Abstract
Purpose :
Reactivity of astrocytes in the retina and optic nerve head (ONH) is observed in glaucoma and other optic nerve injuries. Astrocytes are densely interconnected by gap junction (GJs) formed by connexin 43 (Cx43). The role of astrocytic connectivity on glial reactivity and RGC survival in optic nerve degeneration is not fully understood. Here, we assessed expression of Cx43 with sustained intraocular pressure (IOP) elevation and after optic nerve crush injury. We also examined whether deletion of Cx43 in astrocytes impacted RGC loss and ONH astrocyte reactivity in the two injury models.
Methods :
The astrocyte-specific Cx43 knockout (KO) mouse was established using the Cre-loxP recombination system. Experimental glaucoma was induced in C57BL/6 (WT) and Cx43KO mice by intracameral injection of polystyrene microbeads. Optic nerve of one eye was crushed for 10 seconds using fine curved forceps ~ 1 mm behind the eyeball. Immunohistochemistry was used to assess Cx43 expression, RGC survival (using Brn3a labeling), and astrocyte reactivity in the ONH (using GFAP labeling), after 4 and 8 weeks of IOP elevation and 7 days post nerve crush. Student’s t-test was used for statistical analyses.
Results :
IOP elevation for 8 weeks produced a 35% increase in astrocytic Cx43 expression in the retina (n=5, p=0.001) and ONH (n=3, p=00001). Similarly, Cx43 expression was increased by 30% after optic nerve crush injury (n=5, p=0.0001). Deletion of Cx43 in astrocytes was neuroprotective in glaucomatous injury, with RGC survival increasing by 57% at 4 weeks (n=5-8, p=0.03) and 63% by 8 weeks (n=7-8, p=0.0003) compared to control eyes. The absence of Cx43 also increased viable RGCs after optic nerve crush by twofold compared to control eyes (n=8, p=0.003). Finally, Cx43 deletion caused a decrease in astrocytic reactivity in the ONH of bead injected eyes compared to WT (n=3, p<0.05). However, astrocyte reactivity in the retina was unaffected by Cx43 deletion (n=5-6, p=0.55).
Conclusions :
Results in two different models of optic nerve injury indicate that astrocytic Cx43 channels contribute significantly to RGC death. There is an upregulation of Cx43 in astrocytes in both models, and a reduction in astrocyte reactivity in the ONH upon Cx43 deletion, suggesting that GJ coupling might lead to the spread of signals associated with astrocyte reactivity.
This is a 2021 ARVO Annual Meeting abstract.