Ablation Of Microglia Research Articles

Sphingosine-1-phosphate signaling regulates the ability of Müller glia to become neurogenic, proliferating progenitor-like cells.

The purpose of these studies is to investigate how Sphingosine-1-phosphate (S1P) signaling regulates glial phenotype, dedifferentiation of Müller glia (MG), reprogramming into proliferating MG-derived progenitor cells (MGPCs), and neuronal differentiation of the progeny of MGPCs. We found that S1P-related genes are highly expressed by retinal neurons and glia, and levels of expression were dynamically regulated following retinal damage. S1PR1 is highly expressed by resting MG and is rapidly downregulated following acute retinal damage. Drug treatments that activate S1PR1 or increase levels of S1P suppressed the formation of MGPCs, whereas treatments that inhibit S1PR1 or decreased levels of S1P stimulated the formation of MGPCs. Inhibition of S1PR1 or SPHK1 significantly enhanced the neuronal differentiation of the progeny of MGPCs. Further, ablation of microglia from the retina, wherein the formation of MGPCs in damaged retinas is impaired, has a significant impact upon expression patterns of S1P-related genes in MG. Inhibition of S1PR1 and SPHK1 partially rescued the formation of MGPCs in damaged retinas missing microglia. Finally, we show that TGFβ/Smad3 signaling in the resting retina maintains S1PR1 expression in MG. We conclude that the S1P signaling is dynamically regulated in MG and MGPCs and activation of S1P signaling depends, in part, on signals produced by reactive microglia.

The functional role of locus coeruleus microglia in the female stress response.

Neuropsychiatric disorders that result from stress exposure are highly associated with central inflammation. Our previous work established that females selectively exhibit heightened proinflammatory cytokine production within the noradrenergic locus coeruleus (LC) along with a hypervigilant behavioral phenotype in response to witnessing social stress, and ablation of microglia using pharmacological techniques prevents this behavioral response. These studies were designed to further investigate the impact of stress-induced neuroimmune signaling on the long-term behavioral and neuronal consequences of social stress exposure in females using chemogenetics. We first characterized the use of an AAV-CD68-Gi-DREADD virus targeted to microglia within the LC and confirmed viral transduction, selectivity, and efficacy. Clozapine-n-oxide (CNO) was used for the suppression of microglial reactivity during acute and chronic exposure to vicarious/witness social defeat in female rats. Chemogenetic-mediated inhibition of microglial reactivity during stress blunted the neuroimmune response to stress and prevented both acute and long-term hypervigilant behavioral responses. Further, a history of microglial suppression during stress prevented the heightened LC activity typically observed in response to stress cues. These studies are among the first to use a chemogenetic approach to inhibit microglia within the female brain in vivo and establish LC inflammation as a key mechanism underlying the behavioral and neuronal responses to social stress in females.

Gut microbiota-derived metabolite trimethylamine N-oxide aggravates cognitive dysfunction induced by femoral fracture operation in mice.

An increasing number of elderly individuals are experiencing postoperative cognitive dysfunction (POCD) problems after undergoing hip replacement surgery, with gut microbiota metabolites playing a role in its pathogenesis. Among these, the specific effects of trimethylamine N-oxide (TMAO) on POCD are still unclear. This study aimed to explore the role of TMAO on cognitive dysfunction and underlying mechanisms in mice. The POCD model was created through femoral fracture surgery in elderly mice, followed by cognitive function assessments using the Morris Water Maze and Novel Object Recognition tests. The gut microbiota depletion and fecal microbiota transplantation were performed to examine the relationship between TMAO levels and cognitive outcomes. The effects of TMAO treatment on cognitive dysfunction, microglial activation, and inflammatory cytokine levels in the brain were also evaluated, with additional assessment of the role of microglial ablation in reducing TMAO-induced cognitive impairment. Elevated TMAO levels were found to be associated with cognitive decline in mice following femoral fracture surgery, with gut microbiota depletion mitigating both TMAO elevation and cognitive dysfunction. In contrast, fecal microbiota transplantation from postoperative mice resulted in accelerated cognitive dysfunction and TMAO accumulation in germ-free mice. Furthermore, TMAO treatment worsened cognitive deficits, neuroinflammation, and promoted microglial activation, which were reversed through the ablation of microglia. TMAO exacerbates cognitive dysfunction and neuroinflammation in POCD mice, with microglial activation playing a crucial role in this process. Our findings may provide new therapeutic strategies for managing TMAO-related POCD and improving the quality of life for elderly patients.

