FIGURE 3.

Valproic acid reduces astrocyte morphological changes associated with inflammation and gliosis. (A) Retinal explants were exposed to LPS, VPA, or untreated and assessed at D1 in comparison to D0 controls. Individual GFAP fibers were reconstructed to generate astrocyte networks using Imaris software. Areas enclosed by these fibers were generated in Imaris to measure space filling (individual areas are demonstrated by a unique color). (B) At D1 there was a significant increase in the number of filaments and the normalized volume, with no significant change in total filament length. This suggests a thickening of GFAP filaments and an increase in filament intersection. Supporting this, the number of enclosed areas was significantly increased, and these were significantly smaller (decreased enclosed area) suggesting an increase in filament volume and space filling by the astrocyte network, as is typical in gliosis. Addition of LPS to the media had no significant effect on astrocyte morphology compared to those in untreated D1 retinas. Addition of VPA significantly attenuated changes to the astrocyte network, with a reduction in the number of filaments, total filament length, and the normalized volume relative to untreated D1 retinas. The number of enclosed areas was significantly reduced, and the area of these was significantly higher than in untreated D1 retinas. There was no significant difference to D0 controls. Together, these suggest a protection against astrocyte morphological change and attenuation of pro-inflammatory and gliotic astrocyte changes. GFAP = astrocyte specific marker in NFL when labeled as flat mounts, n = 4 retinas for all conditions, scale bars = 20 μm, * = p < 0.05, ** = p < 0.01, *** = p < 0.001, NS = non-significant. Individual enclosed areas demonstrated by condition as violin plots.