Onset of microglial entry into developing quail retina coincides with increased expression of active caspase-3 and is mediated by extracellular ATP and UDP

Abstract

Microglial cell precursors located in the area of the base of the pecten and the optic nerve head (BP/ONH) start to enter the retina of quail embryos at the 7th day of incubation (E7), subsequently colonizing the entire retina by central-to-peripheral tangential migration, as previously shown by our group. The present study demonstrates a precise chronological coincidence of the onset of microglial cell entry into the retina with a striking increase in death of retinal cells, as revealed by their active caspase-3 expression and TUNEL staining, in regions dorsal to the BP/ONH area, suggesting that dying retinal cells would contribute to the microglial cell inflow into the retina. However, the molecular mechanisms involved in this inflow are currently unclear. Extracellular nucleotides, such as ATP and UDP, have previously been shown to favor migration of microglia towards brain injuries because they are released by apoptotic cells and stimulate both chemotaxis and chemokinesis in microglial cells via signaling through purinergic receptors. Hence, we tested here the hypothesis that ATP and UDP play a role in the entry and migration of microglial precursors into the developing retina. For this purpose, we used an experimental model system based on organotypic cultures of E6.5 quail embryo retina explants, which mimics the entry and migration of microglial precursors in the in situ developing retina. Inhibition of purinergic signaling by treating retina explants with either apyrase, a nucleotide-hydrolyzing enzyme, or suramin, a broad spectrum antagonist of purinergic receptors, significantly prevents the entry of microglial cells into the retina. In addition, treatment of retina explants with either exogenous ATP or UDP results in significantly increased numbers of microglial cells entering the retina. In light of these findings, we conclude that purinergic signaling by extracellular ATP and UDP is necessary for the entry and migration of microglial cells into the embryonic retina by inducing chemokinesis in these cells.

Microglia in the developing retina couple phagocytosis with the progression of apoptosis via P2RY12 signaling

“…Similar systems (cell‐type‐specific promoter driving bacterial NTR) have been used to ablate selected cell types in the zebrafish . Zebrafish microglia express mpeg1 ‐driven transgenes, and we confirmed that microglia in retinas of mpeg1 :nfsBeYFP zebrafish are YFP+ (Figure A,B), indicating that these microglial populations are susceptible to ablation by metronidazole (Mtz) treatment.…”

“…In the embryonic mouse retina, detection of microglia, the resident phagocytic macrophage population, coincides temporally with detection of cell death . In the zebrafish, apoptotic cells can be detected in the brain and retina by 2 days postfertilization, which is also coincident with colonization of the central nervous system (CNS) by microglia . In addition, using the zebrafish model organism, recent studies indicate that normal developmental apoptosis is a significant factor driving macrophage entry into and establishment of microglia in the brain …”

Microglia in the developing retina

“…They are present in human retina by 10 weeks gestation and in mouse retina by E11.5, though it is likely they arrive even earlier [17, 18]. Similar timing has been documented in other species (E7 in quail, [19]; and at E12 in rat, [20]). Two waves of retinal microglia infiltration have been proposed based on the spatiotemporal localization of these cells.…”

BAM15 attenuates transportation-induced apoptosis in iPS-differentiated retinal tissue

“…These immunofluorescence results indicated cleaved caspase-3 expressed occasionally in normal retinal tissues in the developing stage [37,[40][41][42]. In the control group after transportation, cups of 30 days and 60 days manifested a remarkably higher number of apoptotic cells.…”