Biology subjects

MacVicar, B. A.

Publications and source records attributed to MacVicar, B. A..

6 recordsLinked to original sources

bioRxiv2026

Brain pericytes are an oxidative stress-sensitive cell population damaged through ROS-1 dependent TRPA1 activation.

Oxidative stress is a pathological process driven by an imbalance between the production and clearance of reactive oxygen species (ROS). It is a hallmark of numerous brain disorders, driving neuroinflammation, blood-brain barrier (BBB) disruption, and neurodegeneration. Here, we used two-photon microscopy in rodent brain slices to evaluate cell-type vulnerability to oxidative stress. We identified a microvascular cell population displaying pronounced sensitivity to H2O2 and thiol oxidation. These Pdgfr-{beta} expressing cells were identified as pericytes and perivascular fibroblasts. Because pericytes regulate capillary perfusion and BBB integrity, we examined mechanisms driving their injury. Pericyte damage was independent of apoptosis or necroptosis, but required Ca2+ influx. While blocking voltage-gated Ca2+, TRPC, Orai channels was ineffective, inhibitors of TRPA1 channels reduced pericyte damage. Immunostaining and western blotting showed TRPA1 associated with brain capillaries and pericytes. Functionally, TRPA1 regulated uptake of the pericyte-selective marker NeuroTrace, whereas pericyte GCaMP6f imaging demonstrated that TRPA1 was not the primary driver of cytosolic Ca2+ elevations during oxidative stress. These findings establish pericytes as vulnerable targets of oxidative stress and implicate TRPA1 as a mediator of pericyte structural degeneration. This sensitivity to oxidative stress, mediated by ROS-TRPA1 coupling, could contribute to the pericyte stress and degeneration observed in brain pathology.

neuroscience↗

bioRxiv2025

Single cell approaches define the murine leptomeninges:cortical brain interface as a distinct cellular neighborhood comprised of neural and nonneural cell types

The interface between the brain surface and the adjacent meninges is a selective barrier regulating fluid, protein and immune cell exchange between the CNS and periphery. However, the cell types that form this important interface are not yet fully defined. To address this limitation, we have used single cell RNA-sequencing (scRNA-seq) and single cell spatial transcriptomics together with morphological lineage tracing and immunostaining to analyze the adult murine cortex. We show that the cortical interface is comprised of three major cell types, leptomeningeal cells, border astrocytes and tissue-resident macrophages. On the peripheral side the interface is comprised of transcriptionally-distinct PDGFR-positive leptomeningeal mesenchymal cells that are intermingled with macrophages. This leptomeningeal pial layer is lined by a population of transcriptionally-distinct border astrocytes. The interface neighborhood is rich in growth factor mRNAs, including many leptomeningeal ligands predicted to act on both the border astrocytes and macrophages. On the CNS side of the interface is the relatively cell-sparse cortical layer one containing interneurons, microglia, parenchymal astrocytes, oligodendrocyte precursor cells and oligodendrocytes. Except for the border astrocytes, layer one cells are not closely-associated with the interface, suggesting that secreted ligands may be the major way the brain interface communicates with the underlying cortical parenchyma. Thus, our data provide a molecular/cellular resource describing the brain interface cell types and their interactions, thereby enabling future studies asking how this distinct cellular compartment regulates CNS:periphery interactions. Significance StatementRecent years have seen significant progress in identifying the diverse cell types within the meningeal space. However, the mechanisms by which these cells interact with glial and neuronal cells in layer one of the adult murine cortex remain poorly understood. During development, communication between radial precursors and meningeal layers is crucial for proper brain formation, but the role of this interaction in adulthood is still unclear. Additionally, how resident immune cells in the leptomeningeal space signal to layer one cortical cells or meningeal mesenchymal cells during homeostasis remains an open question. Understanding the identity, location, and interactions of these cells is essential for unraveling the complex dynamics at this critical brain interface.

neuroscience↗

bioRxiv2024

Microglia-astrocyte interplay mitigates Aβ toxicity in a novel human 3D neurosphere model of Alzheimer's Disease

