Abstract
Purpose :
A study in mice postulated the existence of a glymphatic-like system connecting eye to optic nerve based on finding intravitreally injected labeled amyloid beta (Ab) outside of axons in the optic nerve. To determine whether Ab might have instead gotten there through the axonal transcellular degradation pathway we previously described for mitochondria, we carried out similar experiments using an experimental system that allows for optic nerve live imaging.
Methods :
Dye labeled Ab and/or Mitotracker dyes were injected intravitreally in 1 week-old Xenopus laevis tadpoles expressing combinations of transgenes: a retinal ganglion cell promoter driving a membrane or mitochondria reporter; a astrocyte promoter expressing a membrane reporter, or Aqp4-GFP, Mfge8-mCherry or CD63-mCherry fusion constructs; a microglia/myeloid cell promoter driving a membrane reporter. Optic nerves in live animals were imaged by spinning disc confocal microscopy. Movement of Ab and mitochondria, imaged for 1 min at 1Hz, were analyzed by kymographs. The fraction of intravitreally injected Ab or mitotracker found in the optic nerve outside of axons, and within astrocytes or microglia/myeloid cells, was assessed by imaging the full thickness of the optic nerve at 1 µm steps, followed by measurements using Imaris software.
Results :
Within the optic nerve, 93% of intravitreally injected Ab co-localized with mitochondria, with 50% being stopped together. 10% of Ab- and 8% of mitochondria-dyes were found outside axons, mostly together, of these approximately 10% fully outside nerve parenchyma. The majority of extra-axonal Ab or Mitochondria were found, largely together, in a discrete set of hyperphagocytic astrocytes, uniquely identified by their redistribution of a Aqp4-GFP transgene and their high expression of Mfge8- and CD63- transgenes, or alternatively were found, largely together, in microglia/myeloid cells, where they also partially co-localized with astrocytic membrane markers.
Conclusions :
At least in X. laevis tadpoles, intravitreally injected Ab reaches the outside of the optic nerve by traveling together with mitochondria within axons, and then leaving axons through the axonal transcellular degradation process that we first described in the mouse optic nerve head. Discrete astrocytes assume a hyperphagocytic state and are responsible for most clearance of axonal mitochondria and Ab.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.