Axons of retinal ganglion cells are insulted in the optic nerve early in DBA/2J glaucoma

Abstract

Here, we use a mouse model (DBA/2J) to readdress the location of insult(s) to retinal ganglion cells (RGCs) in glaucoma. We localize an early sign of axon damage to an astrocyte-rich region of the optic nerve just posterior to the retina, analogous to the lamina cribrosa. In this region, a network of astrocytes associates intimately with RGC axons. Using BAX-deficient DBA/2J mice, which retain all of their RGCs, we provide experimental evidence for an insult within or very close to the lamina in the optic nerve. We show that proximal axon segments attached to their cell bodies survive to the proximity of the lamina. In contrast, axon segments in the lamina and behind the eye degenerate. Finally, the Wlds allele, which is known to protect against insults to axons, strongly protects against DBA/2J glaucoma and preserves RGC activity as measured by pattern electroretinography. These experiments provide strong evidence for a local insult to axons in the optic nerve.

Distal axonopathy with structural persistence in glaucomatous neurodegeneration

“…Labeling of the retina in whole-mount preparations, sections of optic nerve, and brain was performed as previously described (35). We used antibodies against detyrosinated α-tubulin (monoclonal, 1:200; Millipore) and phosphorylated heavy-chain neurofilament (SMI31, 1:1,000; Sternberger Monoclonal) (8,15) to visualize RGCs; myelin basic protein (SMI99, 1:1,000; Sternberger Monoclonal) to delineate the optic nerve head; ERRβ (polyclonal, 1:500; Sigma) (29) to label the RGC SC projection; hyperphosphorylated heavy-chain neurofilament (SMI34, 1:1.000; Sternberger Monoclonal) (15) to highlight axonal dystrophies; and VGluT2 (polyclonal, 1:500; Synaptic Systems) (32) to visualize RGC axon terminals in the SC. We used Alexa Fluor-(Invitrogen) or Cy5-(Jackson ImmunoResearch) conjugated secondary antibodies (1:200).…”

“…DBA/2 mice present agedependent elevations in IOP as a result of pigmentary dispersion in the anterior eye that are linked to RGC degeneration (14,15). Retrograde transport from the SC persists as late as 18 months of age in the DBA/2 mouse (16,17).…”

Early immune responses are independent of RGC dysfunction in glaucoma with complement component C3 being protective

“…Importantly, glaucoma in D2 mice is an axonopathy, as evidenced by substantial RGC protection in the presence of the Wallerian degeneration slow allele (D2.Wld s ) (20). RGCs from D2.Wld s mice can survive high IOP and have an intact pattern electroretinogram (PERG) (20). Given that PERG amplitude is a sensitive measure of RGC health (32, 33), Wld s prevents very early changes.…”

“…It is not known whether the earliest immune responses are protective or damaging, or which events irreversibly damage the optic nerve. Moreover, it is not known whether the immune responses are secondary to RGC dysfunction or occur independently, possibly as a more direct result of IOP elevation.In glaucoma, an early insult to RGC axons occurs where they exit the eye in the optic nerve head (ONH) (16)(17)(18)(19)(20). Modeling shows that an increase in IOP inside the eye leads to increased strain in this critical region (21,22).…”

“…In glaucoma, an early insult to RGC axons occurs where they exit the eye in the optic nerve head (ONH) (16)(17)(18)(19)(20). Modeling shows that an increase in IOP inside the eye leads to increased strain in this critical region (21,22).…”