BACKGROUND: Regular physical activity (PA) has shown substantial cardiac benefits. We sought to investigate whether habitual PA is associated with changes of the electrical action potential duration, as it is represented by the QT duration on a rest ECG, in a population based sample of middle-aged and elderly individuals of Ikaria island. METHODS: In a cross-sectional survey 1071 inhabitants of Ikaria Island (65 ± 13 years, 47% males) were enrolled. PA was estimated by means of IPAQ classifying the participants into low, moderate and vigorous group. QT duration was measured from a surface electrocardiogram; while using Bazett's formula the heart-rate-corrected QT (QTc) was calculated. RESULTS: Among participants, 85% reported at least moderate PA levels. Women in the 'vigorous' and 'moderate' PA level compared to those in the 'low' PA level had significantly shorter QTc (408 ± 2 ms vs. 411 ± 1 ms vs. 419 ± 2 ms, P = 0.001, respectively). In contrast, no significant difference in QTc according to PA levels was observed in men (P = 0.053). Linear regression analysis revealed that PA level was significantly associated with shorter QTc in women after adjustment for established confounders; while no such association was evident in men. Furthermore, compared to the 'low' PA group, women in the 'vigorous' PA group were 5.5-times less likely to have QTc interval above 450 ms (P = 0.031). CONCLUSION: Increased PA is associated with shorter QTc interval only in middle-aged and elderly women of Ikaria Island irrespectively of participant's habits or medical conditions, illustrating gender differences in the cardioprotective effect of habitual exercise.