TPP test performance was impaired by FC injection in the IH. In sum, we show that glial cells, especially astrocytes in specific functional regions of the hippocampus, play distinct roles in processing aversive and rewarding experiences and contribute to the functional organization of the hippocampal longitudinal axis. [ABSTRACT FROM AUTHOR]

• Neuroinflammation significantly contributes to the pathophysiology and chronicity of migraines. • Microglia and astrocytes are central to neuroinflammatory responses in migraine development. • Cytokines and chemokines are important in glia-neuron crosstalk during migraine pathogenesis. • [11C] PBR28-targeted imaging and single-cell sequencing advance understanding of neuroinflammation mechanisms in migraine. • Estrogen and TRPM8 emerge as promising personalized therapeutic targets. Migraine is a complex neurological disorder with neuroinflammation playing a crucial role in its pathogenesis. This review provides an overview of the neuroinflammation mechanisms in migraine, focusing on both cellular and molecular aspects. At the cellular level, we examine the role of glial cells, including astrocytes, microglia, oligodendrocytes in the central nervous system, and Schwann cells and satellite glial cells in the peripheral nervous system. On the molecular level, we explore the signaling pathways, including IL-1β, TNF-α, IL-6, and non-coding RNAs, that mediate cell interactions or independent actions. Some of the molecular signaling pathways mentioned, such as TNF-α and IL-1β, have been investigated as druggable targets. Recent advancements, such as [11C] PBR28-targeted imaging for visualizing astrocyte activation and single-cell sequencing for exploring cellular heterogeneity, represent breakthroughs in understanding the mechanisms of neuroinflammation in migraine. By considering factors for personalized treatments, estrogen and TRPM8 emerge as promising therapeutic targets regarding sexual dimorphism. These advancements may help bridge the gap between preclinical findings and clinical applications, ultimately leading to more precise and personalized options for migraine patients. [ABSTRACT FROM AUTHOR]

Major depressive disorder (MDD) is a prevalent mental health condition characterized by persistent feelings of sadness and hopelessness, affecting millions globally. The precise molecular mechanisms underlying MDD remain elusive, necessitating comprehensive investigations. Our study integrates transcriptomic analysis, functional assays, and computational modeling to explore the molecular landscape of MDD, focusing on the DLPFC. We identify key genomic alterations and co-expression modules associated with MDD, highlighting potential therapeutic targets. Functional enrichment and protein–protein interaction analyses emphasize the role of astrocytes in MDD progression. Machine learning is employed to develop a predictive model for MDD risk assessment. Single-cell and spatial transcriptomic analyses provide insights into cell type–specific expression patterns, particularly regarding astrocytes. We have identified significant genomic alterations and co-expression modules associated with MDD in the DLPFC. Key genes involved in neuroactive ligand-receptor interaction pathways, notably in astrocytes, have been highlighted. Additionally, we developed a predictive model for MDD risk assessment based on selected key genes. Single-cell and spatial transcriptomic analyses underscored the role of astrocytes in MDD. Virtual screening of compounds targeting GPR37L1, KCNJ10, and PPP1R3C proteins has identified potential therapeutic candidates. In summary, our comprehensive approach enhances the understanding of MDD's molecular underpinnings and offers promising opportunities for advancing therapeutic interventions, ultimately aiming to alleviate the burden of this debilitating mental health condition. Key messages: Our investigation furnishes insightful revelations concerning the dysregulation of astrocyte-associated processes in MDD. We have pinpointed specific genes, namely KCNJ10, PPP1R3C, and GPR37L1, as potential candidates warranting further exploration and therapeutic intervention. We incorporate a virtual screening of small molecule compounds targeting KCNJ10, PPP1R3C, and GPR37L1, presenting a promising trajectory for drug discovery in MDD. [ABSTRACT FROM AUTHOR]

Alzheimer´s disease (AD) is characterized by neuronal function loss and degeneration. The integrity of the axon initial segment (AIS) is essential to maintain neuronal function and output. AIS alterations are detected in human post-mortem AD brains and mice models, as well as, neurodevelopmental and mental disorders. However, the mechanisms leading to AIS deregulation in AD and the extrinsic glial origin are elusive. We studied early postnatal differences in AIS cellular/molecular mechanisms in wild-type or APP/PS1 mice and combined neuron-astrocyte co-cultures. We observed AIS integrity alterations, reduced ankyrinG expression and shortening, in APP/PS1 mice from P21 and loss of AIS integrity at 21 DIV in wild-type and APP/PS1 neurons in the presence of APP/PS1 astrocytes. AnkyrinG decrease is due to mRNAs and protein reduction of retinoic acid synthesis enzymes Rdh1 and Aldh1b1, as well as ADNP (Activity-dependent neuroprotective protein) in APP/PS1 astrocytes. This effect was mimicked by wild-type astrocytes expressing ADNP shRNA. In the presence of APP/PS1 astrocytes, wild-type neurons AIS is recovered by inhibition of retinoic acid degradation, and Adnp-derived NAP peptide (NAPVSIPQ) addition or P2X7 receptor inhibition, both regulated by retinoic acid levels. Moreover, P2X7 inhibitor treatment for 2 months impaired AIS disruption in APP/PS1 mice. Our findings extend current knowledge on AIS regulation, providing data to support the role of astrocytes in early postnatal AIS modulation. In conclusion, AD onset may be related to very early glial cell alterations that induce AIS and neuronal function changes, opening new therapeutic approaches to detect and avoid neuronal function loss. [ABSTRACT FROM AUTHOR]

