Fig. 3. Apical neurogenesis is altered in RNAi cnox-2 knocked-down hydra. (A) RT-PCR assay showing cnox-2 expression levels in whole polyps (upper panel) or upper halves collected immediately after bisection (lower panel), exposed 5x and 9x to control (lanes 1,3,5,7) or cnox-2 (lanes 2,4,5,8) dsRNAs. (B-E) cnox-2 expression pattern in hydra exposed to Kazal1 (B) and cnox-2 (C-E) dsRNAs. Note the reduced size of hydra in C. (F,G) Number of apical (F) and gastric (G) cnox-2⁺ cells per hydra exposed once or 5x to control (lanes 1,4), Kazal1 (lanes 2,5), cnox-2 (lanes 3,6) dsRNAs. (H-N) Disorganization of the ANS upon cnox-2 silencing detected by anti-ß-tubulin staining (see Fig. S2 and Movie S1 in the supplementary material). (H,I,K,M) In control and Kazal1(-) hydra, the ANS formed of parallel sensory neurons (arrows) and meridien multipolar neurons (arrowheads) extends from the tentacle zone up to the mouth opening. (J,L) Reduced and disorganized ANS after 5x and 7x cnox-2 dsRNAs exposures. (M,N) Apical neurons are no longer detected after 9x cnox-2 dsRNA exposures. H,J,K,L: maximum projection confocal views; I,M,N: axioplan views. Scale bars: 400 µm in B,C; 200 µm in D,E; 50 µm in H-N. mo, mouth opening; tent, tentacules; tz, tentacle zone.