Fig. 4

(A-A’) Hair cells from wildtype 5 dpf fish before and during a 2 s fluid-jet stimulus. The spatial patterns of the evoked calcium influx during stimulation (A’ GCaMP6s ΔF, indicated via the heatmaps) compared to prestimulus (A) reveal synaptically active cells (white stars). (A”-A’’’) Hair cells depicted in A and A’ after a 30-min treatment with 75 µM neomycin solution. Magenta star demarcates an active cell that did not survive neomycin treatment. (B) A higher percentage of active hair cells survive a 30-min treatment with two different neomycin concentrations (75 and 100 µM) compared to inactive cells. Neuromasts were examined at 5 dpf immediately after washout of neomycin solution and application of FM 4–64. (C) Outline of DAPI labeling protocol used to differentiate between older and younger cells within a given neuromast. DAPI is used to label mature hair cells at 3 dpf. At 5 dpf, the hair cells are assessed. (D-D’) Hair cells from wildtype 5 dpf fish before and during a 2 second fluid-jet stimulus. The spatial patterns of the evoked calcium influx during stimulation (D’ GCaMP6s ΔF, indicated via the heatmaps) compared to prestimulus (D) reveal synaptically active cells (white stars). (D’’-D’’’) Hair cells depicted in D and D’ showing DAPI-positive older and DAPI-negative younger cells. (E) DAPI-positive older cells make up a much greater percentage of the inactive cell population than the active cell population. A minimum of five animals were examined per experimental group at 5 dpf. Each dot in B and E represents one neuromast. Error bars: SEM. For comparisons, a two-way ANOVA with a Sidak’s correction for multiple comparisons was used in B. A Mann-Whitney test was used in E. **** p<0.0001. Scale bar = 5 µm.