Figure 4

Progenitors marked with Shh- and Gli1-GIFM give rise to DA and RN neurons over several days of embryonic development. (A-K) Shh-GIFM; (L,M) Gli1-GIFM. (A-F) Immunofluorescent staining for DA neurons (TH, green) and β-gal-positive fate-mapped cells (red) on E18.5 coronal sections. Examples shown are located in the SN or the VTA. Arrows indicate double-labeled cells. Note that there are distinct contributions of marked cells to the SN or VTA at different fate-mapping time points. (G-I') Immunofluorescent staining for RN neurons (Pou4f1, green) and β-gal-positive fate-mapped cells (red) on E18.5 coronal sections. Arrows indicate double-labeled cells. Scale bar: 20 μm. (J-M) Quantification of the contribution of cells marked with Shh- or Gli1-GIFM to DA and RN neurons. Analysis was performed at E18.5. Cells positive for TH and β-gal (J,L) or Pou4f1 and β-gal (K,M) were counted and normalized for the number of counted sections. The peak contribution of cells marked with Shh-GIFM to DA neurons is with TM9.5, and to RN neurons with TM9.5 and TM10.5. These TM time points correlate with the broadest Shh-expressing domain. The peak contributions of cells marked with Gli1-GIFM to DA and RN neurons are one to two days earlier than observed for cells labeled with Shh-GIFM, consistent with medial Gli1 expression preceding Shh expression. Error bars indicate standard deviations; asterisks indicate the P-value as determined by Student's t-test (*P < 0.05; **P < 0.01). (N) Schematic of fate mapping strategy.