Network formation of motoneurons

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We previously found that two of the Eph receptor tyrosine kinases, EphA3 and EphA4, are transiently expressed in subsets of chick embryonic motoneurons and that the Eph ligands, ephrin-A2 and ephrin-A5, inhibit neurite outgrowth from the motoneurons in culture. Here we developed functional blocking monoclonal antibodies against EphA3, EphA4 and ephrin-A2 in order to examine their functional roles in axonal guidance of motoneurons in vivo. We raised many monoclonal antibodies against the chimeric proteins of each extracellular domain and Fc region of human IgG.Among them some antibodies had blocking activity for ligand-receptor binding and functional blocking activities in stripe assay of retinotectal system. The epitopes of anti-EphA4 and anti-ephrin-A2 were determined to be localized at cysteine rich domain and N-terminal region, respectively. These domains appeared to be essential for Eph receptor-ligand binding. We are now examining the effects of these functional blocking antibodies on motoneuron axonal growth in chick embryos.