(A, F) Timelines of single-ablation (A) and double-ablation (F) proliferation experiments. (B) Maximum projections of mock- (Mock) and neomycin-treated (Neo) neuromasts. EdU-positive cells are shown in magenta, anti-Parvalbumin-stained hair cells are shown in green, and DAPI-stained nuclei are shown in blue. Arrowheads indicate EdU-positive support cells. Scale bar = 10 μm. (C) Percentage of hair cells per neuromast labeled by EdU. Mock: 6.11 ± 8.69, n = 50 neuromasts (10 fish); Neo: 78.24 ± 20.69, n = 45 neuromasts (nine fish); mean ± SD; Mann Whitney U test, p<0.0001. (D) Total EdU-positive cells per neuromast. Mock: 2.78 ± 1.84, n = 50 neuromasts (10 fish); Neo: 8.18 ± 3.07, n = 45 neuromasts (nine fish); mean ± SD; Mann Whitney U test, p<0.0001. (E) Percentage of EdU-positive cells per neuromast that are either hair cells or support cells. Mock: 29.73% hair cells, 70.27% support cells, n = 50 neuromasts (10 fish); Neo: 72.02% hair cells, 27.98% support cells, n = 45 neuromasts (nine fish); mean ± SD. (G–N) Individual slices of a neuromast following two regenerations at two different planes: apical hair cell layer (G–J) and basal support cell layer (K–N). EdU (visualized by a Click-iT reaction) is labeled in magenta, BrdU (anti-BrdU) is labeled in cyan, and myo6b:GFP hair cells are labeled in green. Arrowheads indicate EdU/BrdU-positive hair cells, and asterisks indicate EdU-positive support cells. Scale bar = 10 μm.
Figure 1—source data 1. Hair cell progenitors are replenished via proliferation of other support cells.