Gamma-aminobutyric acid (GABA) inhibition shapes motor cortex output, gates synaptic plasticity in the form of long-term potentiation, and plays an important role in motor learning. Remarkably, recent studies have shown that acute cardiovascular exercise can improve motor memory, but the cortical mechanisms are not completely understood. We investigated whether an acute bout of lower-limb high-intensity interval (HIT) exercise could promote motor memory formation in humans through changes in cortical inhibition within the hand region of the primary motor cortex. We used TMS to assess the input–output relationship, along with inhibition involving GABAA and GABAB receptors. Measures were obtained before and after a 20-min session of HIT cycling (exercise group) or rest (control group). We then had the same participants learn a new visuomotor skill and perform a retention test 5 hr later in the absence of sleep. No differences were found in corticomotor excitability or GABAB inhibition; however, synaptic GABAA inhibition was significantly reduced for the exercise group but not the control group. HIT exercise was found to enhance motor skill consolidation. These findings link modification of GABA to improved motor memory consolidation after HIT exercise and suggest that the beneficial effects of exercise on consolidation might not be dependent on sleep.