Multiple sclerosis: Neuroimmune crosstalk and therapeutic targeting.

Multiple sclerosis (MS) is a chronic inflammatory and degenerative disease of the central nervous system afflicting nearly three million individuals worldwide. Neuroimmune interactions between glial, neural, and immune cells play important roles in MS pathology and offer potential targets for therapeutic intervention. Here, we review underlying risk factors, mechanisms of MS pathogenesis, available disease modifying therapies, and examine the value of emerging technologies, which may address unmet clinical needs and identify novel therapeutic targets.

Cholesterol and matrisome pathways dysregulated in astrocytes and microglia.

The impact of apolipoprotein E ε4 (APOE4), the strongest genetic risk factor for Alzheimer's disease (AD), on human brain cellular function remains unclear. Here, we investigated the effects of APOE4 on brain cell types derived from population and isogenic human induced pluripotent stem cells, post-mortem brain, and APOE targeted replacement mice. Population and isogenic models demonstrate that APOE4 local haplotype, rather than a single risk allele, contributes to risk. Global transcriptomic analyses reveal human-specific, APOE4-driven lipid metabolic dysregulation in astrocytes and microglia. APOE4 enhances de novo cholesterol synthesis despite elevated intracellular cholesterol due to lysosomal cholesterol sequestration in astrocytes. Further, matrisome dysregulation is associated with upregulated chemotaxis, glial activation, and lipid biosynthesis in astrocytes co-cultured with neurons, which recapitulates altered astrocyte matrisome signaling in human brain. Thus, APOE4 initiates glia-specific cell and non-cell autonomous dysregulation that may contribute to increased AD risk.

Astrocytic trans-Differentiation Completes a Multicellular Paracrine Feedback Loop Required for Medulloblastoma Tumor Growth.

The tumor microenvironment (TME) is critical for tumor progression. However, the establishment and function of the TME remain obscure because of its complex cellular composition. Using a mouse genetic system called mosaic analysis with double markers (MADMs), we delineated TME evolution at single-cell resolution in sonic hedgehog (SHH)-activated medulloblastomas that originate from unipotent granule neuron progenitors in the brain. First, we found that astrocytes within the TME (TuAstrocytes) were trans-differentiated from tumor granule neuron precursors (GNPs), which normally never differentiate into astrocytes. Second, we identified that TME-derived IGF1 promotes tumor progression. Third, we uncovered that insulin-like growth factor 1 (IGF1) is produced by tumor-associated microglia in response to interleukin-4 (IL-4) stimulation. Finally, we found that IL-4 is secreted by TuAstrocytes. Collectively, our studies reveal an evolutionary process that produces a multi-lateral network within the TME of medulloblastoma: a fraction of tumor cells trans-differentiate into TuAstrocytes, which, in turn, produce IL-4 that stimulates microglia to produce IGF1 to promote tumor progression.

Glia Accumulate Evidence that Actions Are Futile and Suppress Unsuccessful Behavior.

When a behavior repeatedly fails to achieve its goal, animals often give up and become passive, which can be strategic for preserving energy or regrouping between attempts. It is unknown how the brain identifies behavioral failures and mediates this behavioral-state switch. In larval zebrafish swimming in virtual reality, visual feedback can be withheld so that swim attempts fail to trigger expected visual flow. After tens of seconds of such motor futility, animals became passive for similar durations. Whole-brain calcium imaging revealed noradrenergic neurons that responded specifically to failed swim attempts and radial astrocytes whose calcium levels accumulated with increasing numbers of failed attempts. Using cell ablation and optogenetic or chemogenetic activation, we found that noradrenergic neurons progressively activated brainstem radial astrocytes, which then suppressed swimming. Thus, radial astrocytes perform a computation critical for behavior: they accumulate evidence that current actions are ineffective and consequently drive changes in behavioral states. VIDEO ABSTRACT.

Multi-sensory Gamma Stimulation Ameliorates Alzheimer's-Associated Pathology and Improves Cognition.