LRRK2 is a relatively common genetic risk factor for Parkinson's disease (PD), with six coding variants known to cause familial PD. Non-coding variation at the same locus is also associated with sporadic PD. LRRK2 plays a role in many different intracellular signaling cascades including those involved in endolysosomal function, cytoskeletal dynamics, and Ca2+ homeostasis. PD-causing LRRK2 mutations cause hyperactive LRRK2 kinase activity, resulting in altered cellular signaling. Importantly, LRRK2 is lowly expressed in neurons and prominently expressed in non-neuronal cells in the brain. In this review, we will summarize recent and novel findings on the effects of PD-causing LRRK2 mutations in different nervous system cell types. This review will also provide novel insight into future areas of research at the intersection of LRRK2 cell biology, cell type specificity, and PD. [ABSTRACT FROM AUTHOR]

This review is an attempt to compile existing hypotheses on the mechanisms underlying the initiation and progression of Alzheimer's disease (AD), starting from sensory impairments observed in AD and concluding with molecular events that are typically associated with the disease. These events include spreading of amyloid plaques and tangles of hyperphosphorylated tau and formation of Hirano and Biondi bodies as well as the development of oxidative stress. We have detailed the degenerative changes that occur in several neuronal populations, including the cholinergic neurons in the nucleus basalis of Meynert, the histaminergic neurons in the tuberomammillary nucleus, the serotonergic neurons in the raphe nuclei, and the noradrenergic neurons in the locus coeruleus. Furthermore, we discuss the potential role of iron accumulation in the brains of subjects with AD in the disease progression which served as a basis for the idea that iron chelation in the brain may mitigate oxidative stress and decelerate disease development. We also draw attention to possible role of sympathetic system and, more specifically, noradrenergic neurons of the superior cervical ganglion in triggering of the disease. We also explore the alternative possibility of compensatory protective changes that may occur in these neurons to support cholinergic function in the forebrain of subjects with AD. [ABSTRACT FROM AUTHOR]

[Display omitted] • Activation of astrocytic NMDARs rescued Aβ-induced reduction of BDNF in astrocytes. • Activation of astrocytic NMDARs inhibited Aβ-induced elevation of astrocytic GFAP. • Activation of astrocytic NMDARs inhibited Aβ-induced elevation of astrocytic C3. • Activation of astrocytic NMDARs inhibited Aβ-induced increase of astrocytic pNF-κB. N-methyl-D-aspartate receptors (NMDARs) play a crucial role in mediating Amyloid-β (Aβ) synaptotoxicity. Our previous studies have demonstrated an opposite (neuroprotection and neurotoxicity) effect of activating astrocytic and neuronal NMDARs with higher dose (10 μM) of NMDA, an agonist of NMDARs. By contrast, activating neuronal or astrocyitc NMDARs with lower dose (1 μM) of NMDA both exerts neuroprotective effect in Aβ-induced neurotoxicity. However, the underlying mechanism of activating astrocytic NMDARs with lower dose of NMDA to protect against Aβ neurotoxicity remains unclear. Based on our previous related work, in this study, using a co-cultured cell model of primary hippocampal neurons and astrocytes, we further investigated the possible factors involved in 1 μM of NMDA activating astrocytic NMDARs to oppose Aβ-induced synaptotoxicity. Our results showed that activation of astrocytic NMDARs by 1 μM NMDA rescued Aβ-induced reduction of brain-derived neurotrophic factor (BDNF), and inhibited Aβ-induced increase of GFAP, complement 3 (C3) and activation of NF-κB. Furthermore, blockade of astrocytic GluN2A with TCN201 abrogated the ability of 1 μM NMDA to counteract the effects of Aβ decreasing BDNF, and increasing GFAP, C3 and activation of NF-κB. These findings suggest that activation of astrocytic NMDARs protect against Aβ-induced synaptotoxicity probably through elevating BDNF and suppressing GFAP and C3. Our present research provides valuable insights for elucidating the underlying mechanism of astrocytic NMDARs activation resisting the toxic effects of Aβ. [ABSTRACT FROM AUTHOR]

