Abstract
Purpose :
We recently demonstrated that microglia regulate astrocyte and superficial retinal vascular spatial patterning through complement activation. Here, we aimed to define the role of microglia in the maturation of Müller cells and subsequent formation of the inner retinal vascular network.
Methods :
C57BL6J retinal flatmounts and cryosections were examined at postnatal day (P) 7, 10, and 15 to determine Müller cell-microglial-endothelial interactions during inner retinal vascular development by immunohistochemistry (IHC). Microglia were depleted in C57BL6J litters using the Csf1r-antagonist (PLX5622). Müller marker expression/distribution and inner vascular layer formation were examined by IHC. Gene expression profiles of microglia-depleted and control P10 retinas were examined by RT-PCR and bulk RNA-seq. Statistical differences between the groups were determined by unpaired student’s t-test.
Results :
We analyzed retinal cross-sections during the defined phases of inner retinal vascular growth (P7, P10, and P15) to determine microglial interactions with differentiating Müller cells and the growing inner retinal vasculature. Data showed close association of P2ry12+ microglia with Müller cells (glutamine synthetase (GS)+) and the growing inner retinal vasculature (CD31+). Depletion of microglia showed significant reduction in the development of the intermediate (P=0.004) and deep (P=0.001) vascular plexuses at P10. Further examination of P10 retinas by IHC and transcriptomic studies revealed significant decreases in Müller cell maturation markers such as GS, aquaporin-4, Kir4.1, and Kir2.4 compared to the controls. Intriguingly, even at P15 (vascular growth completion phase), GS expression and intermediate (P=0.007) and deeper (P=0.002) retinal vascular densities were significantly reduced in microglia depleted retinas compared to controls.
Conclusions :
Taken together our study suggests that microglia have a role in promoting Müller cell maturation and the subsequent formation of interconnected vascular networks in the retina.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.