Microglial over-pruning of synapses during development in autism-associated SCN2A-deficient mice and human cerebral organoids.

Autism spectrum disorder (ASD) is a major neurodevelopmental disorder affecting 1 in 36 children in the United States. While neurons have been the focus of understanding ASD, an altered neuro-immune response in the brain may be closely associated with ASD, and a neuro-immune interaction could play a role in the disease progression. As the resident immune cells of the brain, microglia regulate brain development and homeostasis via core functions including phagocytosis of synapses. While ASD has been traditionally considered a polygenic disorder, recent large-scale human genetic studies have identified SCN2A deficiency as a leading monogenic cause of ASD and intellectual disability. We generated a Scn2a-deficient mouse model, which displays major behavioral and neuronal phenotypes. However, the role of microglia in this disease model is unknown. Here, we reported that Scn2a-deficient mice have impaired learning and memory, accompanied by reduced synaptic transmission and lower spine density in neurons of the hippocampus. Microglia in Scn2a-deficient mice are partially activated, exerting excessive phagocytic pruning of post-synapses related to the complement C3 cascades during selective developmental stages. The ablation of microglia using PLX3397 partially restores synaptic transmission and spine density. To extend our findings from rodents to human cells, we established a microglia-incorporated human cerebral organoid model carrying an SCN2A protein-truncating mutation identified in children with ASD. We found that human microglia display increased elimination of post-synapse in cerebral organoids carrying the SCN2A mutation. Our study establishes a key role of microglia in multi-species autism-associated models of SCN2A deficiency from mouse to human cells.

Visualization, Fate Mapping, Ablation, and Mutagenesis of Microglia in the Mouse Brain.

Since the classical studies of Pío del Río-Hortega, microglia research has come a long way. In particular, recent advances in bulk and single-cell (sc) transcriptomics have yielded many fascinating new insights into these intriguing immune cells at the interface with the central nervous system (CNS), both in small animal models and human samples. In parallel, tools developed by advanced mouse genetics have revealed the unique ontogeny of microglia and their striking dynamic interactions with other cells in the brain parenchyma. In this chapter, we will discuss various applications of the Cre/loxP-based approach that have enabled the study of microglia in their physiological context of the mouse brain. We will highlight selected key findings that have shaped our current understanding of these cells and discuss the technical intricacies of the Cre/loxP approach and some remaining challenges.

Ablation of microglia following infection of the central nervous system with a neurotropic murine coronavirus infection leads to increased demyelination and impaired remyelination

Intracranial inoculation of susceptible mice with a glial-tropic strain of mouse hepatitis virus (JHMV), a murine coronavirus, results in an acute encephalomyelitis followed by viral persistence in white matter tracts accompanied by chronic neuroinflammation and demyelination. Microglia are the resident immune cell of the central nervous system (CNS) and are considered important in regulating events associated with neuroinflammation as well as influencing both white matter damage and remyelination. To better understand mechanisms by which microglia contribute to these immune-mediated events, JHMV-infected mice with established demyelination were treated with the small molecular inhibitor of colony stimulating factor 1 receptor (CSF1R), PLX5622, to deplete microglia. Treatment with PLX5622 did not affect viral replication within the CNS yet the severity of demyelination was increased and remyelination impaired compared to control mice. Gene expression analysis revealed that targeting microglia resulted in altered expression of genes associated with immune cell activation and phagocytosis of myelin debris. These findings indicate that microglia are not critical in viral surveillance in persistently JHMV-infected mice yet restrict white matter damage and remyelination, in part, by influencing phagocytosis of myelin debris.

Melatonin protects against NMDA-induced retinal ganglion cell injury by regulating the microglia-TNFα-RGC p38 MAPK pathway.

Glaucoma, one of the most common ocular neurodegenerative diseases worldwide, is characterized by retinal ganglion cell (RGC) loss. There is a large body of literature that describes the neuroprotective role of melatonin against neurodegenerative diseases by regulating neuroinflammation, although the exact mechanism through which melatonin acts on RGC is still uncertain. This study assessed the protective effects of melatonin using a NMDA-induced RGC injury model, and studied the possible mechanisms involved in this process. Melatonin promoted RGC survival, improved retinal function, and inhibited the apoptosis and necrosis of retinal cells. To understand the mechanism of the neuroprotective effects of melatonin on RGC, microglia and inflammation-related pathways were assessed after melatonin administration and microglia ablation. Melatonin promoted RGC survival by suppressing microglia-derived proinflammatory cytokines, in particular TNFα, which in turn inhibited the activation of p38 MAPK pathway. Inhibiting TNFα or manipulating p38 MAPK pathway protected damaged RGC. Our results suggest that melatonin protects against NMDA-induced RGC injury by inhibiting the microglial TNFα-RGC p38 MAPK pathway. It should be considered a candidate neuroprotective therapy against retinal neurodegenerative diseases.

