Netrin-1 is required for commissural axon guidance in the developing vertebrate nervous system.

During nervous system development, spinal commissural axons project toward floor plate cells and trochlear motor axons extend away from these cells. Netrin-1, a diffusible protein made by floor plate cells, can attract spinal commissural axons and repel trochlear axons in vitro, but its role in vivo is unknown. Netrin-1 deficient mice exhibit defects in spinal commissural axon projections that are consistent with netrin-1 guiding these axons. Defects in several forebrain commissures are also observed, suggesting additional guidance roles for netrin-1. Trochlear axon projections are largely normal, predicting the existence of additional cues for these axons, and evidence is provided for a distinct trochlear axon chemorepellent produced by floor plate cells. These results establish netrin-1 as a guidance cue that likely collaborates with other diffusible cues to guide axons in vivo.

The axonal chemoattractant netrin-1 is also a chemorepellent for trochlear motor axons.

Extending axons are guided in part by diffusible chemoattractants that lure them to their targets and by diffusible chemorepellents that keep them away from nontarget regions. Floor plate cells at the ventral midline of the neural tube express a diffusible chemoattractant, netrin-1, that attracts a group of ventrally directed axons. Here we report that floor plate cells also have a long-range repulsive effect on a set of axons, trochlear motor axons, that grow dorsally away from the floor plate in vivo. COS cells secreting recombinant netrin-1 mimic this effect, suggesting that netrin-1 is a bifunctional guidance cue that simultaneously attracts some axons to the floor plate while steering others away. This bifunctionality of netrin-1 in vertebrates mirrors the dual actions of UNC-6, a C. elegans homolog of netrin-1, which is involved in guiding both dorsal and ventral migrations in the nematode.

The role of the floor plate in axon guidance.

The data reviewed above have implicated the floor plate in directing axonal growth towards the midline, in directing the behavior of axons at the midline, and finally, in directing the longitudinal growth of axons alongside the midline. In the case of growth to the midline, there is clear evidence for the existence of two types of cues that collaborate to direct growth--short-range cues that can direct axons along the edge of the spinal cord and a long-range chemoattractant secreted by the floor plate cells whose main function may be to direct later-extending commissural axons that must migrate through the complex environment of the developing motor column. Determining the precise contribution of these cues will require identifying them and perturbing them in vivo. The cues that direct growth along the edge are unknown; their identification in the spinal cord would be of quite general significance, since the growth of axons parallel to (but not in contact with) the pial surface is a widespread feature of early axon growth at all axial levels of the neural tube. A strong candidate for the long-range chemoattractant is netrin-1, a homologue of the UNC-6 protein of C. elegans and a distant relative of laminin, which is expressed by floor plate cells and which can both promote and orient commissural axon outgrowth. Netrin-1 may also influence growth of other populations of neurons that exhibit stereotyped behaviors near the ventral midline. Much less is known about the exact role of the floor plate in directing axon growth at the midline, though it is clearly required for accurate guidance. In the absence of the floor plate, a range of errors has been found, the most prominent of which are aberrant midline crossings and errors in longitudinal growth near the ventral midline. The severity of these errors varies with species, which could result from either the variable importance of the floor plate in the different species or the fact that, so far, quite different manipulations of the ventral midline region have been performed in different species. The most specific perturbation of the ventral midline occurs in the zebrafish cyc-1 mutant, where the selective loss of the floor plate leads to stereotyped misrouting events. Perhaps surprisingly, virtually all axons that grow to the midline turn longitudinally (although sometimes in the wrong direction).(ABSTRACT TRUNCATED AT 400 WORDS)

7-Chlorokynurenate Blocks NMDA Receptor-Mediated Neurotoxicity in Murine Cortical Culture.

We examined the neuroprotective actions of the glycine site N-methyl-D-aspartate (NMDA) antagonist, 7-chlorokynurenate, in murine neocortical cell cultures. Cultures exposed for 5 min to 100 - 500 microM NMDA in the absence of added glycine developed substantial neuronal degeneration over the next 24 h. The addition of 10 microM glycine did not increase submaximal NMDA-induced neuronal injury, suggesting that endogenous glycine levels were sufficient to saturate its receptor sites on NMDA receptor complexes. Addition of 3 - 300 microM 7-chlorokynurenate produced concentration-dependent reduction in this neuronal damage with an IC50 of approximately 30 microM. Some injury reduction was seen even if the drug was added after completion of the NMDA exposure. The protective effect of 100 microM 7-chlorokynurenate could be overcome by adding 10 - 1000 microM glycine (glycine median effective concentration (EC50) approximately 100 microM) or 1 mM D-serine. As predicted by its ability to block NMDA receptor-mediated injury, 10 - 300 microM 7-chlorokynurenate also produced concentration-dependent reduction in the neuronal loss induced by 50 - 60 min exposure to combined glucose and oxygen deprivation. These data support the suggestion that pharmacologic interference with the binding of glycine to the NMDA receptor complex represents a potentially effective approach to blocking NMDA receptor-induced neurotoxicity in ischemia.