Figure 5

Tubastatin A treatment rescues visual function and retinal morphology defects in the dye zebrafish model of inherited blindness. (a) Dye mutants and siblings were treated with 100 μM tubastatin A (TST) from 3 to 5 dpf, lysates were prepared from 50 larval eyes each and subjected to immunoblot analysis using antibodies against α-tubulin (α-Tub), acetylated α-tubulin (ac Tub) and actin as a loading control, n=3. (b) dye mutants treated with TST revealed significantly improved OKR when compared with vehicle (0.1% DMSO)-treated mutants, error bars show standard error of the mean. (c) Summary of VMR activities. Treatment with 100 μM TST significantly improves the MAX OFF (black bars) and MAX ON (yellow bars) activities in dye mutants. For visual function data, N=12 (number of larvae per replicate), n=3 (number of replicates), error bars show standard error of the mean. (d–i) Light micrographs of retinal sections derived from TST (g–i) and vehicle-treated dye larvae (d–f). Treatment with 100 μM TST reduces the number of dying cells in the ciliary marginal zone (red square in g and h) indicated by the presence of pyknotic nuclei compared with control larvae (red square in d, red arrow in e), scale bars represent 100 μm. The number of pyknotic nuclei present in multiple sections through the central retina is also reduced (bar chart in j). Photoreceptor outer segment length is marginally improved following TST treatment (i) compared with controls (f), n=5. Statistical analyses were performed using a multiple comparison Student’s t-test with unequal variances comparing dye, 0.1% DMSO groups to TST-treated larvae, ***P<0.001, **P<0.01 *P<0.05