Microglia Maintain Homeostatic Conditions in the Developing Rostral Migratory Stream (P10-9.005)

<h3>Objective:</h3> This work examines how microglia maintain the homeostatic conditions permissive to neuroblast migration in the rostral migratory stream (RMS) during the early postnatal period. <h3>Background:</h3> Microglia phagocytosis is essential during critical periods of development to clear the results of over-exuberant neurogenesis, including defective and apoptotic cells. The importance of microglia in mediating homeostasis during neurodevelopment was underscored by the case of a child born without microglia, who exhibited neuronal heterotopias and ventriculomegaly (Oosterhof et al., 2019). <h3>Design/Methods:</h3> To illustrate microglia phagocytosis of neuroblasts in the postnatal RMS, P7 <i>CX3CR-1</i><i>^GFP/+</i><i>;DCX</i><i>^DsRed/+</i> mice were stained with primary antibodies against CD68, CLEC7A, and MERTK. Fluorescent liposomes were injected into the cerebral lateral ventricles of P1 <i>CX3CR-1</i><i>^GFP/+</i> mice to demonstrate microglia phagocytic capacity. To selectively deplete microglia <i>in vivo, Cx3cr1</i><i>^CreER/+</i><i>; ROSA26</i><i>^eGFP-DTA/+</i> mice were injected with tamoxifen i.p. at P0 and were either sacrificed at P3 or injected with tamoxifen every 4 days until sacrifice at P14. To label microglia, neuroblasts and apoptotic cells, immunohistochemistry was performed with primary antibodies against Iba1, doublecortin (DCX) and cleaved caspase-3 (CC3), respectively. <h3>Results:</h3> Microglia express phagocytic markers, engulf extracellular material and “wrap” migrating neuroblasts in the early postnatal RMS. Microglia depletion was associated with a significant increase in the density of CC3+ apoptotic neuroblasts in the RMS start and RMS descent at both P3 and P14. All five mice that expressed the DTA subunit showed patent olfactory ventricles following 14 days of tamoxifen injection. <h3>Conclusions:</h3> Microglia expressing phagocytic markers closely interact with migrating neuroblasts in the developing RMS, suggesting phagocytosis of neuroblasts. Microglia ablation compromised the homeostasis of the early postnatal RMS, causing an accumulation of apoptotic neuroblasts that broadened the RMS domain and an extended patency of the lateral ventricle extension. Microglia may thus regulate neuroblast migratory corridors through the elimination of surplus neuroblasts and phagocytosis of apoptotic debris. <b>Disclosure:</b> Dr. Meller has received research support from National Institutes of Health Grant NIDCD F31DC018469-01. The institution of Prof. GREER has received research support from NIH.

Microglia Maintain Homeostatic Conditions in the Developing Rostral Migratory Stream.

Microglia invade the neuroblast migratory corridor of the rostral migratory stream (RMS) early in development. The early postnatal RMS does not yet have the dense astrocyte and vascular scaffold that helps propel forward migrating neuroblasts, which led us to consider whether microglia help regulate conditions permissive to neuroblast migration in the RMS. GFP-labeled microglia in CX3CR-1GFP/+ mice assemble primarily along the outer borders of the RMS during the first postnatal week, where they exhibit predominantly an ameboid morphology and associate with migrating neuroblasts. Microglia ablation for 3 d postnatally does not impact the density of pulse labeled BrdU+ neuroblasts nor the distance migrated by tdTomato electroporated neuroblasts in the RMS. However, microglia wrap DsRed-labeled neuroblasts in the RMS of P7 CX3CR-1GFP/+;DCXDsRed/+ mice and express the markers CD68, CLEC7A, MERTK, and IGF-1, suggesting active regulation in the developing RMS. Microglia depletion for 14 d postnatally further induced an accumulation of CC3+ DCX+ apoptotic neuroblasts in the RMS, a wider RMS and extended patency of the lateral ventricle extension in the olfactory bulb. These findings illustrate the importance of microglia in maintaining a healthy neuroblast population and an environment permissive to neuroblast migration in the early postnatal RMS.

