Fig. 5. Defects in visual axon projections and nervous system architecture in intact planarians after long-term Smed-netR RNAi treatment. (A-B) Merged confocal projections (VC-1 staining in green) and Nomarski (DIC) images (in gray), showing the visual axons of control (A) and dsRNA-injected (B) planarians. Arrowheads point to the posteriorly projecting visual axons. (C-J) Cephalic ganglia visualized by anti-phospho-tyrosine and Hoechst staining: (C-F) control animals; (G-J) Smed-netR dsRNA-injected animals. Single confocal sections were taken at the plane in which the ventral region of the brain overlaps with the dorsal portion of the VNCs (asterisks in C). (E,F) Higher magnification of C and D, respectively. The bracket in F indicates the juxtaposition of ventrolateral brain cells and the dorsolateral VNC cells. (G-J) The ventrolateral region of the brain (arrowheads in G and I) appears shifted laterally with respect to the VNCs (asterisks in G and I). (I,J) Higher magnifications of G and H, respectively. The bracket in J indicates the separation between ventrolateral brain cells (at the top of bracket) and the VNC cells (at the bottom of the bracket), yielding two discontinuous rows of nuclei. (K,M) Anti-phospho-tyrosine staining to visualize the submuscular nerve plexus from controls (K) and Smed-netR dsRNA-injected planarians (M). In K and M, arrowheads point to the plexus and the asterisks mark the position of the VNCs out of the focal plane. (L,N) Anti-tubulin staining to visualize the posterior VNCs of controls (L) and Smed-netR dsRNA-injected planarians (N). In N, arrowheads point to ectopic processes. All the samples shown were analyzed 4 weeks after RNAi treatment; C-J, L and N show single confocal planes; K and M show confocal projections through 2.5 µm. (A-B,K-N) Anterior to the top. (C-J) Anterior to the left. Scale bar: in B, 100 µm for A,B; in N, 100 µm for C,D,G,H,L,N and 50 µm for E,F,I,J,K,M.