Motor skill acquisition is associated with changes in the cortical representation of specific muscles involved in the task (Pascual-Leone et al. 1994; Karni et al. 1995; Classen et al. 1998; Lotze et al. 2003; Perez et al., 2004). It is less well understood whether plastic changes associated to motor skill acquisition also take place in the spinal cord. Here we report a series of experiments in which we investigated the effect of motor skill training on leg motor cortical and spinal neuronal circuits. In healthy volunteers we tested the effect of 32 min training of two motor tasks with different degree of difficulty, a novel motor skill task involving the ankle muscles and a control task involving simple voluntary dorsi- and plantarflexion movements. We assessed tibialis anterior motor evoked potentials (MEPs) and soleus H-reflex size before and after training. We observed an increase of the MEP size and a decrease in the soleus H-reflex size following the motor skill training session. No changes were observed after the control task. To elucidate the mechanisms underlying the increased MEPs size we tested intracortical inhibition and facilitation using the paired-pulse TMS technique. Our results showed a decrease in intracortical inhibition with no changes in intracortical facilitation following the motor skill training, suggesting that removal of cortical inhibition may contribute to training-induced cortical plasticity. To explore the mechanisms contributing to the H-reflex depression we measured the size of the long-latency depression of the soleus H-reflex evoked by peroneal nerve stimulation (D1 inhibition) and the size of the monosynaptic Ia facilitation of the soleus H-reflex evoked by femoral nerve stimulation. The D1 inhibition was increased and the femoral nerve facilitation was decreased following the motor skill training, suggesting an increase in presynaptic inhibition of Ia afferents. Together these observations suggest that selective changes in cortical as well as spinal neuronal mechanisms occur during acquisition of a novel motor skill involving the ankle muscles in healthy humans. The increased leg motor cortical excitability observed in our study in relation to motor skill acquisition likely reflects adaptations in the motor cortex related to the difficulty in the motor task. The increased presynaptic gating of sensory inputs to spinal motorneurones during motor skill acquisition may ensure that the motor cortex could control the movement based on integration of visual and proprioceptive input relatively undisturbed by the influence of the sensory feedback at the spinal level. In other words these finding could reflect a shift of importance in the control of the movement from spinal feedback mechanisms to direct cortical control.