Microglia depletion exacerbates retinal ganglion cell loss in a mouse model of glaucoma.

To test whether depletion of microglia in the optic nerve head has a beneficial effect on retinal ganglion cell numbers and function, we depleted microglia by oral administration of the CSF1R antagonist PLX5622. Then, ocular hypertension was induced by unilateral injection of magnetic microbeads into the anterior chamber. Visual function was assessed with pattern electroretinography and measurement of the optomotor reflex. Retinal ganglion cell bodies and axons were counted and gene expression patterns in optic nerve head astrocytes were tested on freshly dissociated astrocytes. PLX5622 efficiently depleted microglia in the retina and the optic nerve head, but about 20% of microglia persisted in the myelinated optic nerve proper even after prolonged exposure to the drug. PLX5622 did not affect ganglion cell function by itself. Elevation of the IOP for four weeks led to the expected decrease in visual acuity and pattern ERG amplitude. Microglia ablation did not affect these parameters. Ganglion cell and axon numbers were counted histologically post mortem. Mice in the microglia depletion group showed a moderate but significantly greater loss of ganglion cells than the control group. At four weeks post microbead injection, gene expression patterns in optic nerve head astrocytes are consistent with an A2 (or neuroprotective) pattern. Microglia depletion blunted the up-regulation of A2 genes in astrocytes. In conclusion, microglia depletion is unlikely to protect retinal ganglion cells in early glaucoma.

Microglia-derived PDGFB promotes neuronal potassium currents to suppress basal sympathetic tonicity and limit hypertension

Although many studies have addressed the regulatory circuits affecting neuronal activities, local non-synaptic mechanisms that determine neuronal excitability remain unclear. Here, we found that microglia prevented overactivation of pre-sympathetic neurons in the hypothalamic paraventricular nucleus (PVN) at steady state. Microglia constitutively released platelet-derived growth factor (PDGF) B, which signaled via PDGFRα on neuronal cells and promoted their expression of Kv4.3, a key subunit that conducts potassium currents. Ablation of microglia, conditional deletion of microglial PDGFB, or suppression of neuronal PDGFRα expression in the PVN elevated the excitability of pre-sympathetic neurons and sympathetic outflow, resulting in a profound autonomic dysfunction. Disruption of the PDGFBMG-Kv4.3Neuron pathway predisposed mice to develop hypertension, whereas central supplementation of exogenous PDGFB suppressed pressor response when mice were under hypertensive insult. Our results point to a non-immune action of resident microglia in maintaining the balance of sympathetic outflow, which is important in preventing cardiovascular diseases.

Reduced microglia activation following metformin administration or microglia ablation is sufficient to prevent functional deficits in a mouse model of neonatal stroke

BackgroundNeonatal stroke is a devastating insult that can lead to life-long impairments. In response to hypoxic–ischaemic injury, there is loss of neurons and glia as well as a neuroinflammatory response mediated by resident immune cells, including microglia and astrocytes, which can exacerbate damage. Administration of the antidiabetic drug metformin has been shown to improve functional outcomes in preclinical models of brain injury and the cellular basis for metformin-mediated recovery is unknown. Given metformin’s demonstrated anti-inflammatory properties, we investigated its role in regulating the microglia activation and used a microglia ablation strategy to investigate the microglia-mediated outcomes in a mouse model of neonatal stroke.MethodsHypoxia-ischaemia (H-I) was performed on post-natal day 8. Metformin was administered for one week, starting one day after injury. Immunohistochemistry was used to examine the spatiotemporal response of microglia and astrocytes after hypoxia-ischaemia, with or without metformin treatment. To evaluate the effects of microglia depletion after hypoxia-ischaemia, we delivered Plexxikon 5622 for 1 or 2 weeks post-injury. The regional pattern of microglia and astrocyte depletion was assessed through immunohistochemistry. Motor behaviour was assessed with the righting reflex, hindlimb suspension, grip strength and cylinder tests.ResultsHerein, we revealed a spatiotemporally regulated response of microglia and astrocytes after hypoxia-ischaemia. Metformin treatment after hypoxia-ischaemia had no effect on microglia number and proliferation, but significantly reduced microglia activation in all regions examined, concomitant with improved behavioural outcomes in injured mice. Plexxikon 5622 treatment successfully ablated microglia, resulting in a > 90% depletion in microglia in the neonatal brain. Microglia rapidly repopulated upon treatment cessation of Plexxikon. Most interesting, microglia ablation was sufficient to reduce functional deficits after hypoxia-ischaemia, mimicking the effects of 1 week of metformin treatment post-injury.ConclusionThese results highlight the importance of regulating the neuroinflammatory response after neonatal stroke to promote recovery.

