The multiple functions of a neuron depend on the proper assembly of axonal connections during the development of the nervous systems. This assembly involves the motile behavior of growth cones at the ends of elongating axons. The growth cones express receptors that bind to specific guidance molecules in the local environment. In turn, this initiates the attractive and repulsive forces required to give the appropriate direction to the elongating axon. The process implicates a tightly regulated remodeling of the actin cytoskeleton in response to the activation of the Rho GTPases, Cdc42, Rac and RhoA. In this article, we will review how the ephrin-Eph receptor system regulates the activity of the Rho GTPases, to modulate the mechanics of growth cone activity and then axon guidance.