Fig. 3

Glucose dynamics in photoreceptors with chronically active HIFs. a Preparation of acute retinal slices from flat mounted half retinas and imaging of the glucose sensor FLIIP by TPLSM. b Expression of FLIIP in PRs after subretinal AAV application (left) or in cells of the inner retina after intravitreal AAV application (right). Scale bar, 50 \(\upmu\)m. c Schematic representation of glucose transport inhibition by cytochalasin B allowing to monitor glycolytic rate through the hexokinases. d FLIIP traces (mean ± SD) in PRs from \(rod^{\varDelta \ Vhl}\) (orange; 19 retinal slices from 5 mice) and ctrl mice (black, 16 retinal slices from 4 mice) during inhibition of glucose transport with cytochalasin B followed by aglycemia (zero glucose). Traces were normalized to zero glucose (values at the end of experiment). e FLIIP traces (mean ± SD) in cells of the INL (blue; 10 retinal flatmounts from 7 mice) and GCL (green; 16 retinal flatmounts from 7 mice) during inhibition of glucose transport with cytochalasin B followed by aglycemia (zero glucose). Traces were normalized to zero glucose (values at the end of experiment). f Basal glucose levels in PRs and cells of the INL and GCL in \(rod^{\varDelta \ Vhl}\) and ctrl mice as calculated from the FLIIP traces shown in (d) and (e). Shown are individual values and means ± SD. g Glucose consumption rate in PRs and cells of the INL and GCL in \(rod^{\varDelta \ Vhl}\) and ctrl mice as calculated from the FLIIP traces shown in (d) and (e). Shown are individual values and means ± SD. h Comparison of FLIIP raw FRET signals in PRs and cells of the INL and GCL in \(rod^{\varDelta \ Vhl}\) (orange) and ctrl (black) mice at zero intracellular glucose. Statistics: Mann-Whitney nonparametric test for comparison of \(rod^{\varDelta \ Vhl}\) and ctrl mice. Kruskal-Wallis nonparametric test and Dunnett’s multiple comparison test for comparison of PRs, GCL and INL cells. PS: photoreceptor segments. ONL: outer nuclear layer. OPL: outer plexiform layer. INL: inner nuclear layer. GCL: ganglion cell layer