NFkB-signaling promotes glial reactivity and suppresses Müller glia-mediated neuron regeneration in the mammalian retina.

Müller glia (MG) in mammalian retinas are incapable of regenerating neurons after damage, whereas the MG in lower vertebrates regenerate functional neurons. Identification of cell signaling pathways and gene regulatory networks that regulate MG-mediated regeneration is key to harnessing the regenerative potential of MG. Here, we study how NFkB-signaling influences glial responses to damage and reprogramming of MG into neurons in the rodent retina. We find activation of NFkB and dynamic expression of NFkB-associated genes in MG after damage, however damage-induced NFkB activation is inhibited by microglia ablation. Knockout of NFkB in MG suppressed the accumulation of immune cells after damage. Inhibition of NFkB following NMDA-damage significantly enhanced the reprogramming of Ascl1-overexpressing MG into neuron-like cells. scRNA-seq of retinal glia following inhibition of NFkB reveals coordination with signaling via TGFβ2 and suppression of NFI and Id transcription factors. Inhibition of Smad3 signal transducer or Id transcription factors increased numbers of neuron-like cells produced by Ascl1-overexpressing MG. We conclude that NFkB is a key signaling hub that is activated in MG after damage, mediates the accumulation of immune cells, and suppresses the neurogenic potential of MG.

Innate Immune Pathways Promote Oligodendrocyte Progenitor Cell Recruitment to the Injury Site in Adult Zebrafish Brain.

The oligodendrocyte progenitors (OPCs) are at the front of the glial reaction to the traumatic brain injury. However, regulatory pathways steering the OPC reaction as well as the role of reactive OPCs remain largely unknown. Here, we compared a long-lasting, exacerbated reaction of OPCs to the adult zebrafish brain injury with a timely restricted OPC activation to identify the specific molecular mechanisms regulating OPC reactivity and their contribution to regeneration. We demonstrated that the influx of the cerebrospinal fluid into the brain parenchyma after injury simultaneously activates the toll-like receptor 2 (Tlr2) and the chemokine receptor 3 (Cxcr3) innate immunity pathways, leading to increased OPC proliferation and thereby exacerbated glial reactivity. These pathways were critical for long-lasting OPC accumulation even after the ablation of microglia and infiltrating monocytes. Importantly, interference with the Tlr1/2 and Cxcr3 pathways after injury alleviated reactive gliosis, increased new neuron recruitment, and improved tissue restoration.

Diphtheria toxin induced but not CSF1R inhibitor mediated microglia ablation model leads to the loss of CSF/ventricular spaces in vivo that is independent of cytokine upregulation

