Transcranial DC stimulation (tDCS) is a plasticity-inducing noninvasive brain stimulation tool with various potential therapeutic applications in neurological and psychiatric diseases. Currently, the duration of the aftereffects of stimulation is restricted. For future clinical applications, stimulation protocols are required that produce aftereffects lasting for days or weeks. Options to prolong the effects of tDCS are further prolongation or repetition of tDCS. Nothing is known thus far about optimal protocols in this behalf, although repetitive stimulation is already performed in clinical applications. Thus we explored the effects of different break durations on cathodal tDCS-induced cortical excitability alterations. In 12 subjects, two identical periods of cathodal tDCS (9-min duration; 1 mA) with an interstimulation interval of 0 (no break), 3, or 20 min or 3 or 24 h were performed. The results indicate that doubling stimulation duration without a break prolongs the aftereffects from 60 to 90 min after tDCS. When the second stimulation was performed during the aftereffects of the first, a prolongation and enhancement of tDCS-induced effects for ≤ 120 min after stimulation was observed. In contrast, when the second stimulation followed the first one after 3 or 24 h, the aftereffects were initially attenuated, or abolished, but afterwards re-established for up to 120 min after tDCS in the 24-h condition. These results suggest that, for prolonging the aftereffects of cathodal tDCS, stimulation interval might be important.