Re-assessing the role of the floor plate as an intermediate target for commissural axons in the embryonic mouse spinal cord

Abstract

In the developing nervous system, axons navigate great distances to reach their final targets. Along their path, they encounter intermediate targets, which contain cues to guide them along the next leg of their journey. Spinal commissural axons comprise a population of axons that cross the floor plate (FP), an intermediate target at the ventral midline (VM) of the embryonic spinal cord. Here, we describe contralateral commissural projections and examine the role of FP contact and B-class Eph/ephrins on commissural axon pathfinding.;We describe four distinct contralateral commissural projections. Intermediate longitudinal commissural (ILC) axons travel diagonally away from the FP into intermediate regions of the spinal cord, while (MLC) axons grow alongside the FP. Bifurcating longitudinal commissural (BLC) axons split into rostral and caudal projections. Forked transverse commissural (FTC) extend transverse to the FP.;In addition, we assess the role of FP contact on contralateral commissural pathfinding. FP contact has been shown to be important in altering the responsiveness of commissural axons to midline-associated cues (e.g. Slits) so they can exit the FP and turn longitudinally. Here, we demonstrate that numerous decussated axons elaborate wildtype-like projections without contacting FP cells in vivo and in vitro. Interestingly, slit1-3 mRNA is expressed in the ventral ventricular zone and their receptors (Robo1/2) are expressed on longitudinal axons in FP-lacking mice. Supporting an active role for Robo-Slit signaling, blocking Robo-Slit interactions in FP-lacking explants prevents commissural axons from decussating and turning longitudinally. Together, these findings show that Robo-Slit interactions, but not FP contact, are required for the proper elaboration of many contralateral commissural projections.;Finally, we examine the in vivo role of B-class Eph/ephrins on commissural axon guidance. EphrinB3 is enriched in FP cells, decussated commissural axons express EphB receptors, and B-class ephrins are expressed in the dorsal spinal cord. EphB/ephrin mutants elaborate wild-type-like contralateral commissural projections, but ephrinB3/EphB mutants were observed to exhibit a significant number of commissural axons exhibiting midline guidance errors, including an increased frequency of FTC projections, as compared to wild-type littermates. Interestingly, these errors were not observed in ephrinB3 lacz mice, which are incapable of reverse signaling.