BackgroundTwo recently developed novel rodent models have been reported to ablate microglia, either by genetically targeting microglia (via Cx3cr1-creER: iDTR + Dtx) or through pharmacologically targeting the CSF1R receptor with its inhibitor (PLX5622). Both models have been widely used in recent years to define essential functions of microglia and have led to high impact studies that have moved the field forward.MethodsUsing either Cx3cr1-iDTR mice in combination with Dtx or via the PLX5622 diet to pharmacologically ablate microglia, we compared the two models via MRI and histology to study the general anatomy of the brain and the CSF/ventricular systems. Additionally, we analyzed the cytokine profile in both microglia ablation models.ResultsWe discovered that the genetic ablation (Cx3cr1-iDTR + Dtx), but not the pharmacological microglia ablation (PLX5622), displays a surprisingly rapid pathological condition in the brain represented by loss of CSF/ventricles without brain parenchymal swelling. This phenotype was observed both in MRI and histological analysis. To our surprise, we discovered that the iDTR allele alone leads to the loss of CSF/ventricles phenotype following diphtheria toxin (Dtx) treatment independent of cre expression. To examine the underlying mechanism for the loss of CSF in the Cx3cr1-iDTR ablation and iDTR models, we additionally investigated the cytokine profile in the Cx3cr1-iDTR + Dtx, iDTR + Dtx and the PLX models. We found increases of multiple cytokines in the Cx3cr1-iDTR + Dtx but not in the pharmacological ablation model nor the iDTR + Dtx mouse brains at the time of CSF loss (3 days after the first Dtx injection). This result suggests that the upregulation of cytokines is not the cause of the loss of CSF, which is supported by our data indicating that brain parenchyma swelling, or edema are not observed in the Cx3cr1-iDTR + Dtx microglia ablation model. Additionally, pharmacological inhibition of the KC/CXCR2 pathway (the most upregulated cytokine in the Cx3cr1-iDTR + Dtx model) did not resolve the CSF/ventricular loss phenotype in the genetic microglia ablation model. Instead, both the Cx3cr1-iDTR + Dtx ablation and iDTR + Dtx models showed increased activated IBA1 + cells in the choroid plexus (CP), suggesting that CP-related pathology might be the contributing factor for the observed CSF/ventricular shrinkage phenotype.ConclusionsOur data, for the first time, reveal a robust and global CSF/ventricular space shrinkage pathology in the Cx3cr1-iDTR genetic ablation model caused by iDTR allele, but not in the PLX5622 ablation model, and suggest that this pathology is not due to brain edema formation but to CP related pathology. Given the wide utilization of the iDTR allele and the Cx3cr1-iDTR model, it is crucial to fully characterize this pathology to understand the underlying causal mechanisms. Specifically, caution is needed when utilizing this model to interpret subtle neurologic functional changes that are thought to be mediated by microglia but could, instead, be due to CSF/ventricular loss in the genetic ablation model.

Microglial MERTK eliminates phosphatidylserine-displaying inhibitory post-synapses.

Glia contribute to synapse elimination through phagocytosis in the central nervous system. Despite the important roles of this process in development and neurological disorders, the identity and regulation of the "eat-me" signal that initiates glia-mediated phagocytosis of synapses has remained incompletely understood. Here, we generated conditional knockout mice with neuronal-specific deletion of the flippase chaperone Cdc50a, to induce stable exposure of phosphatidylserine, a well-known "eat-me" signal for apoptotic cells, on the neuronal outer membrane. Surprisingly, acute Cdc50a deletion in mature neurons causes preferential phosphatidylserine exposure in neuronal somas and specific loss of inhibitory post-synapses without effects on other synapses, resulting in abnormal excitability and seizures. Ablation of microglia or the deletion of microglial phagocytic receptor Mertk prevents the loss of inhibitory post-synapses and the seizure phenotype, indicating that microglial phagocytosis is responsible for inhibitory post-synapse elimination. Moreover, we found that phosphatidylserine is used for microglia-mediated pruning of inhibitory post-synapses in normal brains, suggesting that phosphatidylserine serves as a general "eat-me" signal for inhibitory post-synapse elimination.

The role of macrophage scavenger receptor 1 (Msr1) in prion pathogenesis

The progression of prion diseases is accompanied by the accumulation of prions in the brain. Ablation of microglia enhances prion accumulation and accelerates disease progression, suggesting that microglia play a neuroprotective role by clearing prions. However, the mechanisms underlying the phagocytosis and clearance of prion are largely unknown. The macrophage scavenger receptor 1 (Msr1) is an important phagocytic receptor expressed by microglia in the brain and is involved in the uptake and clearance of soluble amyloid-β. We therefore asked whether Msr1 might play a role in prion clearance and assessed the scavenger function of Msr1 in prion pathogenesis. We found that Msr1 expression was upregulated in prion-infected mouse brains. However, Msr1 deficiency did not change prion disease progression or lesion patterns. Prion deposition in Msr1 deficient mice was similar to their wild-type littermates. In addition, prion-induced neuroinflammation was not affected by Msr1 ablation. We conclude that Msr1 does not play a major role in prion pathogenesis.Key messagesMsr1 expression is upregulated in prion-infected mouse brains at the terminal stageMsr1 deficiency does not affect prion disease progressionMsr1 does not play a major role in prion clearance or prion pathogenesisMicroglia-mediated phagocytosis and clearance of Aβ and prion may adopt distinct molecular pathways

Microglia Control CNS T Regulatory Cell Activity During Remission From EAE Pathology

