Axon guidance at the midline: of mice and flies

Abstract

In bilaterally symmetric organisms, the midline is a critical organizing center for the developing central nervous system. There is a striking conservation of the molecules and mechanisms that control axon path finding at the midline in vertebrate and invertebrate nervous systems. The majority of axons in the CNS cross the midline before projecting to their contralateral synaptic targets and this crossing decision is under exquisite spatial and temporal regulation. Growing commissural axons initially respond to attractive signals, while inhibiting responses to repulsive signals. Once across, repulsion dominates, allowing axons to leave and preventing them from re-entering the midline. Here we review recent advances in flies and mice that illuminate the molecular mechanisms underlying the establishment of precise connectivity at the midline.

Figure 1

Ligands and receptors that mediate midline attraction and repulsion in fly and mouse. Diagrams show schematics of the Drosophila embryonic ventral nerve cord (left) and an open-book preparation of a mouse spinal cord (right). For both panels, anterior is up; the mediallateral axis of the fly CNS corresponds to the dorsal-ventral axis of the mouse spinal cord. In flies, Frazzled (Fra) and Dscam are the only known attractive midline receptors. Fra responds to a midline source of Netrin, while the attractive ligand for Dscam is unknown. Roundabout (Robo) receptors mediate midine repulsion in response to Slit in ipsilateral and post-crossing contralateral axons. Vertebrate homologs of Fra (DCC), Dscam, and Robo mediate similar guidance responses; however, in mice Dscam is an attractive Netrin receptor. In vertebrates, additional ligand-receptor systems influence midline crossing, including the Sonic hedgehog (Shh) morphogen, which promotes midline attraction via the Boc receptor, and the Ephrin-B3 (a transmembrane ephrin) and Sema3B (a secreted semaphorin) repellants, which signal through the EphA4 and Neuropilin-2 (Npn-2) receptors, respectively. For simplicity, schematics indicate guidance decisions of commissural axons and do not depict other components of Shh and Sema receptor complexes, but note that some repulsive ligand-receptor systems (for example Slit-Robo in fly and vertebrate, and EphrinB3-EphA4 in vertebrate) have been implicated in repelling both ipsilateral and post-crossing contralateral axons. FP, floor plate.

Figure 2

Pre-crossing and post-crossing regulation of attractive and repulsive guidance Top, schematics showing commissural axons before, during, and after crossing the CNS midline (grey). Bottom, enlarged schematics of pre- and post-crossing neurons indicating regulation and signaling output of axon guidance receptors. In pre-crossing fly commissural axons, Frazzled (Fra) mediates attractive signaling in response to midline-produced Netrin (Net). Fra also signals independently of Net to induce expression of commissureless (comm), a transmembrane sorting receptor that directs Robo-containing vesicles to the late endocytic pathway. After crossing, comm expression ceases and Robo is trafficked to the growth cone where it can mediate a Slit-dependent repulsive response and prevent re-crossing. Robo and Fra independently regulate crossing in fly axons. In vertebrates, the Fra homolog DCC signals Net-dependent attraction as axons approach the CNS ventral midline. Pre-crossing Slit sensitivity is antagonized by the Robo3.1 isoform of the Robo3/Rig-1 receptor. After crossing, Robo3.1 is replaced by Robo3.2, which functions alongside Robo1 (and Robo2, not shown) to prevent re-crossing. By analogy to Xenopus spinal axons, Robo1 could potentially act in post-crossing vertebrate commissural axons to silence the attractive output of DCC.