Rodent Anterior Ischemic Optic Neuropathy (rAION) Induces Regional Retinal Ganglion Cell Apoptosis with a Unique Temporal Pattern

Abstract

Purpose Nonarteritic anterior ischemic optic neuropathy (NAION) results in optic nerve damage with retinal ganglion cell (RGC) loss. An NAION model, rodent anterior ischemic optic neuropathy (rAION), was used to determine AION-associated mechanisms of RGC death and associated regional retinal changes. Methods rAION was induced in male Wistar rats, and the retinas analyzed at various times after induction. RGCs were positively identified by both retrograde fluorogold labeling and brain-expressed X-linked protein-1/2 (Bex1/2) immunoreactivity. RGC death was analyzed by fluorescein-tagged annexin-V labeling (FITC-annexin-V), as well as by terminal nucleotide nick-end labeling (TUNEL). Retinal flatmount preparations enabled regional retinal analysis of labeled dying cells. Apoptosis pathway activation was confirmed by Western analysis, with an antibody that recognizes cleaved caspase-3. Results Post-rAION, RGCs die by apoptosis over a longer period than previously recognized. Cleaved caspase-3 immunoreactivity was greatest between 11 and 15 days. rAION-induced RGC death occurs regionally, with sparing of large contiguous regions of RGCs. Conclusions rAION results in later RGC death than in traumatic optic nerve damage models. Apoptosis, measured by FITC-annexin, occurs maximally in the second to third week after infarct. Cleaved caspase-3 activation confirms that after rAION, RGCs undergo apoptosis by the caspase activation pathway. The regional pattern in dying RGCs after rAION implies that a measure of retinotopic organization occurs in the rodent optic nerve. The prolonged period from insult to death suggests that the window for successful treatment after ON infarct may be longer than previously recognized.

Subretinal fluid is common in experimental non-arteritic anterior ischemic optic neuropathy

“…This has been achieved in mice, rats, as well as primates. [10][11][12][13][14][15] Subretinal fluid has not been described in the animal models of NAION.…”

Functional rescue of experimental ischemic optic neuropathy with αB-crystallin

“…[5][6][7][8]10,11,17 After rose bengal (1.25 mM in PBS, 2 ml per g body weight) injection via tail vein, the ON head was exposed to 20 light spots (frequency doubled Nd:YAG laser, Pascal, OptiMedica, Santa Clara, CA, USA), each with 100 mm diameter (equivalent to 39 mm on the nerve), at a power of 35 mW and duration of 500 ms. We delivered small laser spots at low power and shorter duration circumferentially around the ON head in order to focus more precisely while covering the entire ON head. The degree and extent of whitening due to ischemia 18 are similar to that of the Bernstein model in our direct comparison.…”

“…Photochemical thrombosis animal models of AION have been instrumental in elucidating the events following ON head ischemia. [5][6][7][8][9] In experimental AION, there is early glial and axonal dysfunction, with ON oligodendrocyte apoptosis occurring by day 6 and progressive nerve demyelination within 2 weeks. 5,8,9 Retinal ganglion cell (RGC) loss peaks during weeks 2 and 3.…”

“…5,8,9 Retinal ganglion cell (RGC) loss peaks during weeks 2 and 3. 7,8,10,11 During these 3 weeks of critical period for treatment considerations, there are important signs of self repair, such as the upregulation of retinal heat shock proteins HSP 70, 84, and 86. 5,8 Heat shock proteins act as molecular chaperones and are thought to have important roles in optic neuropathies, retinal diseases, and central nervous system inflammatory and neurodegenerative diseases.…”

“…5,8 Heat shock proteins act as molecular chaperones and are thought to have important roles in optic neuropathies, retinal diseases, and central nervous system inflammatory and neurodegenerative diseases. [12][13][14][15][16] In this study, we used a mouse model for AION via photochemical thrombosis [5][6][7][8]10,11,17 to examine the early changes following ON head ischemia. We then assessed the effectiveness of a small heat shock protein named aB-crystallin (aBC) in the treatment of experimental AION, using functional and histological approaches.…”