<b>Knock-down </b><i>foxi3a</i> Expression Severely Reduces Epidermal Ionocyte Progenitor Number and Abolishes the Later Differentiation Program.

<p>(A–G) Interfering with <i>foxi3a</i> and <i>foxi3b</i> functions of epidermal ionocyte differentiation by a morpholino (0.5 mM/embryo) injection. Morphants were fixed at 24 hours post fertilization (hpf) and stained with <i>atp1b1b</i> (green), <i>ca2a</i> (red), and P63 (blue) to detect Na<i>⁺</i>,K<i>⁺</i>-ATPase rich cells (NaRCs), H<i>⁺</i>-ATPase rich cells (HRCs), and epidermal stem cells, respectively. (H–I) A rescue experiment to show the specificity of <i>foxi3a</i> morpholinos. The <i>foxi3a</i> mRNA used for the rescue experiment does not contain a binding site for MO2. (J–L) Comparison of the vital dye uptake ability between the wild type (wt), <i>foxi3a</i> morphants, and <i>foxi3b</i> morphants. NaRCs and HRCs in either wild-types or <i>foxi3b</i> morphants can absorb MitoTracker (red) and Con-A (green) through to their apical openings. For <i>foxi3a</i> morphants, no MitoTracker or Con-A staining was detected due to blockage of the entire differentiation program. (M–O) Detection of the apical opening of epidermal ionocytes in wild-types, <i>foxi3a</i> morphants, and <i>foxi3b</i> morphants by scanning electron microscopy. The apical openings of NaRCs and HRCs in wild-types are shaped as deep holes (green box) or a mesh (red box), respectively. The apical openings were totally undetected in <i>foxi3a</i> morphants due to blockage of the entire differentiation program. For <i>foxi3b</i> morphants, the apical openings for both NaRCs and HRCs were reduced. Embryos in (A–I) were scored at 24 hpf, while in (J–O), they were scored at 72 hpf. In I, embryos are orientated in a dorsal-up and anterior-top position.