Polarized neurites (axons and dendrites) form the functional circuitry of the nervous system. Secreted guidance cues often control the polarity of neuron migration and neurite outgrowth by regulating ion channels. Here, we show that secreted semaphorin 3A (Sema3A) induces the neurite identity of Xenopus spinal commissural interneurons (xSCINs) by activating Ca_V2.3 channels (Ca_V2.3). Sema3A treatment converted the identity of axons of cultured xSCINs to that of dendrites by recruiting functional Ca_V2.3. Inhibition of Sema3A signalling prevented both the expression of Ca_V2.3 and acquisition of the dendrite identity, and inhibition of Ca_V2.3 function resulted in multiple axon-like neurites of xSCINs in the spinal cord. Furthermore, Sema3A-triggered cGMP production and PKG activity induced, respectively, the expression of functional Ca_V2.3 and the dendrite identity. These results reveal a mechanism by which a guidance cue controls the identity of neurites during nervous system development.