NeuN is expressed at low levels or not at all in postnatal Gfrα1-TLZ+/Hb9::GFP- motor neurons. (A) Confocal image of a P5 ventral horn showing Gfrα1-TLZ expression (Cy3, red), Hb9::GFP (green) and NeuN immunoreactive neurons (Cy5, white). All confocal planes through the 70 μm thick section and all fluorescent signals are superimposed. The lamina IX region enclosed by the dotted yellow box is shown in (A1-3) with Gfrα1-TLZ expression superimposed on Hb9::GFP (A1), Gfrα1-TLZ expression on NeuN (A2) and Hb9::GFP on NeuN (A3). (B,C) Similar image series for P10 (B,B1-3) and P20 (C,C1-3). At all ages, NeuN immunoreactivity is very low or not present at all in small MNs that are Gfrα1-TLZ+ and Hb9::GFP- (white arrows). In contrast, large Hb9::GFP+ MNs (yellow arrows) almost always express high levels of NeuN immunoreactivity. Scale bars: (A,B,C) 100 μm.