Noninvasive Quantification of Retinal Microglia Using Widefield Autofluorescence Imaging.

Abstract:

Purpose:To validate widefield autofluorescence (AF) in vivo imaging of the retina in mice expressing green fluorescent protein (gfp) in microglia, and to monitor retinal microglia reconstitution in vivo after lethal irradiation and bone marrow transplantation. Methods:Transgenic Cx3cr1gfp/gfp and wildtype Balb/c mice were used in this study. A confocal scanning laser ophthalmoscope was used for AF imaging with a 55° and a widefield 102° lens. Intrasession reproducibility was assessed for each lens. To investigate reconstitution in vivo, bone marrow from Cx3cr1gfp/gfp mice was used to rescue lethally irradiated wildtype mice. Data were compared to confocal microscopy of retinal flat mounts. Results:Both the 55° and the 102° lens produced high resolution images of retinal microglia with similar microglia density. However, compared to the 55° lens, the widefield 102° lens captured approximately 3.6 times more microglia cells (1515 ± 123 cells versus 445 ± 76 cells [mean ± SD], for 102° and 55°, respectively, P < 0.001). No statistical difference in the number of gfp positive cells within corresponding areas was observed within the same imaging session. Imaging of microglia reconstitution showed a similar time course compared to flat mount preparations with an excellent correlation between microglia cell numbers in AF and gfp-stained flat mounts (R = 0.92, P < 0.0001). Conclusions:Widefield AF imaging of mice with gfp expressing microglia can be used to quantify retinal microglia. In vivo microglia counts corresponded very well with ex vivo counts on retinal flat mounts. As such, AF imaging can largely replace ex vivo quantification.

authors

Kokona D,Schneider N,Giannakaki-Zimmermann H,Jovanovic J,Ebneter A,Zinkernagel M

doi

10.1167/iovs.16-20916

subject

Has Abstract

pub_date

2017-04-01 00:00:00

pages

2160-2165

issue

4

eissn

0146-0404

issn

1552-5783

pii

2618350

journal_volume

58

pub_type

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