Larger Image Fig. 5. The inhibition of Shh, Fgf and Wnt-8 signaling affects eye, brain, and placode development in tadpole stage embryos. (A) Overexpression of mHip1 and Shh. (a–g′) Microinjection of mHip1 (750 pg) into one cell of two-cell stage embryos. (h–n′) Overexpression of Shh (500 pg) into one cell of two-cell stage embryos. (b, e′, f, l′, m) Frontal view. (a, c, d, e, h, i, j, k, l) Lateral view. (g, n) Dorsal view. (a′–d′, f′, g′, h′–k′, m′, n′) Transversal sections. (a, a′) Formation of a giant eye. Note Xrx1-positive cells extend into the prospective midbrain region (white arrow in a′, 67%, n = 18). (b, b′) Repression of optic stalk marker Vax1 within the ventral fore- and midbrain (white arrows, 63%, n = 28). (c, c′) Displaced Rhodopsin positive cells in the enlarged eye (76%, n = 21). (d, d′) Abnormal lens formation revealed by Crystallin α expression (50%, n = 12). (e, e′) Enlarged olfactory placode as revealed by Emx2 expression, indicated by dashed open rectangles (65%, n = 17). (f, f′) Ventral shift of the dorsal limit of Nkx-2.1 expression (white arrows in f′, 55%, n = 11). (g, g′) Ventral expansion of Gsh-1 expression (38%, n = 8). (h–h′) Note the reduction of retinal tissue (Xrx1, dark brown) and lens tissue (Pitx3, red, 50%, n = 10). (i, i′) Expanded expression domain of the optic stalk marker Vax1 (white arrow in i′, 62%, n = 13). (j, j′) Loss of Rhodopsin positive cells (83%, n = 16). (k, k′) Absence of lens specific Crystallin α expression (57%, n = 14). (l, l′) Suppression of olfactory placode development as revealed by the loss of Emx2 expression, indicated by a red arrow (65%, n = 17). The dashed open rectangle marks the olfactory placode on the non-injected side (l′). (m, m′) Dorsal shift of ventral forebrain tissue as revealed by Nkx-2.1 expression (56%, n = 16), the eye structures are emphasized by dashed ellipses. (n, n′) Reduced expression of the dorsal neural tube marker Gsh-1 in the prospective midbrain (black arrow, 42%, n = 12). LacZ mRNA was co-injected as a lineage tracer (light blue in b′, c, c′, e, e′, f′, g, g′, l, l′, n′). (B) Influence of dnWnt-8, XFD and Fgf-8 overexpression on lens and olfactory placode development. (a′, b′, c′, d′, e′, f′) Frontal views, dashed open rectangles indicate the size of the olfactory and lens placodes, respectively. (a, a′, b, b′, c, c′, d, d′, e, e′, f, f′) Lateral views. (a–b′) Overexpression of dnWnt-8 (500 pg) resulted in enlarged olfactory (Emx2; 76%, n = 22) and lens placodes (Pitx3; 78%, n = 18). (c–d′) XFD (1 ng) injection did not alter the size of the olfactory placode but led to enlarged lens placodes (83%, n = 16). (e–f′) Additional Fgf-8 (10 pg) inhibits olfactory and lens placode induction (red arrows in e′ and f′; 76%, n = 19). (a–a′, c–c′, e–e′) Krox-20 and En-2 expression was followed to control hindbrain and midbrain–hindbrain formation, respectively. LacZ mRNA was co-injected as a lineage tracer (light blue in a′, a′, b′, b′, c′, c′, d′, d′, e′, e′, f′, f′). Abbreviations: ey, eye cup; mhb, midbrain–hindbrain boundary; olp, olfactory placode; r3, rhombomere 3; r5, rhombomere 5; tel, telencephalon. Image published in: Cornesse Y et al. (2005) Copyright © 2005. Image reproduced with permission of the Publisher, Elsevier B. V. Permanent Image Page Printer Friendly View XB-IMG-43415