Fig. 6

Small-Molecule-Mediated Inhibition of the Gdf3 Receptor Alk4 Phenocopies the Pharyngeal Defects Observed in tbx1 Mutants

(A) Experimental timeline of SB-505124 treatment and phenotypic analyses.

(B–D) Bright-field z-stacks of 38 hr post-fertilization (hpf) wild-type embryos treated with DMSO or SB-505124 between 75% epiboly and 24 hpf, and processed by in situ hybridization with a tie1 riboprobe. Images of representative animals with unaffected (B; 3 or more clusters), reduced (C; 1–2 clusters), or absent (D; 0 clusters) tie1⁺ pharyngeal arch artery (PAA) clusters are shown. The numbers in (B) and (C) identify each PAA cluster. The asterisks highlight clusters of smaller size in (C). Lateral views, anterior left.

(E) Bar graph showing the percentages of embryos with unaffected, reduced, or absent PAA clusters after treatment with DMSO (n = 22), 40 μM SB-505124 (n = 27), or 80 μM SB-505124 (n = 12).

(F–K) Confocal z-stacks of 72 hpf wild-type embryos treated between 75% epiboly and 24 hpf with DMSO or SB-505124 and co-immunostained with antibodies recognizing striated muscle (MF20, red) or outflow flow tract (OFT) smooth muscle (Eln2, green). Images of representative animals with unaffected (F and I), reduced (G and J), or absent (H and K) head muscles and OFTs (white arrows, F and G) are shown. (F–H) Ventral views, anterior up. (I–K) Lateral views, anteior left.

(L and M) Bar graphs showing the percentages of animals with unaffected, reduced, or absent head muscles (L) or OFTs (M) after treatment with DMSO (n = 49), 20 μM SB-505124 (n = 40), or 40 μM SB-505124 (n = 53).

Scale bars, 25 μm. PA1-derived head muscles: am, abductor mandibulae; do, dilator operculi; ima, intermandibularis anterior; imp, intermandibularis posterior; lap, levator arcus palatini; PA2-derived head muscles: ah, adductor hyoideus; ao, adductor operculi; hh, hyohyoidus; ih, interhyoidus; lo, levator operculi. LDA, lateral dorsal aorta. OFT, outflow tract; V, ventricle.