Abstract
PurposeMicroglia represent the primary resident immune cells in the CNS, and have been implicated in the pathology of neurodegenerative diseases. Under basal or “resting” conditions, microglia possess ramified morphologies and exhibit dynamic surveying movements in their processes. Despite the prominence of this phenomenon, the function and regulation of microglial morphology and dynamic behavior are incompletely understood. We investigate here whether and how neurotransmission regulates “resting” microglial morphology and behavior.MethodsWe employed an ex vivo mouse retinal explant system in which endogenous neurotransmission and dynamic microglial behavior are present. We utilized live-cell time-lapse confocal imaging to study the morphology and behavior of GFP-labeled retinal microglia in response to neurotransmitter agonists and antagonists. Patch clamp electrophysiology and immunohistochemical localization of glutamate receptors were also used to investigate direct-versus-indirect effects of neurotransmission by microglia.ResultsRetinal microglial morphology and dynamic behavior were not cell-autonomously regulated but are instead modulated by endogenous neurotransmission. Morphological parameters and process motility were differentially regulated by different modes of neurotransmission and were increased by ionotropic glutamatergic neurotransmission and decreased by ionotropic GABAergic neurotransmission. These neurotransmitter influences on retinal microglia were however unlikely to be directly mediated; local applications of neurotransmitters were unable to elicit electrical responses on microglia patch-clamp recordings and ionotropic glutamatergic receptors were not located on microglial cell bodies or processes by immunofluorescent labeling. Instead, these influences were mediated indirectly via extracellular ATP, released in response to glutamatergic neurotransmission through probenecid-sensitive pannexin hemichannels.ConclusionsOur results demonstrate that neurotransmission plays an endogenous role in regulating the morphology and behavior of “resting” microglia in the retina. These findings illustrate a mode of constitutive signaling between the neural and immune compartments of the CNS through which immune cells may be regulated in concert with levels of neural activity.
Highlights
Microglia constitute the resident tissue macrophage and primary immune cell of the CNS
These neurotransmitter influences on retinal microglia were unlikely to be directly mediated; local applications of neurotransmitters were unable to elicit electrical responses on microglia patchclamp recordings and ionotropic glutamatergic receptors were not located on microglial cell bodies or processes by immunofluorescent labeling
Our results demonstrate that neurotransmission plays an endogenous role in regulating the morphology and behavior of ‘‘resting’’ microglia in the retina
Summary
Microglia constitute the resident tissue macrophage and primary immune cell of the CNS. Under basal conditions, ‘‘resting’’ microglia demonstrate ramified morphologies and extend fine processes through nearby neural parenchyma in a non-overlapping manner. Termed ‘‘resting’’, microglia in the retina and the brain exhibit rapid and extensive dynamism in their processes, enabling each microglial cell to come into close proximity and establish repeated contact with nearby neurons, macroglia, and blood vessels [1,2,3]. While a ramified morphology and dynamic process behavior are key phenotypes of ‘‘resting’’ microglia, knowledge on how these are influenced and regulated by extracellular signals is largely incomplete [4]. As a result of their distribution and morphology, microglia are in constant and intimate contact with multiple signals originating from nearby neurons and macroglia. The transformation of a microglial cell from a ‘‘resting’’ state through gradations of ‘‘activated’’ states is thought to depend on the overall balance between ‘‘on’’ signals that promote activation, and ‘‘off’’ signals that repress it [5,6]
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