Fig. 3

Fig. 3

Pgrn-a knockdown results in microphthalmia and diminished neuronal differentiation. (A–R) Sections through central retina of uninjected (UI, A–C); standard control morpholino-injected (SC, D–F); SS MM (G–I) and 5′UTR MM (J–L), embryos injected with 5-nucleotide mismatch control morpholinos; SS MO (M–O) and 5′UTR MO (P–R), embryos injected with e3i3 splice site-targeted or 5′UTR-targeted morpholinos, respectively, at 72 hpf. Sections are immunolabeled (cyan) with markers of differentiated ganglion cells (Zn5, top row), amacrine cells (HPC1, middle row), and red–green double cone photoreceptor cells (ZPR1, bottom row), EdU (fusia), and DAPI (gray). (S) Histogram showing relative retinal area in UI (7945.8 ± 1319.6 μm²; n = 11; CI, 7945 ± 886), SC-injected (7594.3 ± 475.2 μm²; n = 9; 95% CI: 7594 (7229–7959)), SS MM MO-injected (7354 ± 1062.6 μm²; n = 11; 95% CI: 7353 (6640-8066)), and 5′UTR MM MO-injected (7610.8 ± 547.6 μm²; n = 10; 95% CI: 7610 (7219–8001)), SS MO (6044.7 ± 956.2 μm²; n = 10; 95% CI: 6044 (5360–6782)), and 5′UTR MO (5188.7 ± 891.8 μm²; n = 16; 95% CI: 5188 (4713–5663)) retinas at 72 hpf; ***p≤0.001. (T) Histogram showing the percent of the retina labeled with EdU in UI (9.3 ± 4.3%; n = 11; 95% CI: 9.3 (6.4–12.2)), SC-injected (12 ± 3.1%; n = 9; 95% CI: 12.0 (8.7–15.3)), SS MM MO-injected (10.8 ± 3.5%; n = 11; 95% CI: 10.8 (8.7–12.9)), 5′UTR MM MO-injected (9.8 ± 2.3%; n = 10; 95% CI: 9.8 (7.3–12.3)), SS MO-injected (27.5 ± 11.5%; n = 10; 95% CI: 27.6 (25.9–29.3)), and 5′UTR MO-injected (41.1 ± 13%; n = 16; 95% CI: 41.1 (35–47.2)) retinas at 72 hpf; ***p≤0.001. (U) Histogram showing the number of microglia in uninjected (UI; 55.4 ± 21.5 cells; n = 20; 95% CI: 62.4 (51.7–73.1)), 5′UTR mismatch morpholino injected (5′UTR MM; 43.4 ± 4.6 cells; n = 8; 95% CI: 43.4 (39.6–47.2)), and Pgrn-a morphant (5′UTR MO; 4.4 ± 3.4 cells; n = 11; 95% CI: 4.4 (2.1–6.7)) retinas at 72 hpf; ***p≤0.001. Quantitative data are represented as mean; error bars represent the standard deviation. Outer nuclear layer (ONL), inner nuclear layer (INL), and ganglion cell layer (GCL); ciliary marginal zone (CMZ, brackets). Scale bar equals 50 µm.