Wnt5a induces simultaneous cortical axon outgrowth and repulsive turning through distinct signaling mechanisms.

Wnt5a is thought to propel cortical axons down the corticospinal tract and through the corpus callosum by repulsive mechanisms. We cultured dissociated early postnatal cortical neurons from hamsters and exposed them to a gradient of Wnt5a as a model for studying the mechanism of Wnt5a effects. Turning assays indicated that cortical axons were repelled away from a point source of Wnt5a. Surprisingly, during the 1-hour turning assay, axons exposed to Wnt5a also increased their growth rates by almost 50%. Ryk receptors but not Frizzled (Fz) receptors were required for Wnt5a-promoted axon outgrowth, whereas both Ryk and Fz receptors were required for repulsive growth-cone turning. Both Ryk and Fz receptors mediated calcium (Ca(2+)) signaling, which is required for axon outgrowth and repulsive turning. Treatments with pharmacological inhibitors revealed that distinct Ca(2+) signaling mechanisms were involved in Wnt5a-dependent axon outgrowth versus repulsive guidance. Ca(2+) release from intracellular stores through inositol 1,4,5-trisphosphate receptors was required for Wnt5a-induced axon outgrowth but not for repulsive turning. In contrast, Ca(2+) entry through transient receptor potential channels was required for both repulsive growth-cone turning and Wnt5a-increased axon outgrowth. Taken together, these results showed that a guidance cue can induce increased rates of axon outgrowth simultaneously with repulsive guidance and may provide an understanding of how cortical axons may be repelled down the spinal cord in vivo. Moreover, we demonstrate that previously unidentified Wnt signaling pathways differentially mediate these growth-cone behaviors.