We previously reported that inducing gamma oscillations with a non-invasive light flicker (gamma entrainment using sensory stimulus or GENUS) impacted pathology in the visual cortex of Alzheimer's disease mouse models. Here, we designed auditory tone stimulation that drove gamma frequency neural activity in auditory cortex (AC) and hippocampal CA1. Seven days of auditory GENUS improved spatial and recognition memory and reduced amyloid in AC and hippocampus of 5XFAD mice. Changes in activation responses were evident in microglia, astrocytes, and vasculature. Auditory GENUS also reduced phosphorylated tau in the P301S tauopathy model. Furthermore, combined auditory and visual GENUS, but not either alone, produced microglial-clustering responses, and decreased amyloid in medial prefrontal cortex. Whole brain analysis using SHIELD revealed widespread reduction of amyloid plaques throughout neocortex after multi-sensory GENUS. Thus, GENUS can be achieved through multiple sensory modalities with wide-ranging effects across multiple brain areas to improve cognitive function.

Metabolic Control of Astrocyte Pathogenic Activity via cPLA2-MAVS.

Metabolism has been shown to control peripheral immunity, but little is known about its role in central nervous system (CNS) inflammation. Through a combination of proteomic, metabolomic, transcriptomic, and perturbation studies, we found that sphingolipid metabolism in astrocytes triggers the interaction of the C2 domain in cytosolic phospholipase A2 (cPLA2) with the CARD domain in mitochondrial antiviral signaling protein (MAVS), boosting NF-κB-driven transcriptional programs that promote CNS inflammation in experimental autoimmune encephalomyelitis (EAE) and, potentially, multiple sclerosis. cPLA2 recruitment to MAVS also disrupts MAVS-hexokinase 2 (HK2) interactions, decreasing HK enzymatic activity and the production of lactate involved in the metabolic support of neurons. Miglustat, a drug used to treat Gaucher and Niemann-Pick disease, suppresses astrocyte pathogenic activities and ameliorates EAE. Collectively, these findings define a novel immunometabolic mechanism that drives pro-inflammatory astrocyte activities, outlines a new role for MAVS in CNS inflammation, and identifies candidate targets for therapeutic intervention.

Development and Cell Biology of the Blood-Brain Barrier.

The vertebrate vasculature displays high organotypic specialization, with the structure and function of blood vessels catering to the specific needs of each tissue. A unique feature of the central nervous system (CNS) vasculature is the blood-brain barrier (BBB). The BBB regulates substance influx and efflux to maintain a homeostatic environment for proper brain function. Here, we review the development and cell biology of the BBB, focusing on the cellular and molecular regulation of barrier formation and the maintenance of the BBB through adulthood. We summarize unique features of CNS endothelial cells and highlight recent progress in and general principles of barrier regulation. Finally, we illustrate why a mechanistic understanding of the development and maintenance of the BBB could provide novel therapeutic opportunities for CNS drug delivery.

Multitasking: Dual Leucine Zipper-Bearing Kinases in Neuronal Development and Stress Management.

The dual leucine zipper-bearing kinase (DLK) and leucine zipper-bearing kinase (LZK) are evolutionarily conserved MAPKKKs of the mixed-lineage kinase family. Acting upstream of stress-responsive JNK and p38 MAP kinases, DLK and LZK have emerged as central players in neuronal responses to a variety of acute and traumatic injuries. Recent studies also implicate their function in astrocytes, microglia, and other nonneuronal cells, reflecting their expanding roles in the multicellular response to injury and in disease. Of particular note is the potential link of these kinases to neurodegenerative diseases and cancer. It is thus critical to understand the physiological contexts under which these kinases are activated, as well as the signal transduction mechanisms that mediate specific functional outcomes. In this review we first provide a historical overview of the biochemical and functional dissection of these kinases. We then discuss recent findings on regulating their activity to enhance cellular protection following injury and in disease, focusing on but not limited to the nervous system.

