September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Microglial dynamics and phenotype are altered by Angiotensin II in the mouse retina
Author Affiliations & Notes
  • Joanna Phipps
    Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia
  • Kirstan Vessey
    Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia
  • Nupur Nag
    Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia
  • Andrew Ian Jobling
    Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia
  • Erica L Fletcher
    Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia
  • Footnotes
    Commercial Relationships   Joanna Phipps, None; Kirstan Vessey, None; Nupur Nag, None; Andrew Jobling, None; Erica Fletcher, None
  • Footnotes
    Support  ARC DECRA Grant DE140100099
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 2232. doi:
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    • Get Citation

      Joanna Phipps, Kirstan Vessey, Nupur Nag, Andrew Ian Jobling, Erica L Fletcher; Microglial dynamics and phenotype are altered by Angiotensin II in the mouse retina. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2232.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : Microglia are the principal immune cells of the central nervous system and have dynamic, ramified processes that extend and recede whilst surveying the retinal environment. Regulation of microglial responses are mediated via cell surface receptors, and there is evidence that microglia can produce and release angiogenic factors involved in neovascularisation. We investigated whether the pro-angiogenic factor Angiotensin II (AngII) had a direct effect on the dynamics and activation state of retinal microglia.

Methods : We used transgenic heterozygote Cx3cr1+/GFP mice which are normal with the exception that microglia are labeled with Enhanced Green Fluorescent Protein (eGFP). Three month old Cx3Cr1+/GFP mice received 1µl intravitreal injection of 10mM AngII, with fellow eyes receiving 1µl saline. Retinae were collected after 24hrs and processed for immunocytochemistry. For live cell imaging, retinae from 3-6 week old Cx3Cr1+/GFP mice were placed in a temperature controlled recording chamber perfused with Ames’ medium. Retinae were imaged with a Leica SP5 confocal microscope and 10s z-series time-lapse images collected for 20 minutes (10min baseline, followed by 10min perfusion of 5µm AngII or Ames’ medium). NIH ImageJ and Metamorph Offline® software were used for morphological analyses.

Results : Retinal microglia in Cx3cr1+/GFP mice expressed the Angiotensin type 1 receptor (AT1R). Intravitreal injections of AngII resulted in altered microglial morphology consistent with activation, characterised by increased soma size (n=6, 23% p<0.001) decreased process length (n=6, 20% p<0.05), and an increase in M2 phenotype (n=3). Live microglia imaged in retinal exmounts showed characteristic dynamic movement of processes under control conditions. Perfusion of AngII induced an immediate change in retinal microglia with decreased process length and increased soma size compared to controls (n=7, p<0.05) consistent with altered morphology in fixed tissue. The short latency (1-2min) of the AngII effect on microglia is suggestive of a direct effect.

Conclusions : We show that retinal microglia dynamics and activation state are significantly altered by angiogenic factors. Specifically, AngII may directly activate AT1Rs on microglia and contribute to retinal inflammation. This may have implications for diseases like diabetic retinopathy.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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