Using Nogo antibodies with defined binding specificity, Nogo-B, but not Nogo-A, was localized on radial glia in the floor plate of mouse embryos. The presence of Nogo-B was confirmed in Nogo-A knockout mice. In explant cultures of embryonic day (E) 11 and E12 spinal cord, blocking of NgR function with antagonist peptide NEP1-40 reduced the crossing of newly arrived commissural axons, resulting in an accumulation of growth cones in the floor plate. Analysis of growth cone morphology demonstrated an increase in size of growth cones in the floor plate after peptide treatment, which was not detected in axons growing toward the midline. In knockout embryos, midline crossing was not affected by absence of Nogo-A. In co-culture experiments using collagen gel, floor plate showed a strong inhibitory effect on the extension of post-commissural neurites from the spinal cord. This effect was abolished by NEP1-40, and was observed neither in pre-commissural neurites, nor in post-commissural neurites grown with floor plate derived from Nogo-A knockout embryo. Furthermore, western blot analysis of conditioned medium from floor plates showed a truncated form of Nogo with molecular weight of 37 kDa, which could mediate the diffusible effect to axon growth. We conclude that Nogo-B is expressed in the floor plate of mouse embryo, which probably mediates axon crossing in the spinal cord by repelling axons out of the midline when they start upregulate NgR. Nogo acts on axon growth not only through a contact-mediated mechanism, but also through a diffusible mechanism.
J Comp Neurol
2915 - 2928
Nogo receptor, RRID: AB_10000211, RRID: AB_138404, RRID: AB_141357, RRID: AB_143165, RRID: AB_144696, RRID: AB_213621, RRID: AB_2183731, RRID: AB_2313567, RRID: AB_2313773, RRID: AB_2534102, RRID: AB_531775, axon guidance, floor plate, spinal cord, Age Factors, Animals, Axon Guidance, Axons, Coculture Techniques, Contactin 2, Culture Media, Conditioned, Embryo, Mammalian, Embryoid Bodies, Female, Gene Expression Regulation, Developmental, Growth Cones, In Vitro Techniques, Male, Mice, Mice, Inbred C57BL, Mice, Knockout, Netrin-1, Nogo Proteins, Nogo Receptors, Organ Culture Techniques, Pregnancy, Spinal Cord, Tubulin