Repulsive guidance of axons of spinal sensory neurons in Xenopus laevis embryos: roles of Contactin and notochord-derived chondroitin sulfate proteoglycans.

An immunoglobulin superfamily neuronal adhesion molecule, Contactin, has been implicated in axon guidance of spinal sensory neurons in Xenopus embryos. To identify the guidance signaling molecules that Contactin recognizes in tailbud embryos, an in situ binding assay was performed using recombinant Contactin-alkaline phosphatase fusion protein (Contactin-AP) as a probe. In the assay of whole-mount or sectioned embryos, Contactin-AP specifically bound to the notochord and its proximal regions. This binding was completely blocked by either digestion of embryo sections with chondroitinase ABC or pretreatment of Contactin-AP with chondroitin sulfate A. When the spinal cord and the notochord explants were co-cultured in collagen gel, growing Contactin-positive spinal axons were repelled by notochord-derived repulsive activity. This repulsive activity was abolished by the addition of either a monoclonal anti-Contactin antibody, chondroitin sulfate A or chondroitinase ABC to the culture medium. An antibody that recognizes chondroitin sulfate A and C labeled immunohistochemically the notochord in embryo sections and the collagen gel matrix around the cultured notochord explant. Addition of chondroitinase ABC into the culture eliminated the immunoreactivity in the gel matrix. These results suggest that the notochord-derived chondroitin sulfate proteoglycan acts as a repulsive signaling molecule that is recognized by Contactin on spinal sensory axons.