Zebrafish Hair Cell Development and Regeneration

For hair cell development analysis, the embryos at 5 dpf were used for hair cell staining and counting. For hair cell regeneration analysis, the embryos at 5 dpf were exposed to 10 μM of copper sulfate (Sigma, Cat# 451657) for 2 h, allowed to recover for 48 h except when otherwise indicated, and then used for hair cell staining and counting. Hair cell staining by YO-PRO-1 (Thermo Fisher Scientific, Cat# Y3603) was done as previously described [18 (link)]. The stained embryos were oriented for lateral views in a 96-well plate for counting and imaging with a fluorescent microscope. Hair cells per neuromast presented in graphs were obtained by averaging the hair cell counts from four neuromast per embryos at the P1, P2, P4, and P5 positions [45 (link)] from multiple embryos. P2 neuromast was not counted, since it had a greater variation in the number of hair cells. For studying genotype and phenotype correlation, 24–32 embryos obtained from a single pair of heterozygote incross were analyzed and then genotyped. For studying the effect of chemical treatment, the number of wild-type embryos used for each data point was 10, except when otherwise indicated.