Late subtype specification is altered and boundary cap cell identity is lost in mutant animals. (A) Illustration of a transverse section at E11.5 displaying cells, which contribute to the DRG in red. Green box represents caption area for subfigures B-G. (B-G) Triple immunohistochemistry at E11.5 for β-gal expressed by the R26R reporter and for markers for subtype specification of sensory neurons: tyrosine receptor kinases RET and TrkA, TrkB and TrkC. Dashed lines surround the border of R26R-positive cells contributing to the DRG. (H) Quantification of late sensory subtypes per DRG displays an altered distribution of sensory neuron subtypes in mutant animals. (I) Illustration of a transverse section at E12.5 displaying boundary cap cells in red. Green box represents caption area for subfigures J-O, respectively. (J-L) Double immunohistochemistry for Krox20 as a marker for differentiated boundary cap cells and neurofilament (NF) at E12.5. (M-O) Triple immunohistochemistry for Krox20, NF and Islet1/2 as a marker for motor neurons at E12.5. (J’,M’) In control animals, sensory axons invading the dorsal entry zone of the neural tube and motor axons penetrating the prospective motor exit point of the neural tube are surrounded by boundary cap cells. (K’,L’,N’,O’) Dorsal as well as ventral Krox20 expression is lost in both mutant animals. Arrows show migrating motor neurons, which are exiting or have already exited the ventral neural tube due to the loss of functional boundary cap cells. NT, neural tube; DEZ, dorsal entry zone; MEP, motor exit point; Scale bars: 25 μm. DRG, dorsal root ganglia; E; embryonic day.