Zebrafish NECs are not neural crest-derived: analysis of neural crest-deficient zebrafish mutants.
(a) 7-dpf tfap2amob;foxd3mos zebrafish lack all neural crest derivatives, including melanophores and jaw skeleton (arrowhead). Dotted lines: section planes in b-e. (b,c) At 7-dpf, tfap2amob;foxd3mos orobranchial epithelium retains innervated serotonergic (5-HT+) cells (putative NECs), identified by cytoplasmic serotonin surrounded by a ring of HNK1 epitope-immunoreactive neurites. (d,e) At 7-dpf, putative NECs (arrowheads) persist in tfap2amob;foxd3mos zebrafish in the region ventral to the orobranchial cavity where the pharyngeal arches would be located in wild-type fish. (NB The hypothalamus [Hyp] extends caudally beneath the midbrain and rostral hindbrain, and often separates from the overlying brain on sections, as here.) (f) The mean number per 7-dpf larva of putative NECs in the orobranchial epithelium does not differ between tfap2amob;foxd3mos (98.0 ± 49.7 s.d.; n = 3) and wild-type zebrafish (81.3 ± 24.0 s.d.; n = 3) (p=0.63, unpaired two-tailed Student’s t-test). All such cells in the orobranchial epithelium were counted for each embryo. Error bars indicate s.d. (g–i) Wild-type sibling at 7-dpf. Dotted line: section plane in h-i. Putative NECs are present in the orobranchial epithelium (arrowheads; dashed box in i, magnified without DAPI in top right corner). 5-HT, serotonin; Gf, gill filament; Hyp, hypothalamus; Nt, neural tube; Obc, orobranchial cavity. Scale-bars: 50 μm in b,d,h; 25 μm in c,e,i.