Fig. 5.

Müllerian duct regression in mutant urogenital ridges is inhibited in organ culture. Urogenital ridges from E13.5 control and mutant female fetuses were isolated, incubated for 48 h with 5 μg/mL MIS, and subjected to whole mount in situ hybridization with Wnt7a riboprobe detected by BM purple. (A) There is no Wnt7a mRNA expression in control female urogenital ridge with MIS, indicating complete regression of MD epithelium. (Inset) Wnt7a expression in a control female urogenital without added MIS (indicated with a white arrowhead). (B) Complete MD retention (indicated with a white arrowhead) with an Amhr2-Cre/+;Ctnnb1^Δ(ex3)/+ female urogenital ridge similarly incubated with MIS. In situ hybridization for Amhr2 mRNA in control (C) and Amhr2-Cre/+;Ctnnb1^Δ(ex3)/+ (D) E16.5 male reproductive tract shows expression in the retained MD tissue in the mutant (indicated with black arrowheads in the epididymis and a black arrow in the vas deferens). pSmad1/5/8 expression was detected by immunofluorescence in control (E) and mutant (F) E14.5 male urogenital ridges. DAPI staining in blue indicates nuclei. O, ovary; T, testis; E, epididymis; MDM, Müllerian duct mesenchyme; MDE, Müllerian duct epithelium; WD, Wolffian duct. (Scale bars, 50 μm.)