Fig. 5

arx is required for the formation of DA neurons in the prethalamus and in the preoptic area. Morpholino-injected embryos were analyzed by WISH to detect th (A-F, 96 hpf), slc6a3/dat (M-P, 72 hpf) or ddc (Q-T, 72 hpf) expression, as well as by anti-TH immunofluorescence (G-L, 72 hpf). Arx knockdown by Morpholino injection results in a reduction in the number of prethalamic group 1 DA neurons (arrows in B, E, H) and of the preoptic group (K, arrowhead points to the postoptic commissure). This phenotype is not caused by Morpholino off-target p53-mediated cell death, as we obtain the same result when co-injecting p53-MO as a control (C, F, I, L). Moreover, the prethalamic expression domains of slc6a3/dat (M–P) and ddc (Q–T) are also reduced in morphant embryos when compared to controls (arrows point to the prethalamus). U: Evaluation of the effect of Arx knockdown on th expression for each DA cluster. Both ATG (left) and splice (right) targeting morpholinos affected DC1 and PO groups development, also when co-injected with the p53-MO (+). The groups of the arx morphant larvae were compared with those of control MO injected larvae, which all developed normally and are summarized here in the left-most lanes (ctr). A–C,M,N,Q,R are lateral views; D–L, O, P, S, T are dorsal views; anterior is to the left. Scale bars: 100 μm. Abbreviations: AC, amacrine cells; DC1-6, diencephalic groups 1–6; LC, locus coeruleus; MO, medulla oblongata; OB, olfactory bulb; PO, preoptic area; poc, postoptic commissure; Pr, pretectum; SP, subpallium; TC, telencephalic cluster; vDC, ventral diencephalic cluster.