Sonic hedgehog expression is locally inhibited by bmp4 induction and inhibition of Shh signaling results in defective RGC projections. (A–L) In situ hybridization in tg(hsp70l:bmp4) embryos and controls heat shocked at 15 hpf. (A,B) shha expression at 21 hpf in the diencephalon is inhibited by bmp4. (C–F) shha expression at 30 hpf in the anterior domain of the diencephalon (arrow) is inhibited by bmp4. (G,H) shhb expression at 30 hpf in the diencephalon (arrow) is inhibited by bmp4. (I,J) midline slit1a expression at 30 hpf is not changed by bmp4. (K,L) slit2 expression at 30 hpf is inhibited by bmp4. (M–P) RGC projection phenotypes in tg(pou4f3:mGFP) embryos treated with Cyclopamine. (M) Ethanol control. (N) Chart showing the distribution of RGC phenotypes in embryos treated with 100 µM Cyclopamine. Controls had exclusively contralateral projections (n = 61). (O) Ipsilateral RGC projections and (P) unilateral innervation of an optic tectum (“lambda”) in treated embryos. (Q–X) In situ hybridization in (R,T,V,X) embryos treated with 100 µM Cyclopamine and (Q,S,U,W) controls treated with ethanol. (Q,R) Expression of rgmd, (S,T) lrig1. (U,V) Expression of sema3d in the anterior diencephalon is sensitive to Cyclopamine treatment (arrow). (W,X) Expression of ephb2b. All images are maximum intensity projections, DAPI counterstaining, transverse view (except E,F, sagittal view). Scale bars 50 µm.
|
