Abstract
Purpose :
Purpose: We previously showed that in a rapid optic nerve (ON) shortening event that occurs during Xenopus laevis development, astrocytes are the primary phagocytes within the ON parenchyma; by contrast, myeloid cells show no increase in the parenchyma and only modest increase in the ON sheath. Here, we used live imaging in very young X. laevis tadpoles to understand how astrocytes and myeloid cells cooperate to clear debris in the normal ON.
Methods :
Methods: Transgenes Tg(Fabp7:mBFPtag2), Tg(Fabp7:Mfge8D91E-mCherry), Tg(Fap7:Aqp4-GFP) and Tg(Coro1a:lynk-GFP) were used to label astrocyte cytoplasm, extracellular membranous debris, water channels, and myeloid cell membranes, respectively. Anesthetized 7-10 day old X. laevis tadpoles were mounted on custom molds and their ONs imaged using a Leica Dragonfly spinning disc confocal microscope either as z-series spanning the entire thickness of the ON, single section t-series, or combination z- t- series (6D-imaging).
Results :
Results: Extensive foci of Mfge8-tagged debris occur within every ON imaged. These foci exhibit nondirectional, seemingly diffusive random movement. Much debris localizes to the periphery of the ON, but some is also found in discrete clusters within the ON parenchyma, despite the tadpole ON parenchyma being avascular. Within the ON parenchyma two discrete types of clusters are apparent: linear structures that appear to be extracellular spaces lined by Aqp4, or larger more amorphous structures that prove to be myeloid cells, as shown by Coro1a labeling. 6D-imaging shows that these phagocytic myeloid cells migrate in and out of the ON parenchyma and extravasate in or out of nearby vessels over the course of minutes.
Conclusions :
Conclusion: Debris within the Xenopus tadpole ON parenchyma accumulates in extracellular spaces lined by Aqp4 and is taken out of the ON by infiltrating myeloid cells. Should a similar debris-clearance mechanism also occur in the normal mammalian ON head, involving astrocyte created water channels and infiltration of blood-borne myeloid cells (a hallmark of inflammation), it would have very significant implications for glaucoma diagnosis and treatment.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.