Microglia, the parenchymal brain macrophages of the central nerve system (CNS), have emerged as critical players in brain development and homeostasis. However, immune functions of these cells remain less well defined. We investigated contributions of microglia in a relapsing remitting (RR) multiple sclerosis paradigm, experimental autoimmune encephalitis (RR-EAE) in C57BL/6 / SJL F1 mice. Fate mapping-assisted translatome profiling during the RR disease course revealed the potential of microglia to interact with T cells through antigen presentation, co-stimulation and co-inhibition. Abundant microglia / T cell aggregates, as observed by histology and flow cytometry, supported the notion of functional interactions during remission, with a bias towards T regulatory cells. Finally, microglia ablation and microglia-restricted Ifnγ receptor mutagenesis, while compatible with EAE onset, significantly affected the T cell compartment of the animals and remission. Collectively, our data establish critical interactions of microglia with T cells during autoimmune neuro-degeneration.

Microglia depletion diminishes key elements of the leukotriene pathway in the brain of Alzheimer\u2019s Disease mice

Leukotrienes (LTs) contribute to the neuropathology of chronic neurodegenerative disorders including Alzheimer’s Disease (AD), where they mediate neuroinflammation and neuronal cell-death. In consequence, blocking the action of Leukotrienes (LTs) ameliorates pathologies and improves cognitive function in animal models of neurodegeneration. Surprisingly, the source of Leukotrienes (LTs) in the brain is largely unknown. Here, we identified the Leukotriene (LT) synthesis rate-limiting enzyme 5-Lipoxygenase (5-Lox) primarily in neurons and to a lesser extent in a subpopulation of microglia in human Alzheimer´s Disease (AD) hippocampus brain sections and in brains of APP Swedish PS1 dE9 (APP-PS1) mice, a transgenic model for Alzheimer´s Disease (AD) pathology. The 5-Lipoxygenase (5-Lox) activating protein (FLAP), which anchors 5-Lipoxygenase (5-Lox) to the membrane and mediates the contact to the substrate arachidonic acid, was confined exclusively to microglia with the entire microglia population expressing 5-Lipoxygenase activating protein (FLAP). To define the contribution of microglia in the Leukotriene (LT) biosynthesis pathway, we ablated microglia using the colony stimulating factor 1 receptor (CSF1R) inhibitor PLX5622 in wildtype (WT) and APP-PS1 mice. Microglia ablation not only diminished the expression of FLAP and of the Leukotriene (LT) receptor Cysteinylleukotriene receptor 1 (CysLTR1), as expected based on their microglia cell type-specific expression, but also drastically reduced 5-Lipoxygenase (5-Lox) mRNA expression in the brain and its protein expression in neurons, in particular in wildtype (WT) mice. In conclusion i) microglia are key in Leukotriene (LT) biosynthesis, and ii) they regulate neuronal 5-Lipoxygenase (5-Lox) expression implying a yet unknown signaling mechanism between neurons and microglia.

Depletion of microglia in developing cortical circuits reveals its critical role in glutamatergic synapse development, functional connectivity, and critical period plasticity

Microglia populate the early developing brain and mediate pruning of the central synapses. Yet, little is known on their functional significance in shaping the developing cortical circuits. We hypothesize that the developing cortical circuits require microglia for proper circuit maturation and connectivity, and as such, ablation of microglia during the cortical critical period may result in a long-lasting circuit abnormality. We administered PLX3397, a colony-stimulating factor 1 receptor inhibitor, to mice starting at postnatal day 14 and through P28, which depletes >75% of microglia in the visual cortex (VC). This treatment largely covers the critical period (P19-32) of VC maturation and plasticity. Patch clamp recording in VC layer 2/3 (L2/3) and L5 neurons revealed increased mEPSC frequency and reduced amplitude, and decreased AMPA/NMDA current ratio, indicative of altered synapse maturation. Increased spine density was observed in these neurons, potentially reflecting impaired synapse pruning. In addition, VC intracortical circuit functional connectivity, assessed by laser scanning photostimulation combined with glutamate uncaging, was dramatically altered. Using two photon longitudinal dendritic spine imaging, we confirmed that spine elimination/pruning was diminished during VC critical period when microglia were depleted. Reduced spine pruning thus may account for increased spine density and disrupted connectivity of VC circuits. Lastly, using single-unit recording combined with monocular deprivation, we found that ocular dominance plasticity in the VC was obliterated during the critical period as a result of microglia depletion. These data establish a critical role of microglia in developmental cortical synapse pruning, maturation, functional connectivity, and critical period plasticity.