Article Nanoscale Surveillance of the Brain by Microglia via cAMP-Regulated Filopodia Graphical Abstract Highlights d Microglia use actin-dependent filopodia to efficiently sample the brain parenchyma d Intracellular cAMP drives filopodia growth but induces large process retraction d Norepinephrine and nitric oxide contribute to cAMP-driven filopodia extension d Fine filopodia and large processes establish dual-scale surveillance by microglia This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). relative to variations in the brain parenchyma environment. STAR+METHODSDetailed methods are provided in the online version of this paper and include the following:TABLE d CONTACT FOR REAGENTS AND RESOURCE SHARING d EXPERIMENTAL MODEL AND SUBJECT DETAILS B Animals B Acute slice preparation and maintenance B Isolation of rat microglia d METHOD DETAILS B Open cranial window surgery and in vivo two-photon imaging B Brain perfusion, immunofluorescence and confocal imaging B Acute slice two-photon imaging B Slice fixation and immunolabelling B Rat primary microglia live imaging B Western blotting d QUANTIFICATION AND STATISTICAL ANALYSIS B Image processing and analysis B Statistical analysis SUPPLEMENTAL INFORMATION Supplemental Information can be found online at https://doi.

Astrocytes display complex morphologies with an array of fine extensions extending from the soma and the primary thick processes. Until the use of genetically encoded calcium indicators (GECIs) selectively expressed in astrocytes, Ca signaling was only examined in soma and thick primary processes of astrocytes where Ca -sensitive fluorescent dyes could be imaged. GECI imaging in astrocytes revealed a previously unsuspected pattern of spontaneous Ca transients in fine processes that has not been observed without chronic expression of GECIs, raising potential concerns about the effects of GECI expression. Here, we perform two-photon imaging of Ca transients in adult CA1 hippocampal astrocytes using a new single-cell patch-loading strategy to image Ca -sensitive fluorescent dyes in the cytoplasm of fine processes. We observed that astrocyte fine processes exhibited a high frequency of spontaneous Ca transients whereas astrocyte soma rarely showed spontaneous Ca oscillations similar to previous reports using GECIs. We exploited this new approach to show these signals were independent of neuronal spiking, metabotropic glutamate receptor (mGluR) activity, TRPA1 channels, and L- or T-type voltage-gated calcium channels. Removal of extracellular Ca almost completely and reversibly abolished the spontaneous signals while IP R2 KO mice also exhibited spontaneous and compartmentalized signals, suggesting they rely on influx of extracellular Ca . The Ca influx dependency of the spontaneous signals in patch-loaded astrocytes was also observed in astrocytes expressing GCaMP3, further highlighting the presence of Ca influx pathways in astrocytes. The mechanisms underlying these localized Ca signals are critical for understanding how astrocytes regulate important functions in the adult brain. GLIA 2016;64:2093-2103.

This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.12/issuetoc.

Microglia are highly motile cells that continuously monitor the brain environment and respond to damage-associated cues. While glucose is the main energy substrate used by neurons in the brain, the nutrients metabolized by microglia to support surveillance of the parenchyma remain unexplored. Here, we use fluorescence lifetime imaging of intracellular NAD(P)H and time-lapse two-photon imaging of microglial dynamics in vivo and in situ, to show unique aspects of the microglial metabolic signature in the brain. Microglia are metabolically flexible and can rapidly adapt to consume glutamine as an alternative metabolic fuel in the absence of glucose. During insulin-induced hypoglycemia in vivo or in aglycemia in acute brain slices, glutaminolysis supports the maintenance of microglial process motility and damage-sensing functions. This metabolic shift sustains mitochondrial metabolism and requires mTOR-dependent signaling. This remarkable plasticity allows microglia to maintain their critical surveillance and phagocytic roles, even after brain neuroenergetic homeostasis is compromised.

No abstract

P2X receptors are ATP-gated nonselective cation channels highly permeable to calcium that contribute to nociception and inflammatory responses. The P2X 4 subtype, upregulated in activated microglia, is thought to play a critical role in the development of tactile allodynia following peripheral nerve injury. Posttranslational regulation of P2X 4 function is crucial to the cellular mechanisms of neuropathic pain, however it remains poorly understood. Here, we show that the phosphoinositides PI(4

Investigations on the role of microglia in the brain have traditionally been focused on their contributions to disease states. However, recent observations have now convincingly shown that microglia in the healthy brain are not passive bystanders, but instead play a critical role in both central nervous system development and homeostasis of synaptic circuits in the adult. Here, we review the various mechanisms by which microglia impact neuronal communication in the healthy adult brain, both by sensing nearby synaptic responses and by actively modulating neuronal function. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 78: 593-603, 2018.

Visual Abstract