Fig. 3

Immunostaining of HuC/D and GFP in the gut of Tg(chata:GGFF2) at embryonic, larval and adult stages. A–F": The expression of GFP is shown in the top row (A–F), that of HuC/D in the middle row (A′–F′), and merged images in the bottom row (A″–F″). The stages are shown at the top right. HuC/D was expressed in most GFP + cells (white arrowheads) at all stages examined, whereas some GFP + cells were HuC/D− (green arrowheads), and some HuC/D+ cells were GFP− (magenta arrowheads). GFP + cells with very weak HuC/D found at 60 hpf (A) are indicated by green arrowheads outlined with white. GFP + cells were decreased in the young (1.5-month-old) adults (E), and not found in mature (∼1-year-old) adults (F). We found that 96% of the GFP + cells were HuC/D+ at 6 dpf (n = 6 animals, 346/360 cells) and that 88% of the GFP + cells were HuC/D+ at 12 dpf (n = 7, 383/434 cells) (B-B″ and C-C″). In the 1.5-month-old adults (E-E″), 100% of the GFP + cells were HuC/D+ (n = 2 fish, 107/107 cells), and the percentage of GFP + cells among HuC/D+ cells was 33% (n = 2, 107 GFP+/326 HuC/D+ cells), which is a much smaller percentage than that observed at 12 dpf (60%, n = 7, 383 GFP+/639 HuC/D+ cells). In the 1-year-old adults (F-F″), only 0.5% of the HuC/D+ cells were GFP+ (n = 2, 2 GFP+/433 HuC/D+). Anterior: to the left. White dotted lines in B″ outline the gut. For A-C and E-F, scale bar in F": 50 μm. For D-D″, scale bar in D″: 20 μm.