• Adolescent male LB mice, but not their female littermates, exhibit significant deficits in contextual fear conditioning (CFC) and synaptic connectivity in the hippocampus. • LB impaired microglial-mediated synaptic pruning in the hippocampus of male and female postnatal day 17 (P17) but not in adolescent P33 mice when synaptic engulfment by microglia is substantially lower. • Transient ablation of microglia caused similar sex-specific behavioral and synaptic abnormalities to those observed in adolescent LB mice. • Chemogenetic activation of microglia from P13-17 reversed the microglial-mediated phagocytic deficits at P17 and restored normal CFC and synaptic connectivity in adolescent LB male mice. • LB female, but not LB male littermates, upregulate the expression of the membrane bound receptor MEGF10 and enhance synaptic pruning in astrocytes. Abnormal development and function of the hippocampus are two of the most consistent findings in humans and rodents exposed to early-life adversity (ELA), with males often being more affected than females. Using the limited bedding (LB) paradigm as a rodent model of ELA, we found that male adolescent mice that had been exposed to LB exhibit significant deficits in contextual fear conditioning and synaptic connectivity in the hippocampus, which are not observed in females. This is linked to altered developmental refinement of connectivity, with LB severely impairing microglial-mediated synaptic pruning in the hippocampus of male and female pups on postnatal day 17 (P17), but not in adolescent P33 mice when levels of synaptic engulfment by microglia are substantially lower. Since the rodent hippocampus undergoes intense synaptic pruning during the second and third weeks of life, we investigated whether microglia are required for the synaptic and behavioral aberrations observed in adolescent LB mice. Indeed, transient ablation of microglia from P13-21 in normally developing mice caused sex-specific behavioral and synaptic abnormalities similar to those observed in adolescent LB mice. Furthermore, chemogenetic activation of microglia during the same period reversed the microglial-mediated phagocytic deficits at P17 and restored normal contextual fear conditioning and synaptic connectivity in adolescent LB male mice. Our data support an additional contribution of astrocytes in the sex-specific effects of LB, with increased expression of the membrane receptor MEGF10 and enhanced synaptic engulfment in hippocampal astrocytes of 17-day-old LB females, but not in LB male littermates. These findings suggest a potential compensatory mechanism that may explain the relative resilience of LB females. Collectively, our study highlights a novel role for glial cells in mediating sex-specific hippocampal deficits in a mouse model of ELA. [ABSTRACT FROM AUTHOR]

• PM 0.2 exposure induced the disorders of α-syn degradation in mice and astrocytes. • LAMP2-mediated autophagy-lysosome pathway was disturbed and underlied α-syn pathology induced by PM 0.2 exposure. • PP242 improved the disorder of α-syn through TFEB/LAMP2 pathway. Atmospheric particulate matter (PM) exacerbates the risk factor for Alzheimer's and Parkinson's diseases (PD) by promoting the alpha-synuclein (α-syn) pathology in the brain. However, the molecular mechanisms of astrocytes involvement in α-syn pathology underlying the process remain unclear. This study investigated PM with particle size <200 nm (PM 0.2) exposure-induced α-syn pathology in ICR mice and primary astrocytes, then assessed the effects of mammalian target of rapamycin inhibitor (PP242) in vitro studies. We observed the α-syn pathology in the brains of exposed mice. Meanwhile, PM 0.2 -exposed mice also exhibited the activation of glial cell and the inhibition of autophagy. In vitro study, PM 0.2 (3, 10 and 30 µg/mL) induced inflammatory response and the disorders of α-syn degradation in primary astrocytes, and lysosomal-associated membrane protein 2 (LAMP2)-mediated autophagy underlies α-syn pathology. The abnormal function of autophagy-lysosome was specifically manifested as the expression of microtubule-associated protein light chain 3 (LC3II), cathepsin B (CTSB) and lysosomal abundance increased first and then decreased, which might both be a compensatory mechanism to toxic α-syn accumulation induced by PM 0.2. Moreover, with the transcription factor EB (TFEB) subcellular localization and the increase in LC3II, LAMP2, CTSB, and cathepsin D proteins were identified, leading to the restoration of the degradation of α-syn after the intervention of PP242. Our results identified that PM 0.2 exposure could promote the α-syn pathological dysregulation in astrocytes, providing mechanistic insights into how PM 0.2 increases the risk of developing PD and highlighting TFEB/LAMP2 as a promising therapeutic target for antagonizing PM 0.2 toxicity. [Display omitted] [ABSTRACT FROM AUTHOR]

Parkinson's disease is typically characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta. Many studies have been performed based on the supplementation of lost dopaminergic neurons to treat Parkinson's disease. The initial strategy for cell replacement therapy used human fetal ventral midbrain and human embryonic stem cells to treat Parkinson's disease, which could substantially alleviate the symptoms of Parkinson's disease in clinical practice. However, ethical issues and tumor formation were limitations of its clinical application. Induced pluripotent stem cells can be acquired without sacrificing human embryos, which eliminates the huge ethical barriers of human stem cell therapy. Another widely considered neuronal regeneration strategy is to directly reprogram fibroblasts and astrocytes into neurons, without the need for intermediate proliferation states, thus avoiding issues of immune rejection and tumor formation. Both induced pluripotent stem cells and direct reprogramming of lineage cells have shown promising results in the treatment of Parkinson's disease. However, there are also ethical concerns and the risk of tumor formation that need to be addressed. This review highlights the current application status of cell reprogramming in the treatment of Parkinson's disease, focusing on the use of induced pluripotent stem cells in cell replacement therapy, including preclinical animal models and progress in clinical research. The review also discusses the advancements in direct reprogramming of lineage cells in the treatment of Parkinson's disease, as well as the controversy surrounding in vivo reprogramming. These findings suggest that cell reprogramming may hold great promise as a potential strategy for treating Parkinson's disease. [ABSTRACT FROM AUTHOR]

