WNT ligand involvement in MbDA neuron development in vivo. Genetic inducible fate mapping (GIFM) shows that distinct spatial and temporal epochs underpin Wnt1 lineage contribution to MbDA

INTRODUCTION The midbrain (Mb) comprises the superior colliculus, which integrates attention and visual stimuli, and the inferior colliculus, which is an obligate auditory processing center. These dorsal Mb (d.Mb) structures are adjacent to the cerebellum (Cb), which controls motor behaviors. The ventral Mb (v.Mb) contains molecularly, physiologically and biochemically distinct neurons that control complex functions. Of particular interest are the Mb dopamine (MbDA) neurons because they modulate brain function and are centrally involved in schizophrenia, addiction and Parkinson’s disease. During embryogenesis, MbDA neurons are derived from multiple lineages originating within the mesencephalon (mes), which is the Mb primordium (Zervas et al., 2004; Joksimovic et al., 2009; Brown et al., 2011; Hayes et al., 2011; Blaess et al., 2011). The molecular identity of the mes is based on a combinatorial code of transcription factors and is patterned by signaling molecules, including WNT1 (reviewed by Zervas et al., 2005). We investigated Wnt1, which is initially expressed broadly in the mes, but becomes restricted to distinct domains over time (Wilkinson et al., 1987; Echelard et al., 1994; Zervas et al., 2004). Mice with a null allele of Wnt1 or with a spontaneous mutation in Wnt1 do not form the Mb and Cb, demonstrating the importance of WNT1 for these domains (McMahon and Bradley, 1990; McMahon et al., 1992; Thomas et al., 1991; Bally-Cuif et al., 1995; Ellisor et al., 2012). WNT1 functions through β-catenin-mediated WNT signaling and β-catenin regulates cell cycle exit and MbDA neuron development (Tang et al., 2009; Tang et al., 2010). In addition, βcatenin binds to the promoter of Lmx1a, which is proposed to be a determinant of MbDA neurons (Andersson et al., 2006; Chung et al., 2009). However, all WNTs that use canonical signaling converge on β-catenin, which precludes an understanding of the specificity of WNT ligand involvement in MbDA neuron development in vivo. Genetic inducible fate mapping (GIFM) shows that distinct spatial and temporal epochs underpin Wnt1 lineage contribution to MbDA neurons and to Cb neurons (Brown et al., 2011; Hagan and Zervas, 2012). Furthermore, dynamic oscillatory genetic networks in human neuronal progenitors are influenced by WNT1 (Wexler et al., 2011). However, the functional requirement of WNT1 in developmental processes is unresolved. Thus, we generated a conditional knockout allele of Wnt1 that we coupled with GIFM to uncover distinct spatial and temporal requirements for Wnt1 in patterning the Mb and Cb, controlling ventral mesencephalon (v.Mes) progenitors and regulating cell cycle exit during MbDA neuron development.