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. Author manuscript; available in PMC: 2018 Feb 17.
Published in final edited form as: Exp Eye Res. 2017 Feb 17;155:121–127. doi: 10.1016/j.exer.2017.02.007

Figure 5.

Figure 5

Increased levels of all-trans retinal after bleaching result in increased formation of lipofuscin precursors in rod photoreceptors isolated from Sv/129 (○, ●; n = 72 cells total) and Abca4−/− (△, ▲; n = 50 cells total) mice. Lipofuscin precursor fluorescence was measured in isolated dark-adapted rods (●, ▲); subsequently, the cells were bleached with long-wavelength (>530 nm) light for 1 min and rod outer segment fluorescence signals from all-trans retinol, all-trans retinal and lipofuscin precursors measured at 30 min after bleaching (○, △). Retinol and retinal fluorescence were obtained by exciting with 340 and 380 nm light and collecting emission >420 nm. Lipofuscin precursor fluorescence was obtained with 490 nm excitation and collecting emission >515 nm. All-trans retinal accumulation was measured as the Fex-340/Fex-380 ratio. The value Fex-340/Fex-380 = 0.55 corresponds to 100% retinal-0% retinol; Fex-340/Fex-380 = 6.95 corresponds to 0% retinal-100% retinol. Different levels of all-trans retinal were brought about by using conditions that suppress the supply of NADPH to the rod outer segment. Conditions were: a, 5 mM glucose; b, 0.5 mM glutamine; c, 50 μM glucose; d, 50 μM glutamine; e, no substrate; f, broken-off rod outer segments. Numbers of cells were: for wild type; a, n = 9; b, n = 7; c, n = 17; d, n = 16; e, n =12; f, n =11; for Abca4−/−; a, n = 9 cells; b, n = 8; c, n = 8; d, n = 9; e, n =9; f, n =7. Experiments at 37 °C. Error bars denote standard errors. The dashed lines through the data points represent linear regression lines.