Fig. 6

Formation of dorso-laterally located commissural nuclei is disrupted in OM2 morphants (number of samples analyzed are the same as in Fig. 5). Later-onset, ascending commissural nuclei within the hindbrain were visualized using Zn-5 immunostaining. Islet-1 positive primary motor neurons are detected by GFP (green). (A, C and E) Control embryos. (B, D and F) OM2 morphants. (A and B) Embryos (28 hpf), showing that pharyngeal arch endodermal pouch (PEP) formation is moderately defective in OM2 morphants (arrows). Dotted line in (B) denotes partially divided endodermal pouches in OM2 morphants. M, mandibular arch; H, hyoid arch. (C and D) Embryos (36 hpf), showing intricate structures of Zn-5 positive cells. Hindbrain commissural neurons (brackets) are almost completely lacking or severely underdeveloped in OM2 morphants, although commissural neurons adjacent to the CN VI do form partially in morphants (dotted circle). PEP formation in OM2 morphants also appears abnormal, with pharyngeal endodermal pouches 2 and 3 possibly fused. g1, first gill endodermal pouch. (E and F) In 54 hpf embryos continued absence of development of hindbrain commissural neurons is observed in OM2 morphants. Gill motor axons from cranial nerve X send their axons through Zn-5 negative areas (inset in E), which is sharply contrasted by morphant embryos where gill motor axons fail to grow through the PEPs (inset in F). All photographs were taken with 200x magnification and z-stacked images were flattened to show structures located at different depths. The scale bar in (F) indicates 100 micrometer.