Figure 4. Decreased mitophagy and autophagy in aging zebrafish retinas. (A) Pink1 gene expression in young and aging zebrafish retinas as measured by quantitative real-time PCR (mean ± SEM, ***P<0.001, n=3). (B) Representative western blot showing the protein expression level of Pink1 in young and aging zebrafish retinas. (C) Graph of the densitometric mean and SEM normalized to the corresponding level of the loading control protein beta-tubulin (**P<0.01, n=3). (D) Representative western blot showing the protein expression levels of LC3B-II and LC3B-I in young and aging zebrafish retina. (E) Graph of the ratio of LC3B-II to LC3B-I protein levels as determined by densitometry (**P<0.01, n=3). (F) Immunofluorescence analysis of Pink1 in the RGC layer of young and aging zebrafish retinas. Retina cross sections prepared from zebrafish eyes were immunostained with Pink1 antibody. All photographs were taken at 40x magnification except the rat sections that were at 20x. Young, 4-6 months old; aging, 19-23 months old; NC, negative control (without primary antibody); Rat, positive control; GCL, ganglion cell layer.