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Fig. 1. Analysis of hypothalamic neurons in fezl/tof^m808 mutant embryos. (A-F) Heterozygous (WT; A,C,E) embryos and their too few^m808 (tof^m808) mutant siblings (B,D,F) were fixed at 52 (A,B), and 48 (C-F) hours post fertilization (hpf) and subjected to whole-mount in situ hybridization with antisense RNA probes directed against either the hypothalamic neuropeptide isotocin, which is the zebrafish ortholog of oxytocin (it; A,B), hypocretin/orexin (hcrt; C,D) or somatostatin (ss; E,F). After probe color development, all specimens were subjected to immunostaining with an anti-tyrosine hydroxylase (TH) antibody to detect DA neurons. WT heterozygous and tof^m808 embryos were scored by TH staining followed by sequencing-based genotyping. Black arrowheads indicate deficiencies in DA and IT neurons in too few embryos. (G,H) Schematic representations of the examined hypothalamic cell types and of fezl/tof expression domains in a 2-day-old WT embryo. All panels show lateral views of the embryo, anterior to the left. Hyp, hypothalamus; LC, locus coeruleus; NPO, neurosecretory preoptic area; PT, posterior tuberculum; Tel, telencephalon. Scale bars: 50 µm in A-D; 100 µm in E,F.

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