Fig. 3

Coablation results in delayed MG and progenitor cell proliferation. (A) Schematic of the assay timeline and the paradigm for rod cell and macrophage/microglia coablation for data shown in B–D (dots on the Histo line indicate time points as in Fig. 2). (B–B′′) Representative retinal section of a 7-dpf rho:YFP-NTR;mpeg1:Tomato;gfap:GFP larva immunostained for PCNA (blue) after 24-h exposure to Mtz. Enlarged views of boxed regions 1 and 2 show evidence of MG (green cells) phagocytosis of microglia (red cells) containing rod cell debris (yellow cells). Boxed region 3 shows an example of a proliferating MG cell following rod cell ablation (blue/green double-positive cell). Note: The prominent red signal outside the ONL in B′ and B′′ (asterisks in B′) is autofluorescence from the retinal pigment epithelium (RPE) layer and blood vessels. (C and D) Quantification of proliferating (PCNA-immunoreactive) MG (GFP⁺/PCNA⁺) and presumptive progenitor cells (PCNA⁺ only) in the INL (C) and ONL (D) following rod photoreceptor ablation (C, green boxes; D, yellow boxes) or rod cell and macrophage/microglia coablation (C and D, red boxes) normalized to the number of cells per 1 mm of retina (measured at the OPL). Note that retinal microglia were eliminated from the analysis of proliferating cells based on expression of the Tomato transgene (arrowheads in B′). (C) An early deficit in INL proliferation rates is evident in the coablated condition during Mtz treatment, but increased proliferation at later time points appears to compensate. (D) Deficits in ONL progenitor proliferation rates in coablated retinas persist through the second day of recovery before matching the levels observed in retinas in which only rod cells were ablated. Statistics (effect sizes, 95% confidence intervals, and P values for paired comparisons) are provided in Table S2; bolded letters denote pairs showing reproducible differences, italicized letters denote pairs with no discernible differences. Co-Abl, coablated retinas; Rod-Abl, rod photoreceptor ablated retinas.