Neuronal hyperexcitability within developing cortical circuits is a common characteristic of several heritable neurodevelopmental disorders, including Fragile X Syndrome (FXS), intellectual disability and autism spectrum disorders (ASD). While this aberrant circuitry is typically studied from a neuron-centric perspective, glial cells secrete soluble factors that regulate both neurite extension and synaptogenesis during development. The nucleotide-mediated purinergic signalling system is particularly instrumental in facilitating these effects. We recently reported that within a FXS animal model, the Fmr1 KO mouse, the purinergic signalling system is upregulated in cortical astrocytes leading to altered secretion of synaptogenic and plasticity-related proteins. In this study, we examined whether elevated astrocyte purinergic signalling also impacts neuronal morphology and connectivity of Fmr1 KO cortical neurons. Here, we found that conditioned media from primary Fmr1 KO astrocytes was sufficient to enhance neurite extension and complexity of both wildtype and Fmr1 KO neurons to a similar degree as UTP-mediated outgrowth. Significantly enhanced firing was also observed in Fmr1 KO neuron-astrocyte co-cultures grown on microelectrode arrays but was associated with large deficits in firing synchrony. The selective P2Y2 purinergic receptor antagonist AR-C 118925XX effectively normalized much of the aberrant Fmr1 KO activity, designating P2Y2 as a potential therapeutic target in FXS. These results not only demonstrate the importance of astrocyte soluble factors in the development of neural circuitry, but also show that P2Y purinergic receptors play a distinct role in pathological FXS neuronal activity. [ABSTRACT FROM AUTHOR]

The activity of HIV-1 and its viral proteins within the central nervous system (CNS) is responsible for a wide array of neuropathological effects, resulting in a spectrum of neurocognitive deficits defined as HIV-associated neurocognitive disorders (HAND). Amongst the various viral proteins, the transactivator of transcription (Tat) remains detectable even with effective antiretroviral therapy (ART) and suppressed viremia, highlighting the significance of this protein in the modern ART era. Tat has been extensively researched in both fundamental and clinical settings due to its role in neuroinflammation, neuronal damage, and neurocognitive impairment amongst people living with HIV (PLHIV). To date, numerous fundamental studies have explored Tat-induced neuroinflammation. However, there is no clear consensus on the most frequently studied inflammatory markers or the consistency in the levels of these Tat-induced inflammatory marker levels across different studies. Therefore, we conducted a scoping review of studies investigating Tat-induced neuroinflammation. We conducted searches in PubMed, Scopus, and Web of Science databases using a search protocol tailored specifically to adhere to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses for scoping reviews (PRISMA-ScR) guidelines. From the 22 included studies, findings suggest that the HIV-1 Tat protein amplifies levels of neuroinflammatory markers. Amongst the vast array of inflammatory markers explored in the included studies, consistent results point to higher levels of CCL2, IL-6, IL-8, and TNF-α in primary cells and cell lines exposed to or transfected with HIV-1 Tat. These markers are regulated by key inflammatory pathways, such as the extracellular signal-regulated kinase (ERK)1/2 mitogen-activated protein kinase (MAPK) pathway, the phosphatidylinositol 3-kinase (PI3K) pathway, the p38 MAPK pathway, and nuclear factor-kB (NF-kB). Furthermore, Tat has been shown to induce neuronal apoptosis, both directly and indirectly. With regards to study designs, utilizing full-length Tat101 at concentrations ranging from 100 to 1000 ng/ml and durations of 24 and 48 h appears optimal for investigating Tat-induced neuroinflammation. In this context, we highlight specific inflammatory markers and pathways that are potentially pivotal in Tat-induced neuroinflammation and subsequent neuronal damage. A deeper investigation into these markers and pathways is crucial to better understand their roles in the development of HAND. [ABSTRACT FROM AUTHOR]

Unlike mammals, species such as fish and amphibians can regenerate damaged spinal cords, offering insights into potential therapeutic targets. This study investigates the structural features of the molly fish spinal cord through light and electron microscopy. The most notable characteristic was the presence of Mauthner cells (M‐cells), which exhibited large cell bodies and processes, as well as synaptic connections with astrocytes. These astrocytic connections contained synaptic vesicles, suggesting electrical transmission at the M‐cell endings. Astrocytes, which were labeled with glial fibrillary acidic protein (GFAP), contained cytoplasmic glycogen granules, potentially serving as an emergency fuel source. Two types of oligodendrocytes were identified: a small, dark cell and a larger, lighter cell, both of which reacted strongly with oligodendrocyte transcription factor 2 (Olig2). The dark oligodendrocyte resembled human oligodendrocyte precursors, while the light oligodendrocyte was similar to mature human oligodendrocytes. Additionally, proliferative neurons in the substantia grisea centralis expressed myostatin, Nrf2, and Sox9. Collectively, these findings suggest that the molly fish spinal cord has advanced structural features conducive to spinal cord regeneration and could serve as an excellent model for studying central nervous system regeneration. Further studies on the functional aspects of the molly fish spinal cord are recommended. Research Highlights: Mauthner cells (M‐cell), with their typical large cell body and processes, were the most characteristic feature in Molly fish spinal cord, where it presented synaptic connections with astrocytes and their ends contained synaptic vesicles indicating an electrical transmission in the M‐cells endings.Two types of oligodendrocytes could be recognized; both reacted intensely with Oligodendrocyte transcription factor 2 (Olig2).The proliferative neurons of the substantia grisea centralis expressed myostatin, Nrf2, and Sox9.The findings of this study suggest that molly fish possess highly developed structural features conducive to spinal cord regeneration. Consequently, they could be deemed an exemplary model for investigating central nervous system regeneration. [ABSTRACT FROM AUTHOR]

