Surgical removal of the developing eye in the chick disrupts extraocular muscles (EOMs). AM indicates anterior mandibulae IO, inferior oblique IR, inferior rectus LR, lateral rectus MR, medial rectus PA, pharyngeal arches SO, superior oblique and SR, superior rectus.įigure 10. By 96 hpf, the sox10/p53 morphants had rudimentary EOM that were severely malformed and not organized around the eye compared with control MO (original magnification ×100). At 72 hpf, the sox10/p53 morphants showed significant delay in EOM development and overall decreased α -actin expression. Morpholino oligonucleotide (MO) against p53 was coinjected with the sox10 MO to decrease nonspecific (off-target) effects but knockdown of p53 did not alter the EOM phenotype. Knockdown of sox10 in the Tg(α -actin::EGFP) embryos demonstrates poor development of EOMs at 72 (A-C) and 96 hours after fertilization (hpf) (G-I) compared with controls (D-F, J-L). Microinjection of antisense oligonucleotide morpholinos at the 1- to 2-cell stage (within 30 minutes of fertilization) effectively knocked down the expression of sox10 and inhibited neural crest development. Neural crest cell migration is necessary for extraocular muscle (EOM) development. IO indicates inferior oblique IR, inferior rectus LR, lateral rectus MR, medial rectus P, pyramidalis SO, superior oblique SR, superior rectus and WT, wild-type.įigure 9. The dotted line in the lateral view corresponds to the coronal cut shown on the sections shown in Figure 5 (original magnification ×200).

Schematics of the rx3/chokh mutant (D-F) and wild-type (J-L) show the relation of the EOMs (red) to the eye (blue) and jaw musculature (purple).

In the Rx3/chokh mutants, muscles of the jaw and pharyngeal arch differentiate and form normally (purple arrows). However, the muscles are not properly organized into 6 EOMs, as seen in wild-type embryos (G-I). A-C, Whole-mount in situ hybridization for myhz2, the gene encoding myosin heavy chain, in Rx3 −/− mutants at 72 hours after fertilization (hpf) demonstrates the presence of differentiated muscle in the anophthalmic sockets (red arrows).

Genetic ablation of the optic vesicle in Rx3/chokh −/− mutant embryos results in poorly formed extraocular muscles (EOMs).

96 hour chick embryo cross section