Stress induces behavioral abnormalities by increasing expression of phagocytic receptor MERTK in astrocytes to promote synapse phagocytosis.

Childhood neglect and/or abuse can induce mental health conditions with unknown mechanisms. Here, we identified stress hormones as strong inducers of astrocyte-mediated synapse phagocytosis. Using in vitro, in vivo, and human brain organoid experiments, we showed that stress hormones increased the expression of the Mertk phagocytic receptor in astrocytes through glucocorticoid receptor (GR). In post-natal mice, exposure to early social deprivation (ESD) specifically activated the GR-MERTK pathway in astrocytes, but not in microglia. The excitatory post-synaptic density in cortical regions was reduced in ESD mice, and there was an increase in the astrocytic engulfment of these synapses. The loss of excitatory synapses, abnormal neuronal network activities, and behavioral abnormalities in ESD mice were largely prevented by ablating GR or MERTK in astrocytes. Our work reveals the critical roles of astrocytic GR-MERTK activation in evoking stress-induced abnormal behaviors in mice, suggesting GR-MERTK signaling as a therapeutic target for stress-induced mental health conditions.

Therapeutic Potential of PTBP1 Inhibition, If Any, Is Not Attributed to Glia-to-Neuron Conversion.

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.

Astrocytic Insulin Signaling Couples Brain Glucose Uptake with Nutrient Availability.

We report that astrocytic insulin signaling co-regulates hypothalamic glucose sensing and systemic glucose metabolism. Postnatal ablation of insulin receptors (IRs) in glial fibrillary acidic protein (GFAP)-expressing cells affects hypothalamic astrocyte morphology, mitochondrial function, and circuit connectivity. Accordingly, astrocytic IR ablation reduces glucose-induced activation of hypothalamic pro-opio-melanocortin (POMC) neurons and impairs physiological responses to changes in glucose availability. Hypothalamus-specific knockout of astrocytic IRs, as well as postnatal ablation by targeting glutamate aspartate transporter (GLAST)-expressing cells, replicates such alterations. A normal response to altering directly CNS glucose levels in mice lacking astrocytic IRs indicates a role in glucose transport across the blood-brain barrier (BBB). This was confirmed in vivo in GFAP-IR KO mice by using positron emission tomography and glucose monitoring in cerebral spinal fluid. We conclude that insulin signaling in hypothalamic astrocytes co-controls CNS glucose sensing and systemic glucose metabolism via regulation of glucose uptake across the BBB.

Astrocyte-immune cell interactions in physiology and pathology.

Astrocytes play both physiological and pathological roles in maintaining central nervous system (CNS) function. Here, we review the varied functions of astrocytes and how these might change in subsets of reactive astrocytes. We review the current understanding of astrocyte interactions with microglia and the vasculature and protective barriers in the central nervous system as well as highlight recent insights into physiologic and reactive astrocyte sub-states identified by transcriptional profiling. Our goal is to stimulate inquiry into how these molecular identifiers link to specific functional changes in astrocytes and to define the implications of these heterogeneous molecular and functional changes in brain function and pathology. Defining these complex interactions has the potential to yield new therapies in CNS injury, infection, and disease.

Dectin-1 limits autoimmune neuroinflammation and promotes myeloid cell-astrocyte crosstalk via Card9-independent expression of Oncostatin M.

Pathologic roles of innate immunity in neurologic disorders are well described, but their beneficial aspects are less understood. Dectin-1, a C-type lectin receptor (CLR), is largely known to induce inflammation. Here, we report that Dectin-1 limited experimental autoimmune encephalomyelitis (EAE), while its downstream signaling molecule, Card9, promoted the disease. Myeloid cells mediated the pro-resolution function of Dectin-1 in EAE with enhanced gene expression of the neuroprotective molecule, Oncostatin M (Osm), through a Card9-independent pathway, mediated by the transcription factor NFAT. Furthermore, we find that the Osm receptor (OsmR) functioned specifically in astrocytes to reduce EAE severity. Notably, Dectin-1 did not respond to heat-killed Mycobacteria, an adjuvant to induce EAE. Instead, endogenous Dectin-1 ligands, including galectin-9, in the central nervous system (CNS) were involved to limit EAE. Our study reveals a mechanism of beneficial myeloid cell-astrocyte crosstalk regulated by a Dectin-1 pathway and identifies potential therapeutic targets for autoimmune neuroinflammation.

