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Fig. 3. Dlx3b function is required for generation of the full complement of otic sensory hair cells and semi-circular canals. (A,C,I) In normal (control) embryos, pax2a expression marks the presumptive otic placode at the three-somite (3s; A) and eight-somite (8s; C) stages. By high-pec stage (43h; I), otic sensory hair cells express Pax2a as indicated by antibody ({alpha}-Pax2; I) labeling. (B,D,J) In embryos injected with dlx3b-MO, pax2a expression is absent early (B; three-somite stage; 16/60 embryos) and reduced later (D; eight-somite stage; 5/17 embryos). By high-pec stage (J; 43 h) about half the normal number of Pax2-expressing otic sensory hair cells are apparent after knock down of dlx3b function by morpholino injection. Hair cell numbers as indicated by {alpha}-Pax2 labeling: wild-type 11.9±0.8 (n=34), dlx3b-MO 6.9±1.5 (n=49), dlx4b-MO 9.4±1.2 (n=7), dlx3b + dlx4b MOs 0.2±0.4 (n=18). (E,F) A Dlx3b antibody ({alpha}-Dlx3b) labels Dlx3b-expressing cells at the six-somite stage in normal embryos (E), but not after injection of dlx3b-MO (F). (G,H,K,L) Knock down of dlx3b function blocks formation of the otoliths (G,H; prim-5 stage, 24 h) and subsequent formation of the semi-circular canals (K,L; pec-fin stage). Twenty-eight percent of 356 injected embryos lacked both otoliths and semi-circular canals; all embryos injected with dlx3b-MO and dlx4b-MO lacked otoliths (data not shown). (A-F) Dorsal views, anterior towards the left; (G-L) side views, anterior towards the left, dorsal towards the top. Scale bar: 107 µm for A-D; 110 µm for E,F; 73 µm for G,H,K,L; 25 µm for I,J.