Figure 3.

Use of two-photon imaging for ophthalmic drug screening. (a) Ret–NH₂ protects RPE of 1-month-old Abca4^−/−Rdh8^−/− mouse from bright light induced accumulation of fluorescent granules. Representative ex vivo images obtained 7 and 14 days after bright light exposure; images obtained with a ‘through the sclera’ configuration are included for comparison. Excitation with 730 nm was used for the upper row images whereas 850 nm was employed for the lower row. (b) Individual rod photoreceptors expressing rhodopsin-GFP fusion protein are visible in photoreceptor layer of 2-month-old hrhoG/hrhoG mice. (c) Two-photon excited emission spectra from fluorescent granules in the RPE of Abca4^−/−Rdh8^−/− mouse obtained through the sclera (black) and pupil (red). Spectrum from photoreceptors in hrhoG/hrhoG mice is shown in gray. (d) Quantification of Ret–NH₂ impact on accumulation of fluorescent granules in the RPE, based on images as shown in (a); ND stands for none detected; error bars indicate S.D., n = 3. (e) Lower zoom image of the RPE in 6-week-old mouse not treated with Ret–NH₂, showing the optic disc is displayed in upper panel. Lower panel shows a magnified view from RPE area outlined with white rectangle in the upper image. Scale bars represent 30 μm in (a, b) and lower panel of (e) and 220 μm in the upper panel of (e).