Astrocytes and Behavior.

Animal behavior was classically considered to be determined exclusively by neuronal activity, whereas surrounding glial cells such as astrocytes played only supportive roles. However, astrocytes are as numerous as neurons in the mammalian brain, and current findings indicate a chemically based dialog between astrocytes and neurons. Activation of astrocytes by synaptically released neurotransmitters converges on regulating intracellular Ca2+ in astrocytes, which then can regulate the efficacy of near and distant tripartite synapses at diverse timescales through gliotransmitter release. Here, we discuss recent evidence on how diverse behaviors are impacted by this dialog. These recent findings support a paradigm shift in neuroscience, in which animal behavior does not result exclusively from neuronal activity but from the coordinated activity of both astrocytes and neurons. Decoding how astrocytes and neurons interact with each other in various brain circuits will be fundamental to fully understanding how behaviors originate and become dysregulated in disease.

Brain Repair by Cell Replacement via In Situ Neuronal Reprogramming.

Neurodegenerative diseases, characterized by progressive neural loss, have been some of the most challenging medical problems in aging societies. Treatment strategies such as symptom management have little impact on disease progression, while intervention with specific disease mechanisms may only slow down disease progression. One therapeutic strategy that has the potential to reverse the disease phenotype is to replenish neurons and rebuild the pathway lost to degeneration. Although it is generally believed that the central nervous system has lost the capability to regenerate, increasing evidence indicates that the brain is more plastic than previously thought, containing perhaps the biggest repertoire of cells with latent neurogenic programs in the body. This review focuses on key advances in generating new neurons through in situ neuronal reprogramming, which is tied to fundamental questions regarding adult neurogenesis, cell source, and mechanisms for neuronal reprogramming, as well as the ability of new neurons to integrate into the existing circuitry.

Metabolic Recruitment in Brain Tissue.

Information processing imposes urgent metabolic demands on neurons, which have negligible energy stores and restricted access to fuel. Here, we discuss metabolic recruitment, the tissue-level phenomenon whereby active neurons harvest resources from their surroundings. The primary event is the neuronal release of K+ that mirrors workload. Astrocytes sense K+ in exquisite fashion thanks to their unique coexpression of NBCe1 and α2ß2 Na+/K+ ATPase, and within seconds switch to Crabtree metabolism, involving GLUT1, aerobic glycolysis, transient suppression of mitochondrial respiration, and lactate export. The lactate surge serves as a secondary recruiter by inhibiting glucose consumption in distant cells. Additional recruiters are glutamate, nitric oxide, and ammonium, which signal over different spatiotemporal domains. The net outcome of these events is that more glucose, lactate, and oxygen are made available. Metabolic recruitment works alongside neurovascular coupling and various averaging strategies to support the inordinate dynamic range of individual neurons.

Astrogenesis in the murine dentate gyrus is a life-long and dynamic process.

Astrocytes are highly abundant in the mammalian brain, and their functions are of vital importance for all aspects of development, adaption, and aging of the central nervous system (CNS). Mounting evidence indicates the important contributions of astrocytes to a wide range of neuropathies. Still, our understanding of astrocyte development significantly lags behind that of other CNS cells. We here combine immunohistochemical approaches with genetic fate-mapping, behavioural paradigms, single-cell transcriptomics, and in vivo two-photon imaging, to comprehensively assess the generation and the proliferation of astrocytes in the dentate gyrus (DG) across the life span of a mouse. Astrogenesis in the DG is initiated by radial glia-like neural stem cells giving rise to locally dividing astrocytes that enlarge the astrocyte compartment in an outside-in-pattern. Also in the adult DG, the vast majority of astrogenesis is mediated through the proliferation of local astrocytes. Interestingly, locally dividing astrocytes were able to adapt their proliferation to environmental and behavioral stimuli revealing an unexpected plasticity. Our study establishes astrocytes as enduring plastic elements in DG circuits, implicating a vital contribution of astrocyte dynamics to hippocampal plasticity.

