Morphological abnormalities and progressive retinal degeneration in mzbbs1−/− mutants starting at 7 dpf.
a, c Representative images of semi-thin plastic sections stained with Richardson solution in wildtype (top) and mzbbs1−/− (bottom) retina at 7 dpf (a) and 10 dpf (c) showing slight morphological abnormalities at 7 dpf and more severe changes at 10 dpf with shorter and misshapen OSs. b, d DiO-staining highlighting OSs reveals progressive morphological alterations of the OSs between 7 dpf (b) and 10 dpf (d). e TEM reveals bulky OSs (*asterisk) with abnormal membrane disc stacking in mutant retina at 7 dpf. Disc membranes in mutants (middle & right image) are less compact in places or stacked vertically compared to controls (red arrowheads). Membrane-filled swirls are frequently observed in the RPE (yellow arrows) at that stage (middle & right image are two images of mzbbs1−/− retinae showing various alterations seen in the mutants). f At 8dpf, UV-Opsin (green) localizes normally to the OS in mzbbs1−/− mutants and controls, despite abnormal ultrastructure and shortened OS revealed by the membrane dye BODIPY (magenta). g Top panel shows bar plots of the maximum b-wave amplitude by electroretinography at 10 dpf. Reduced b-wave intensity at various light intensities demonstrates severely decreased response to light. Unpaired two-tailed multiple T-test; Sig: ns = FDR(q value) = 0.089, **** = FDR(q value) < 0.0001; Sample size (n = 13 WT, n = 8 Mut larvae), Error bars show standard deviation around the mean. The bottom panel shows the average response curve after a 100 ms light flash at log-2 intensity (Light On/Off line below the ERG curve) for mutant (grey) compared to control (black). Scale Bars: (a,c) 12 µm, (b, d, f) 50 µm, (e) 5 µm. Abbreviations: OS outer segment, ERG electroretinography, INL inner nuclear layer, M mitochondria, N nucleus, ONL outer nuclear layer, OPL outer plexiform layer, P pigment, TEM transmission electron microscopy.