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Open Access Highly Accessed Research

A quantitative spatiotemporal analysis of microglia morphology during ischemic stroke and reperfusion

Helena W Morrison and Jessica A Filosa*

Author Affiliations

Georgia Health Sciences University, 1120 15th, St, Augusta, GA, 30912, USA

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Journal of Neuroinflammation 2013, 10:4  doi:10.1186/1742-2094-10-4

Published: 11 January 2013

Abstract

Background

Microglia cells continuously survey the healthy brain in a ramified morphology and, in response to injury, undergo progressive morphological and functional changes that encompass microglia activation. Although ideally positioned for immediate response to ischemic stroke (IS) and reperfusion, their progressive morphological transformation into activated cells has not been quantified. In addition, it is not well understood if diverse microglia morphologies correlate to diverse microglia functions. As such, the dichotomous nature of these cells continues to confound our understanding of microglia-mediated injury after IS and reperfusion. The purpose of this study was to quantitatively characterize the spatiotemporal pattern of microglia morphology during the evolution of cerebral injury after IS and reperfusion.

Methods

Male C57Bl/6 mice were subjected to focal cerebral ischemia and periods of reperfusion (0, 8 and 24 h). The microglia process length/cell and number of endpoints/cell was quantified from immunofluorescent confocal images of brain regions using a skeleton analysis method developed for this study. Live cell morphology and process activity were measured from movies acquired in acute brain slices from GFP-CX3CR1 transgenic mice after IS and 24-h reperfusion. Regional CD11b and iNOS expressions were measured from confocal images and Western blot, respectively, to assess microglia proinflammatory function.

Results

Quantitative analysis reveals a significant spatiotemporal relationship between microglia morphology and evolving cerebral injury in the ipsilateral hemisphere after IS and reperfusion. Microglia were both hyper- and de-ramified in striatal and cortical brain regions (respectively) after 60 min of focal cerebral ischemia. However, a de-ramified morphology was prominent when ischemia was coupled to reperfusion. Live microglia were de-ramified, and, in addition, process activity was severely blunted proximal to the necrotic core after IS and 24 h of reperfusion. CD11b expression, but not iNOS expression, was increased in regions of hyper- and de-ramified microglia during the course of ischemic stroke and 24 h of reperfusion.

Conclusions

Our findings illustrate that microglia activation after stroke includes both increased and decreased cell ramification. Importantly, quantitative analyses of microglial morphology and activity are feasible and, in future studies, would assist in the comprehensive identification and stratification of their dichotomous contribution toward cerebral injury and recovery during IS and reperfusion.

Keywords:
Microglia; Morphology; Mouse; Ischemic stroke; Reperfusion