Navigation of trochlear motor axons along the midbrain-hindbrain boundary by neuropilin 2

Trochlear motor axons project dorsally along the midbrain-hindbrain boundary (MHB) to decussate at the dorsal midline. We report on the roles of neuropilin 2 and its ligands in the molecular mechanisms controlling this trajectory. In chick embryos, neuropilin 2 was expressed in the neuroepithelium of the dorsal isthmus in addition to the trochlear neurons, and Sema3F transcripts were localized along the caudal margin of the midbrain. Misexpression of Sema3F demonstrated that Sema3F displays repulsive activity in vivo that guides the trochlear motor axons along the MHB. An unexpected result was that misexpression of neuropilin 2 canceled the midbrain-evoked repulsion, allowing trochlear motor axons to cross the MHB and invade the tectum. A binding assay with neuropilin 2 ectodomain revealed the existence of neuropilin 2 ligands in the midbrain, which were masked by ectopic neuropilin 2. We therefore propose that neuropilin 2 neutralizes the repulsive activity in order to steer trochlear motor axons towards the dorsal decussation point. Taken together, our results suggest that the interaction of neuropilin 2 with its ligands has crucial roles for establishing trochlear trajectory along the MHB.