Techniques for triggering neural differentiation of embryonic and induced pluripotent stem cells into neural stem cells and neurons have been established. However, neural induction of mesenchymal stem cells, including dental pulp stem cells (DPSCs), has been assessed primarily based on neural-related gene regulation, and detailed studies into the characteristics and differentiation status of cells are lacking. Therefore, this study was aimed at evaluating the cellular components and differentiation pathways of neural lineage cells obtained via neural induction of human DPSCs. Human DPSCs were induced to neural cells in monolayer culture and examined for gene expression and mechanisms underlying differentiation using microarray-based ingenuity pathway analysis. In addition, the neural lineage cells were subjected to single-cell RNA sequencing (scRNA-seq) to classify cell populations based on gene expression profiles and to elucidate their differentiation pathways. Ingenuity pathway analysis revealed that genes exhibiting marked overexpression, post-neuronal induction, such as FABP7 and ZIC1, were associated with neurogenesis. Furthermore, in canonical pathway analysis, axon guidance signals demonstrated maximum activation. The scRNA-seq and cell type annotations revealed the presence of neural progenitor cells, astrocytes, neurons, and a small number of non-neural lineage cells. Moreover, trajectory and pseudotime analyses demonstrated that the neural progenitor cells initially engendered neurons, which subsequently differentiated into astrocytes. This result indicates that the aforementioned neural induction strategy generated neural stem/progenitor cells from DPSCs, which might differentiate and proliferate to constitute neural lineage cells. Therefore, neural induction of DPSCs may present an alternative approach to pluripotent stem cell-based therapeutic interventions for nervous system disorders. [ABSTRACT FROM AUTHOR]

Aim Methods Results Conclusions Calcium‐activated ligand‐gated chloride channels, beyond their role in maintaining anion homeostasis, modulate neuronal excitability by facilitating nonvesicular neurotransmitter release. BEST1, a key member of this family, is permeable to γ‐aminobutyric acid (GABA) and glutamate. While astrocytic BEST1 is well‐studied and known to regulate neurotransmitter levels, its distribution and role in other brain cell types remain unclear. This study aimed to reassess the localization of BEST1 in the mouse cerebral cortex.We examined the localization and distribution of BEST1 in the mouse parietal cortex using light microscopy, confocal double‐labeling with markers for astrocytes, neurons, microglia, and oligodendrocyte precursor cells, and 3D reconstruction techniques.In the cerebral cortex, BEST1 is more broadly distributed than previously thought. Neurons are the second most abundant BEST1+ cell type in the cerebral cortex, following astrocytes. BEST1 is diffusely expressed in neuronal somatic and neuropilar domains and is present at glutamatergic and GABAergic terminals, with a prevalence at GABAergic terminals. We also confirmed that BEST1 is expressed in cortical microglia and identified it in oligodendrocyte precursor cells, albeit to a lesser extent.Together, these findings suggest that BEST1's role in controlling neurotransmission may extend beyond astrocytes to include other brain cells. Understanding BEST1's function in these cells could offer new insights into the molecular mechanisms shaping cortical circuitry. Further research is needed to clarify the diverse roles of BEST1 in both normal and pathophysiological conditions. [ABSTRACT FROM AUTHOR]

Summary Neuroinflammation, characterized by a complex interplay among innate and adaptive immune responses within the central nervous system (CNS), is crucial in responding to infections, injuries, and disease pathologies. However, the dysregulation of the neuroinflammatory response could significantly affect neurons in terms of function and structure, leading to profound health implications. Although tremendous progress has been made in understanding the relationship between neuroinflammatory processes and alterations in neuronal integrity, the specific implications concerning both structure and function have not been extensively covered, with the exception of perspectives on glial activation and neurodegeneration. Thus, this review aims to provide a comprehensive overview of the multifaceted interactions among neurons and key inflammatory players, exploring mechanisms through which inflammation influences neuronal functionality and structural integrity in the CNS. Further, it will discuss how these inflammatory mechanisms lead to impairment in neuronal functions and architecture and highlight the consequences caused by dysregulated neuronal functions, such as cognitive dysfunction and mood disorders. By integrating insights from recent research findings, this review will enhance our understanding of the neuroinflammatory landscape and set the stage for future interventions that could transform current approaches to preserve neuronal integrity and function in CNS‐related inflammatory conditions. [ABSTRACT FROM AUTHOR]

