In certain nocturnal moth species the frequency range of best hearing shifts to higher frequencies during repeated sound stimulation. This could provide the moths with a mechanism to better detect approaching echolocating bats. However, such a dynamic up-tuning would be of little value for day-flying moths that use intra-specific acoustic communication. Here we examined if the ears of day-flying moths provide stable tuning during longer sound stimulation. Contrary to our expectations, dynamic up-tuning was found in the ear of the day-flying species Urania boisduvalii and Empyreuma pugione. Audiograms were measured with distortion-product otoacoustic emissions (DPOAEs). The level of the dominant distortion product (i.e. 2f1-f2) varied as a function of time by as much as 45 dB during ongoing acoustic stimulation, showing a systematic decrease at low frequencies and an increase at high frequencies. As a consequence, within about 2 s of acoustic stimulation, the DPOAEs audiogram shifted from low to high frequencies. Despite the up-tuning, the range of best audition still fell within the frequency band of the species-specific communication signals, suggesting that intra-specific communication should not be affected adversely. Up-tuning could be an ancestral condition in moth ears that in day-flying moths does not underlie larger selection pressure.