BackgroundAlzheimers Disease (AD) is characterized by progressive amyloid beta (A{beta}) deposition in the brain, with eventual widespread neurodegeneration. While the cell-specific molecular signature of end-stage AD is reasonably well characterized through autopsy material, less is known about the molecular pathways in the human brain involved in the earliest exposure to A{beta}. Human model systems that not only replicate the pathological features of AD but also the transcriptional landscape in neurons, astrocytes and microglia are crucial for understanding disease mechanisms and for identifying novel therapeutic targets. MethodsIn this study, we used a human 3D iPSC-derived neurosphere model to explore how resident neurons, microglia and astrocytes and their interplay are modified by chronic amyloidosis induced over 3 to 5 weeks by supplementing media with synthetic A{beta}1-42 oligomers. Neurospheres under chronic A{beta} exposure were grown with or without microglia to investigate the functional roles of microglia. Neuronal activity and oxidative stress were monitored using genetically encoded indicators, including GCaMP6f and roGFP1, respectively. Single nuclei RNA sequencing (snRNA-seq) was performed to profile A{beta} and microglia driven transcriptional changes in neurons and astrocytes, providing a comprehensive analysis of cellular responses. ResultsMicroglia efficiently phagocytosed A{beta} inside neurospheres and significantly reduced neurotoxicity, mitigating amyloidosis-induced oxidative stress and neurodegeneration following different exposure times to A{beta}. The neuroprotective effects conferred by the presence of microglia was associated with unique gene expression profiles in astrocytes and neurons, including several known AD-associated genes such as APOE. These findings reveal how microglia can directly alter the molecular landscape of AD. ConclusionsOur human 3D neurosphere culture system with chronic A{beta} exposure reveals how microglia may be essential for the cellular and transcriptional responses in AD pathogenesis. Microglia are not only neuroprotective in neurospheres but also act as key drivers of A{beta}-dependent APOE expression suggesting critical roles for microglia in regulating APOE in the AD brain. This novel, well characterized, functional in vitro platform offers unique opportunities to study the roles and responses of microglia to A{beta} modelling key aspects of human AD. This tool will help identify new therapeutic targets, accelerating the transition from discovery to clinical applications. HighlightsO_LIWell-characterized functional human iPSC-derived 3D neurospheres (hiNS) consisting of neurons and astrocytes can be supplemented with microglia/macrophages (hiMG) C_LIO_LIChronic amyloidosis in the presence of hiMG recapitulate key features and gene expression profiles of AD C_LIO_LIhiMG within the model phagocytose A{beta} and mitigate A{beta}-induced neurotoxicity, reducing oxidative stress and neuronal damage C_LIO_LIhiMG are essential for A{beta} to upregulate AD-like gene expression signatures in astrocytes C_LIO_LIImmunohistochemical analysis reveals hiMG-dependent colocalization of A{beta} and APOE C_LI

neuroscience↗

bioRxiv2022

Bidirectional and reciprocal control of astrocyte free calcium by modest fluctuations in external potassium

Astrocytes sense and respond to changes in the concentration of extracellular K+, and separately contribute to multiple physiological processes through Ca2+ dependent mechanisms. Yet, whether a modest change in [K+]o impacts astrocyte free Ca2+ remains unclear. Using relative or quantitative two-photon fluorescence Ca2+ imaging in acute brain slices or in vivo in the somatosensory cortex from Sprague Dawley rats and C57Bl/6 mice, we showed that changes to external K+ (+/-1mM to 2.5mM) reciprocally controls the astrocyte Rhod-2 or OGB-1 Ca2+-dependent fluorescence in the soma, major processes and endfeet. The astrocyte Ca2+ decrease when [K+]o was elevated was sensitive to lowering the external concentration of Ca2+, Cl-, and HCO3-, but not Na+. Unexpectedly, the phenomenon was blocked by inhibiting K-Cl cotransport. Picrotoxin induced ictal neural activity drove an analogous decrease of astrocyte Ca2+. K+ mediated cerebral arteriole dilation in brain slices was also sensitive to inhibiting K-Cl cotransport as well as whole-cell patching a peri-arteriole astrocyte which perturbs normal Ca2+, Cl- and HCO3- concentration gradients. These data reveal subtle, bidirectional regulation of astrocyte free Ca2 via fluctuations of [K+]o within the physiological range.

neuroscience↗

bioRxiv2021

Age related susceptibility to grey matter demyelination and neurodegeneration is associated with meningeal neutrophil accumulation in an animal model of Multiple Sclerosis

People living with multiple sclerosis (MS) experience episodic central nervous system (CNS) white matter lesions instigated by autoreactive T cells. With age, MS patients show evidence of grey matter demyelination and experience devastating non-remitting symptomology. What drives progression is unclear and has been hampered by the lack of suitable animal models. Here we show that passive experimental autoimmune encephalomyelitis (EAE) induced by an adoptive transfer of young Th17 cells induces a non-remitting clinical phenotype that is associated with persistent leptomeningeal inflammation and cortical pathology in old, but not young SJL/J mice. While the quantity and quality of T cells did not differ in the brains of old vs young EAE mice, an increase in neutrophils and a decrease in B cells was observed in the brains of old mice. Neutrophils were also found in the leptomeninges of a subset of progressive MS patient brains that showed evidence of leptomeningeal inflammation and subpial cortical demyelination. Taken together, our data show that while Th17 cells initiate CNS inflammation, subsequent clinical symptoms and grey matter pathology are dictated by age and associated with other immune cells such as neutrophils.

immunology↗

bioRxiv2020

Quantitative fluorescence lifetime imaging uncovers a novel role for KCC2 chloride transport in dendritic microdomains

Intracellular chloride ion ([Cl-]i) homeostasis is critical for synaptic neurotransmission yet variations in subcellular domains are poorly understood owing to difficulties in obtaining quantitative, high-resolution measurements of dendritic [Cl-]i. We combined whole-cell patch clamp electrophysiology with simultaneous fluorescence lifetime imaging (FLIM) of the Cl- dye MQAE to quantitatively map dendritic Cl- levels in normal or pathological conditions. FLIM-based [Cl-]i estimates were corroborated by Rubi-GABA uncaging to measured EGABA. Low baseline [Cl-]i in dendrites required Cl- efflux via the K+-Cl- cotransporter KCC2 (SLC12A5). In contrast, pathological NMDA application generated spatially heterogeneous subdomains of high [Cl-]i that created dendritic blebs, a signature of ischemic stroke. These discrete regions of high [Cl-]i were caused by reversed KCC2 transport. Therefore monitoring [Cl-]i microdomains with a new high resolution FLIM-based technique identified novel roles for KCC2-dependent chloride transport to generate dendritic microdomains with implications for disease.

neuroscience↗