BMP/SMAD Pathway Promotes Neurogenesis of Midbrain Dopaminergic Neurons In Vivo and in Human Induced Pluripotent and Neural Stem Cells

The embryonic formation of midbrain dopaminergic (mDA) neurons in vivo provides critical guidelines for the in vitro differentiation of mDA neurons from stem cells, which are currently being developed for Parkinson's disease cell replacement therapy. Bone morphogenetic protein (BMP)/SMAD inhibition is routinely used during early steps of stem cell differentiation protocols, including for the generation of mDA neurons. However, the function of the BMP/SMAD pathway for in vivo specification of mammalian mDA neurons is virtually unknown. Here, we report that BMP5/7-deficient mice (Bmp5(-/-); Bmp7(-/-)) lack mDA neurons due to reduced neurogenesis in the mDA progenitor domain. As molecular mechanisms accounting for these alterations in Bmp5(-/-); Bmp7(-/-) mutants, we have identified expression changes of the BMP/SMAD target genes MSX1/2 (msh homeobox 1/2) and SHH (sonic hedgehog). Conditionally inactivating SMAD1 in neural stem cells of mice in vivo (Smad1(Nes)) hampered the differentiation of progenitor cells into mDA neurons by preventing cell cycle exit, especially of TH(+)SOX6(+) (tyrosine hydroxylase, SRY-box 6) and TH(+)GIRK2(+) (potassium voltage-gated channel subfamily-J member-6) substantia nigra neurons. BMP5/7 robustly increased the in vitro differentiation of human induced pluripotent stem cells and induced neural stem cells to mDA neurons by up to threefold. In conclusion, we have identified BMP/SMAD signaling as a novel critical pathway orchestrating essential steps of mammalian mDA neurogenesis in vivo that balances progenitor proliferation and differentiation. Moreover, we demonstrate the potential of BMPs to improve the generation of stem-cell-derived mDA neurons in vitro, highlighting the importance of sequential BMP/SMAD inhibition and activation in this process.