Fig. 5

Analysis of HCAR expression in D2 and D2G retina and ON. a, b Bar graphs showing Hcar1 (a), ***p = 0.0001, and Hcar2 (b) mRNA normalized to Hprt mRNA by qRT-PCR; n = 8 ON per group. c Immunohistochemical analysis of HCAR1 (red) expression in the untreated (Unt) and keto D2 retina and ON. GFAP (green) labels Müller glia and astrocytes, Iba1 (cyan) labels microglia, and DAPI (blue) stains nuclei. Arrows indicate colocalization of HCAR1 with Iba1+ microglia and GFAP+ astrocytes. d, e Percentage of mean fluorescence of HCAR1 in the inner retina (d), **p = 0.0036, and in the ROI of the proximal ON (e), ***p = 0.0001; five retinae per group, with five sections/retina. f Western blot analysis of HCAR1 protein level in retina. g Quantification by densitometry of HCAR1 protein normalized to β-actin, ***p = 0.0008; n = 3 blots per group, each with independent samples. h HCAR1 protein in the ON normalized to β-actin levels, as determined by capillary electrophoresis, **p = 0.0025; n = 3 ON per group. All bar graphs are presented as the mean ± SEM, n = 5, analyzed by two-tailed unpaired t test. Scale bar, 20 μm