title: Activation of neuronal NMDA receptors triggers transient ATP-mediated microglial process outgrowth.
abstract: Microglia are morphologically dynamic cells that rapidly extend their processes in response to various stimuli including extracellular ATP. In this study, we tested the hypothesis that stimulation of neuronal NMDARs trigger ATP release leading to communication with microglia. We used acute mouse hippocampal brain slices and two-photon laser scanning microscopy to study microglial dynamics and developed a novel protocol for fixation and immunolabeling of microglia processes. Similar to direct topical ATP application in vivo, short multiple applications of NMDA triggered transient microglia process outgrowth that was reversible and repeatable indicating that this was not due to excitotoxic damage. Stimulation of NMDAR was required as NMDAR antagonists, but not blockers of AMPA/kainate receptors or voltage-gated sodium channels, prevented microglial outgrowth. We report that ATP release, secondary to NMDAR activation, was the key mediator of this neuron-microglia communication as both blocking purinergic receptors and inhibiting hydrolysis of ATP to prevent locally generated gradients abolished outgrowth. Pharmacological and genetic analyses showed that the NMDA-triggered microglia process extension was independent of Pannexin 1, the ATP releasing channels, ATP release from astrocytes via connexins, and nitric oxide generation. Finally, using whole-cell patch clamping we demonstrate that activation of dendritic NMDAR on single neurons is sufficient to trigger microglia process outgrowth. Our results suggest that dendritic neuronal NMDAR activation triggers ATP release via a Pannexin 1-independent manner that induces outgrowth of microglia processes. This represents a novel uncharacterized form of neuron-microglial communication mediated by ATP.
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada.
authors.forename: Lasse
authors.initials: L
authors.lastname: Dissing-Olesen
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada.
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: LeDue
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada.
authors.forename: Ravi L
authors.initials: RL
authors.lastname: Rungta
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada.
authors.forename: Jasmin K
authors.initials: JK
authors.lastname: Hefendehl
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada.
authors.forename: Hyun B
authors.initials: HB
authors.lastname: Choi
authors.affiliation: Brain Research Centre, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada bmacvicar@brain.ubc.ca.
authors.forename: Brian A
authors.initials: BA
authors.lastname: MacVicar
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000166:cytology* / Q000187:drug effects; D002135:Calcium-Binding Proteins / Q000378:metabolism; D048708:Cell Growth Processes / Q000187:drug effects / Q000235:genetics; D018683:Excitatory Amino Acid Agents / Q000494:pharmacology; D005260:Female; D066298:In Vitro Techniques; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008840:Microfilament Proteins / Q000378:metabolism; D017628:Microglia / Q000166:cytology / Q000378:metabolism*; D016202:N-Methylaspartate / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism*; D058925:Receptors, Purinergic P2Y12 / Q000378:metabolism; D013997:Time Factors
chemical_list: C491049:Aif1 protein, mouse; D002135:Calcium-Binding Proteins; D018683:Excitatory Amino Acid Agents; D008840:Microfilament Proteins; C548577:P2ry12 protein, mouse; D016194:Receptors, N-Methyl-D-Aspartate; D058925:Receptors, Purinergic P2Y12; D016202:N-Methylaspartate; D000255:Adenosine Triphosphate; D008274:Magnesium
keywords: ATP purinergic receptors; NMDA glutamate receptors; Pannexin; microglia process motility; synaptic surveillance
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