Figure 3.

Immunohistochemistry confirmed alterations in diurnal patterns of autophagic gene expression upon light cycle disruption. Mice were separated into two groups. One group was kept under 12/12-h light/dark cycle and the other group was kept in the dark for 48-h (dark/dark), before being euthanized. Samples were collected every 3 h for 24 h and were analyzed by immunohistochemistry for autophagic markers (A) ATG9 (B) LC3 (C) ATG7 and (D) Beclin1 (BECN). Autophagy protein expression is green, TRITC agglutinin for vessels red and DAPI nuclear staining blue. The arrows indicate autophagy protein localization to retinal vessels. RGC, retina ganglion cell; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; RPE, retinal pigment epithelium. The error bars in the circadian plots represent the mean + SEM and diurnal oscillation had a p-value less than 0.05.