Integration of opposing semaphorin guidance cues in cortical axons.

Previous work demonstrated that members of the semaphorin family, Sema3A and Sema3C, act as repulsive and attractive guidance signals, respectively, for cortical axons. During the development of corticofugal projections, these semaphorins are expressed in adjacent cortical zones, but there is a considerable overlap between Sema3A and Sema3C expression in the subventricular zone. We used different in vitro assays to examine the response of cortical axons exposed to defined mixtures of these opposing guidance cues. Results showed that even at very low concentrations, Sema3A overrides the effects of Sema3C. Moreover, experiments with function-blocking antibodies directed against neuropilin provided insights into how cortical axons integrate disparate guidance signals at the receptor level. These in vitro data suggest that the pathway of corticofugal axons is defined by an attractive cue in the intermediate zone, where Sema3C is expressed alone. To directly test this hypothesis in vivo, we performed axon-tracing experiments in Sema3C-deficient mice. Compared with wild-type animals, corticofugal axons take a more superficial route in Sema3C(-/-) mice, and the corticofugal pathway is more compacted. This phenotype is expected when an attractive cue for cortical axons, Sema3C, is eliminated and a repulsive cue, Sema3A, becomes predominant.

Ephrins guide migrating cortical interneurons in the basal telencephalon.

Cortical interneurons are born in the proliferative zones of the ganglionic eminences in the subpallium and migrate to the developing cortex along well-defined tangential routes. The mechanisms regulating interneuron migration are not completely understood. Here we examine the role of class-A members of the Eph/ephrin system in directing the migration of interneurons. In situ hybridizations demonstrated that ephrin-A3 is expressed in the developing striatum, an area that is strictly avoided by migrating cortical interneurons in vivo, which express the EphA4 receptor. We then examined interneuron migration in grafting experiments, where explants of the medial ganglionic eminence (MGE) from enhanced green fluorescent protein-expressing transgenic mice were homotopically grafted into host slices from wildtype littermate embryos. After blocking ephrin-A ligands, many interneurons invaded the striatal anlage. Moreover, stripe assay experiments revealed that ephrin-A3 acts as a repellent cue for neurons from the medial ganglionic eminence. Downregulation of the EphA4 receptor via siRNA transfection reduced the repulsive effect of ephrin-A3, indicating that EphA4 mediates at least in part the repulsive effect of ephrin-A3 on these cells. Together, these results suggest that ephrin-A3 acts as a repulsive cue that restricts cortical interneurons from entering inappropriate regions and thus contributes to define the migratory route of cortical interneurons.