The extracellular matrix plays a critical role in modulating cell behaviour in the developing and adult central nervous system influencing neural cell morphology, function and growth. Neurons and astrocytes, play vital roles in neural signalling and support respectively and respond to cues from the surrounding matrix environment. However, a better understanding of the impact of specific individual extracellular matrix proteins on both neurons and astrocytes is critical for advancing the development of matrix‐based scaffolds for neural repair applications. This study aimed to provide an in‐depth analysis of how different commonly used extracellular matrix proteins— laminin‐1, Fn, collagen IV, and collagen I—affect the morphology and growth of trophic induced pluripotent stem cell (iPSC)‐derived astrocyte progenitors and mouse motor neuron‐like cells. Following a 7‐day culture period, morphological assessments revealed that laminin‐1, fibronectin, and collagen‐IV, but not collagen I, promoted increased process extension and a stellate morphology in astrocytes, with collagen‐IV yielding the greatest increases. Subsequent analysis of neurons grown on the different extracellular matrix proteins revealed a similar pattern with laminin‐1, fibronectin, and collagen‐IV supporting robust neurite outgrowth. fibronectin promoted the greatest increase in neurite extension, while collagen‐I did not enhance neurite growth compared to poly‐L‐lysine controls. Super‐resolution microscopy highlighted extracellular matrix‐specific nanoscale changes in cytoskeletal organization, with distinct patterns of actin filament distribution where the three basement membrane‐associated proteins (laminin‐1, fibronectin, and collagen‐IV) promoted the extension of fine cellular processes. Overall, this study demonstrates the potent effect of laminin‐1, fibronectin and collagen‐IV to promote both iPSC‐derived astrocyte progenitor and neuronal growth, yielding detailed insights into the effect of extracellular matrix proteins on neural cell morphology at both the whole cell and nanoscale levels. The ability of laminin‐1, collagen‐IV and fibronectin to elicit strong growth‐promoting effects highlight their suitability as optimal extracellular matrix proteins to incorporate into neurotrophic biomaterial scaffolds for the delivery of cell cargoes for neural repair. [ABSTRACT FROM AUTHOR]

The mammalian cerebral cortex is composed of a rich diversity of cell types. Sensory cortical cells are organized into networks that rely on their functional diversity to ultimately carry out a variety of sophisticated cognitive functions for perception, learning, and memory. The auditory cortex (AC) has been most extensively studied for its experience-dependent effects, including for perceptual learning and associative memory. Here, we used single-nucleus RNA sequencing (snRNA-seq) in the AC of the adult rat to investigate the breadth of transcriptionally diverse cell types that likely support the role of AC in experience-dependent functions. A variety of unique excitatory and inhibitory neuron subtypes were identified that harbor unique transcriptional profiles of genes with putative relevance for the adaptive neuroplasticity of cortical microcircuits. In addition, we report for the first time a diversity of astrocytes in AC that may represent functionally unique subtypes, including those that could integrate experience-dependent adult neuroplasticity at cortical synapses. Together, these results pave the way for building models of how cortical neurons work in concert with astrocytes to fulfill dynamic and experience-dependent cognitive functions. [ABSTRACT FROM AUTHOR]

Mitochondrial dysfunction is a well‐established hallmark of Alzheimer's disease (AD). Despite recent documentation of transcellular mitochondrial transfer, its role in the pathogenesis of AD remains unclear. In this study, we report an impairment of mitochondrial quality within the astrocytes and neurons of adult 5 × FAD mice. Following treatment with mitochondria isolated from aged astrocytes induced by exposure to amyloid protein or extended cultivation, cultured neurons exhibited an excessive generation of reactive oxygen species and underwent neurite atrophy. Notably, aerobic exercise enhanced mitochondrial quality by upregulating CD38 within hippocampal astrocytes of 5 × FAD mice. Conversely, the knockdown of CD38 diminished astrocytic–neuronal mitochondrial transfer, thereby abolishing the ameliorative effects of aerobic exercise on neuronal oxidative stress, β‐amyloid plaque deposition, and cognitive dysfunction in 5 × FAD mice. These findings unveil an unexpected mechanism through which aerobic exercise facilitates the transference of healthy mitochondria from astrocytes to neurons, thus countering the AD‐like progression. [ABSTRACT FROM AUTHOR]

Astrocytes are crucial for the functioning of the nervous system as they maintain the ion homeostasis via volume regulation. Pathological states, such as amyotrophic lateral sclerosis (ALS), affect astrocytes and might even cause a loss of such functions. In this study, we examined astrocytic swelling/volume recovery in both the brain and spinal cord of the SOD1 animal model to determine the level of their impairment caused by the ALS-like pathology. Astrocyte volume changes were measured in acute brain or spinal cord slices during and after exposure to hyperkalemia. We then compared the results with alterations of extracellular space (ECS) diffusion parameters, morphological changes, expression of the Kir4.1 channel and the potassium concentration measured in the cerebrospinal fluid, to further disclose the link between potassium and astrocytes in the ALS-like pathology. Morphological analysis revealed astrogliosis in both the motor cortex and the ventral horns of the SOD1 spinal cord. The activated morphology of SOD1 spinal astrocytes was associated with the results from volume measurements, which showed decreased swelling of these cells during hyperkalemia. Furthermore, we observed lower shrinkage of ECS in the SOD1 spinal ventral horns. Immunohistochemical analysis then confirmed decreased expression of the Kir4.1 channel in the SOD1 spinal cord, which corresponded with the diminished volume regulation. Despite astrogliosis, cortical astrocytes in SOD1 mice did not show alterations in swelling nor changes in Kir4.1 expression, and we did not identify significant changes in ECS parameters. Moreover, the potassium level in the cerebrospinal fluid did not deviate from the physiological concentration. The results we obtained thus suggest that ALS-like pathology causes impaired potassium uptake associated with Kir4.1 downregulation in the spinal astrocytes, but based on our data from the cortex, the functional impairment seems to be independent of the morphological state. [ABSTRACT FROM AUTHOR]