YAP and TAZ differentially regulate postnatal cortical progenitor proliferation and astrocyte differentiation.

WW domain-containing transcription regulator 1 (WWTR1, referred to here as TAZ) and Yes-associated protein (YAP, also known as YAP1) are transcriptional co-activators traditionally studied together as a part of the Hippo pathway, and are best known for their roles in stem cell proliferation and differentiation. Despite their similarities, TAZ and YAP can exert divergent cellular effects by differentially interacting with other signaling pathways that regulate stem cell maintenance or differentiation. In this study, we show in mouse neural stem and progenitor cells (NPCs) that TAZ regulates astrocytic differentiation and maturation, and that TAZ mediates some, but not all, of the effects of bone morphogenetic protein (BMP) signaling on astrocytic development. By contrast, both TAZ and YAP mediate the effects on NPC fate of ß1-integrin (ITGB1) and integrin-linked kinase signaling, and these effects are dependent on extracellular matrix cues. These findings demonstrate that TAZ and YAP perform divergent functions in the regulation of astrocyte differentiation, where YAP regulates cell cycle states of astrocytic progenitors and TAZ regulates differentiation and maturation from astrocytic progenitors into astrocytes.

YAP and TAZ differentially regulate postnatal cortical progenitor proliferation and astrocyte differentiation.

WW domain-containing transcription regulator 1 (WWTR1, referred to here as TAZ) and Yes-associated protein (YAP, also known as YAP1) are transcriptional co-activators traditionally studied together as a part of the Hippo pathway, and are best known for their roles in stem cell proliferation and differentiation. Despite their similarities, TAZ and YAP can exert divergent cellular effects by differentially interacting with other signaling pathways that regulate stem cell maintenance or differentiation. In this study, we show in mouse neural stem and progenitor cells (NPCs) that TAZ regulates astrocytic differentiation and maturation, and that TAZ mediates some, but not all, of the effects of bone morphogenetic protein (BMP) signaling on astrocytic development. By contrast, both TAZ and YAP mediate the effects on NPC fate of ß1-integrin (ITGB1) and integrin-linked kinase signaling, and these effects are dependent on extracellular matrix cues. These findings demonstrate that TAZ and YAP perform divergent functions in the regulation of astrocyte differentiation, where YAP regulates cell cycle states of astrocytic progenitors and TAZ regulates differentiation and maturation from astrocytic progenitors into astrocytes.

Microglia refine developing retinal astrocytic and vascular networks through the complement C3/C3aR axis.

Microglia, a resident immune cell of the central nervous system (CNS), play a pivotal role in facilitating neurovascular development through mechanisms that are not fully understood. Previous reports indicate a role for microglia in regulating astrocyte density. This current work resolves the mechanism through which microglia facilitate astrocyte spatial patterning and superficial vascular bed formation in the neuroretina during development. Ablation of microglia increased astrocyte density and altered spatial patterning. Mechanistically, we show that microglia regulate the formation of the spatially organized astrocyte template required for subsequent vascular growth, through the complement C3/C3aR axis during neuroretinal development. Lack of C3 or C3aR hindered the developmental phagocytic removal of astrocyte bodies and resulted in increased astrocyte density. In addition, increased astrocyte density was associated with elevated proangiogenic extracellular matrix gene expression in C3- and C3aR-deficient retinas, resulting in increased vascular density. These data demonstrate that microglia regulate developmental astrocyte and vascular network spatial patterning in the neuroretina via the complement axis.