Glial Cell–Derived Neurotrophic Factor–Independent Ureteric Bud Outgrowth from the Wolffian Duct Akito Maeshima, Hiroyuki Sakurai, Yohan Choi, Shinji Kitamura, Duke A. Vaughn, James B. Tee, and Sanjay K. Nigam Departments of Pediatrics, Medicine, and Cellular & Molecular Medicine, University of California, San Diego, La Jolla, California ABSTRACT The kidney collecting duct system and the ureter derive from the ureteric bud, an outgrowth of the Wolffian duct. It is generally believed that glial cell– derived neurotrophic factor (GDNF) plays a critical role in this earliest stage of kidney development, but 30 to 50% of knockout mice that lack either Gdnf or one of its receptors, such as Ret, have normal ureters. This suggests that an alternative pathway can induce ureteric bud outgrowth from the Wolffian duct. Isolated Wolffian ducts were cultured, and it was found that a combination of fibroblast growth factor 7 (FGF7) and blockade of the TGF- superfamily member activin A induced formation of buds from the Wolffian duct. This occurred even in the presence of a neutralizing anti-GDNF antibody or in Ret-knockout– derived Wolffian ducts, suggesting GDNF- independent induction of bud formation. Similar to wild-type ureteric buds or those induced by GDNF, FGF7/follistatin-induced buds were shown to be functionally competent, as they underwent branching morphogenesis and induced nephron formation upon recombination with metanephric mesenchyme. These in vitro findings suggest that modulation by FGF7 and the activin A signaling pathway, or equivalent pathways, can lead to GDNF-independent induction of ureteric bud outgrowth, possibly explaining the seemingly normal ureteric bud outgrowth in Gdnf or Ret null mice. J Am Soc Nephrol 18: 3147–3155, 2007. doi: 10.1681/ASN.2007060642 The embryonic kidney forms through reciprocal interac- tions between the metanephric mesenchyme (MM) and the ureteric bud (UB). 1 It is thought that the UB emerges from the Wolffian duct (WD) in response to glial cell– derived neurotrophic factor (GDNF) secreted by the MM. Gdnf knockout mice are known to have renal agen- esis, 2– 4 and mice lacking GDNF receptor genes such as Gfra1 5 and Ret 6,7 also exhibit similar phenotypic abnor- malities. In vitro studies have also demonstrated that GDNF signaling through GFR1 and Ret is a central sig- naling system involved in both the UB outgrowth from the WD 8,9 and branching morphogenesis of the UB. 10 Nevertheless, a substantial percentage of knockout ani- mals lacking Gdnf (27% 2 ) or its receptors (40 to 45% of Ret -/- 7 ) have seemingly normal UB formation, suggest- ing that the GDNF-dependent budding pathway of the WD may be bypassed by activation of other signaling pathways. The biological roles of fibroblast growth factor (FGF) have been extensively investigated by target- ing the genes of individual Fgf or FGF receptors (Fgfr) by homologous recombination or by overex- pression of soluble receptors. 11 In some of these mice, embryonic lethality has precluded identifica- tion of a renal phenotype. In other cases, knockouts of FGFs display mild abnormalities late in kidney development that do not seem to result from ab- normal ureteric budding. 12,13 Overexpression of Received June 4, 2007. Accepted October 9, 2007. Published online ahead of print. Publication date available at www.jasn.org. A.M. and H.S. contributed equally to this work. Correspondence: Dr. Sanjay K. Nigam, Department of Pediatrics, Med- icine, Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0693. Phone: 858-822- 3482; Fax: 858-822-3483; E-mail: snigam@ucsd.edu Copyright © 2007 by the American Society of Nephrology BASIC RESEARCH www.jasn.org J Am Soc Nephrol 18: 3147–3155, 2007 ISSN : 1046-6673/18112-3147 3147