The neurobiological mechanisms underlying major depressive disorder (MDD) remain largely unexplored due to the limited availability of study models in humans. Induced pluripotent stem cells (iPSCs) have overcome multiple limitations of retrospective clinical studies, contributing to a more detailed understanding of the molecular pathways that presumably contribute to the manifestation of depression. Despite the significant progress made by these study models, there are still more formidable challenges that will eventually be addressed by these platforms, as further studies may eventually emerge. This review will examine the most recent advances in the comprehension of depression by using human neurons and non-neuronal cells derived from induced pluripotent stem cells of patients with depression. This study highlights the importance of using these platforms to increase our knowledge of depression and address this psychiatric disorder more efficiently. [ABSTRACT FROM AUTHOR]

Stem cell therapies for Parkinson's disease are at an exciting time of development, and several clinical trials have recently been initiated. Human pluripotent stem cells are differentiated into transplantable dopamine (DA) progenitors which are proliferative at the time of grafting and undergo terminal differentiation and maturation in vivo. While the progenitors are homogeneous at the time of transplantation, they give rise to heterogeneous grafts composed not only of therapeutic DA neurons but also of other mature cell types. The mechanisms for graft diversification are unclear. We used single-nucleus RNA-seq and ATAC-seq to profile DA progenitors before transplantation combined with molecular barcode-based tracing to determine origin and shared lineages of the mature cell types in the grafts. Our data demonstrate that astrocytes, vascular leptomeningeal cells, and DA neurons are the main component of the DAergic grafts, originating from a common progenitor that is tripotent at the time of transplantation. [ABSTRACT FROM AUTHOR]

During focal ischemia, neurons can use lactate as an alternative source of energy through its oxidation into pyruvate by the lactate dehydrogenase (LDH). After cardiac arrest, the neurological consequences of this phenomenon are unknown. Experimental study. Experimental laboratory. Male New-Zealand rabbits. Animals were surgically instrumented and randomly divided into five groups receiving short infusion duration of either lactate or pyruvate or a pre-cardiac arrest infusion of oxamate (an inhibitor of the lactate dehydrogenase) or injection of fluorocitrate (an inhibitor of astrocytic tricarboxylic acid), or Saline (lactate, pyruvate, Oxa, FC and Control groups, respectively). After randomization, animals were submitted to 10 min of ventricular fibrillation and subsequent resuscitation. All animals were then either followed during 4 h, for the evaluation of the cerebral net uptake and concentrations of metabolites by microdialysis (n = 6 in each experimental group, n = 12 in control group), or during 48 h for the evaluation of their neurological outcome (n = 7 in each groups and n = 14 in control group). Cardiac arrest was associated with a dramatic increase in cerebral net uptake of lactate during 120 min after resuscitation, which was increased by lactate or pyruvate administration. This was associated with an increase in the mean neurological dysfunction score (66.7 ± 4.7, 79.0 ± 4.5 vs 57.7 ± 1.5 in Lactate, Pyruvate and Control group respectively) at 48 h after cardiac arrest. Oxamate and FC administration were associated with a lower lactate cerebral uptake after cardiac arrest and with an improvement of the neurological recovery (28.85 ± 9.4, 23.86 ± 6.2 vs 57.7 ± 1.5 in Oxa, FC and Control group respectively). After cardiac arrest, immediate isotonic lactate or pyruvate administration is deleterious. Pre-cardiac arrest LDH inhibition was potently neuroprotective in this setting. [ABSTRACT FROM AUTHOR]

The role of long non-coding RNAs (lncRNAs) in malignant cell transformation remains elusive. We previously identified an enhancer-associated lncRNA, LINC01116 (named HOXDeRNA), as a transformative factor converting human astrocytes into glioma-like cells. Employing a combination of CRISPR editing, chromatin isolation by RNA purification coupled with sequencing (ChIRP-seq), in situ mapping RNA-genome interactions (iMARGI), chromatin immunoprecipitation sequencing (ChIP-seq), HiC, and RNA/DNA FISH, we found that HOXDeRNA directly binds to CpG islands within the promoters of 35 glioma-specific transcription factors (TFs) distributed throughout the genome, including key stem cell TFs SOX2, OLIG2, POU3F2, and ASCL1, liberating them from PRC2 repression. This process requires a distinct RNA quadruplex structure and other segments of HOXDeRNA, interacting with EZH2 and CpGs, respectively. Subsequent transformation activates multiple oncogenes (e.g., EGFR, miR-21, and WEE1), driven by the SOX2- and OLIG2-dependent glioma-specific super enhancers. These results help reconstruct the sequence of events underlying the process of astrocyte transformation, highlighting HOXDeRNA's central genome-wide activity and suggesting a shared RNA-dependent mechanism in otherwise heterogeneous and multifactorial gliomagenesis. [Display omitted] • HOXDeRNA transforms astrocytes into glioma-like stem cells • HOXDeRNA directly binds to PRC2-covered CpG islands in the promoters of glioma driver genes • HOXDeRNA removes EZH2 through its RNA quadruplex sequences Deforzh et al. demonstrate the ability of the lncRNA HOXDeRNA to transform astrocytes into glioma-like stem cells by sequestering and protecting key glioma genes from Polycomb2 repression in an RNA quadruplex-dependent manner. [ABSTRACT FROM AUTHOR]

