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Microglial Activation Promotes Cell Survival in Organotypic Cultures of Postnatal Mouse Retinal Explants.

ABSTRACT: The role of microglia during neurodegeneration remains controversial. We investigated whether microglial cells have a neurotoxic or neuroprotective function in the retina. Retinal explants from 10-day-old mice were treated in vitro with minocycline to inhibit microglial activation, with LPS to increase microglial activation, or with liposomes loaded with clodronate (Lip-Clo) to deplete microglial cells. Flow cytometry was used to assess the viability of retinal cells in the explants and the TUNEL method to show the distribution of dead cells. The immunophenotypic and morphological features of microglia and their distribution were analyzed with flow cytometry and immunocytochemistry. Treatment of retinal explants with minocycline reduced microglial activation and simultaneously significantly decreased cell viability and increased the presence of TUNEL-labeled cell profiles. This treatment also prevented the migration of microglial cells towards the outer nuclear layer, where cell death was most abundant. The LPS treatment increased microglial activation but had no effect on cell viability or microglial distribution. Finally, partial microglial removal with Lip-Clo diminished the cell viability in the retinal explants, showing a similar effect to that of minocycline. Hence, cell viability is diminished in retinal explants cultured in vitro when microglial cells are removed or their activation is inhibited, indicating a neurotrophic role for microglia in this system.

SUBMITTER: Ferrer-Martin RM

PROVIDER: S-EPMC4529135 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Microglial Activation Promotes Cell Survival in Organotypic Cultures of Postnatal Mouse Retinal Explants.

Ferrer-Martín Rosa M RM Martín-Oliva David D Sierra-Martín Ana A Carrasco Maria-Carmen MC Martín-Estebané María M Calvente Ruth R Martín-Guerrero Sandra M SM Marín-Teva José L JL Navascués Julio J Cuadros Miguel A MA

PloS one 20150807 8

The role of microglia during neurodegeneration remains controversial. We investigated whether microglial cells have a neurotoxic or neuroprotective function in the retina. Retinal explants from 10-day-old mice were treated in vitro with minocycline to inhibit microglial activation, with LPS to increase microglial activation, or with liposomes loaded with clodronate (Lip-Clo) to deplete microglial cells. Flow cytometry was used to assess the viability of retinal cells in the explants and the TUNE ...[more]

PMID: 26252475

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Project description:We previously found that Mertk and its ligand Gas6, astrocytic genes involved in phagocytosis, are upregulated after acute sleep deprivation. These results suggested that astrocytes may engage in phagocytic activity during extended wake, but direct evidence was lacking. Studies in humans and rodents also found that sleep loss increases peripheral markers of inflammation, but whether these changes are associated with neuroinflammation and/or activation of microglia, the brain's resident innate immune cells, was unknown. Here we used serial block-face scanning electron microscopy to obtain 3D volume measurements of synapses and surrounding astrocytic processes in mouse frontal cortex after 6-8 h of sleep, spontaneous wake, or sleep deprivation (SD) and after chronic (∼5 d) sleep restriction (CSR). Astrocytic phagocytosis, mainly of presynaptic components of large synapses, increased after both acute and chronic sleep loss relative to sleep and wake. MERTK expression and lipid peroxidation in synaptoneurosomes also increased to a similar extent after short and long sleep loss, suggesting that astrocytic phagocytosis may represent the brain's response to the increase in synaptic activity associated with prolonged wake, clearing worn components of heavily used synapses. Using confocal microscopy, we then found that CSR but not SD mice show morphological signs of microglial activation and enhanced microglial phagocytosis of synaptic elements, without obvious signs of neuroinflammation in the CSF. Because low-level sustained microglia activation can lead to abnormal responses to a secondary insult, these results suggest that chronic sleep loss, through microglia priming, may predispose the brain to further damage.SIGNIFICANCE STATEMENT We find that astrocytic phagocytosis of synaptic elements, mostly of presynaptic origin and in large synapses, is upregulated already after a few hours of sleep deprivation and shows a further significant increase after prolonged and severe sleep loss, suggesting that it may promote the housekeeping of heavily used and strong synapses in response to the increased neuronal activity of extended wake. By contrast, chronic sleep restriction but not acute sleep loss activates microglia, promotes their phagocytic activity, and does so in the absence of overt signs of neuroinflammation, suggesting that like many other stressors, extended sleep disruption may lead to a state of sustained microglia activation, perhaps increasing the brain's susceptibility to other forms of damage.

Dataset Information

Microglial Activation Promotes Cell Survival in Organotypic Cultures of Postnatal Mouse Retinal Explants.

Publications

Microglial Activation Promotes Cell Survival in Organotypic Cultures of Postnatal Mouse Retinal Explants.

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