Astrocytes are essential for maintaining brain homeostasis. Alterations in their activity have been associated with various brain pathologies. Sex differences were reported to affect astrocyte development and activity, and even susceptibility to different neurodegenerative diseases. This review aims to summarize the current knowledge on the effects of sex on astrocyte activity in health and disease. [ABSTRACT FROM AUTHOR]

A subpopulation of astrocytes on the brain's surface, known as subpial astrocytes, constitutes the "glia limitans superficialis" (GLS), which is an interface between the brain parenchyma and the cerebrospinal fluid (CSF) in the subpial space. Changes in connexin-43 (Cx43) and aquaporin-4 (AQP4) proteins in subpial astrocytes were examined in the medial prefrontal cortex at postoperative day 1, 3, 7, 14, and 21 after sham operation and sciatic nerve compression (SNC). In addition, we tested the altered uptake of TRITC-conjugated 3 kDa dextran by reactive subpial astrocytes. Cellular immunofluorescence (IF) detection and image analysis were used to examine changes in Cx43 and AQP4 protein levels, as well as TRITC-conjugated 3 kDa dextran, in subpial astrocytes. The intensity of Cx43-IF was significantly increased, but AQP4-IF decreased in subpial astrocytes of sham- and SNC-operated rats during all survival periods compared to naïve controls. Similarly, the uptake of 3 kDa dextran in the GLS was reduced following both sham and SNC operations. The results suggest that both sciatic nerve injury and peripheral tissue injury alone can induce changes in subpial astrocytes related to the spread of their reactivity across the cortical surface mediated by increased amounts of gap junctions. At the same time, water transport and solute uptake were impaired in subpial astrocytes. [ABSTRACT FROM AUTHOR]

The mechanisms of anxiety disorders, the most common mental illness, remain incompletely characterized. The ventral hippocampus (vHPC) is critical for the expression of anxiety. However, current studies primarily focus on vHPC neurons, leaving the role for vHPC astrocytes in anxiety largely unexplored. Here, genetically encoded Ca2+ indicator GCaMP6m and in vivo fiber photometry calcium imaging are used to label vHPC astrocytes and monitor their activity, respectively, genetic and chemogenetic approaches to inhibit and activate vHPC astrocytes, respectively, patch‐clamp recordings to measure glutamate currents, and behavioral assays to assess anxiety‐like behaviors. It is found that vHPC astrocytic activity is increased in anxiogenic environments and by 3‐d subacute restraint stress (SRS), a well‐validated mouse model of anxiety disorders. Genetic inhibition of vHPC astrocytes exerts anxiolytic effects on both innate and SRS‐induced anxiety‐related behaviors, whereas hM3Dq‐mediated chemogenetic or SRS‐induced activation of vHPC astrocytes enhances anxiety‐like behaviors, which are reversed by intra‐vHPC application of the ionotropic glutamate N‐methyl‐d‐aspartate receptor antagonists. Furthermore, intra‐vHPC or systemic application of the N‐methyl‐d‐aspartate receptor antagonist memantine, a U.S. FDA‐approved drug for Alzheimer's disease, fully rescues SRS‐induced anxiety‐like behaviors. The findings highlight vHPC astrocytes as critical regulators of stress and anxiety and as potential therapeutic targets for anxiety and anxiety‐related disorders. [ABSTRACT FROM AUTHOR]

Acute liver dysfunction commonly leads to rapid increases in ammonia concentrations in both the serum and the cerebrospinal fluid. These elevations primarily affect brain astrocytes, causing modifications in their structure and function. However, its impact on neurons is not yet fully understood. In this study, we investigated the impact of elevated ammonium chloride levels (NH4Cl, 5 mM) on synaptic transmission onto CA1 pyramidal neurons in mouse organotypic entorhino-hippocampal tissue cultures. We found that acute exposure to NH4Cl reversibly reduced excitatory synaptic transmission and affected CA3-CA1 synapses. Notably, NH4Cl modified astrocytic, but not CA1 pyramidal neuron, passive intrinsic properties. To further explore the role of astrocytes in NH4Clinduced attenuation of synaptic transmission, we used methionine sulfoximine to target glutamine synthetase, a key astrocytic enzyme for ammonia clearance in the central nervous system. Inhibition of glutamine synthetase effectively prevented the downregulation of excitatory synaptic activity, underscoring the significant role of astrocytes in adjusting excitatory synapses during acute ammonia elevation. [ABSTRACT FROM AUTHOR]