Background: Purkinje fibers plays important role of maintenance of long-duration VF. Acetylcholine-related potassium current has known to induce excitation of purkinje fiber, We assessed the role of acetylcholine in maintenance of VF using atropine. Methods: The rabbit hearts(n=3) were retrogradely perfused and action potentials from the left ventricle free wall were recorded using di-4-ANEPPS with a high speed CMOS camera (100x100 pixels, 1000 f/s). Epicardial optical action potentials were recorded with a photodiode array (26��26 mm). The burst pacing was performed to induce VF. Results: As VF progressed, dominant frequency(DF) analysis showed that the highest local DF was located on anterior wall of LV overlying papillary muscle. At 30min of VF, mean DF was immediately reduced by atropine 100關M/L(14 +/-2, 9 +/-2 Hz), with redistribution of the highest local DF at anterior wall of LV (32 +/-5, 15 +/-2 Hz). DF and frequency gradient was decreased in a dose-dependent manner and VF terminated as APD70 (47 +/-2 -> 70 +/-10ms) increases and conduction velocity(CV) decrease (0.27 +/-0.0.8 -> 0.16 +/-0.07) by high dose atropine(500關M/L). Conclusions: Our results suggest that acetylcholine-related potassium current existing in purkinje fiber plays an significant role in rotor stability and VF dynamics, concomitantly supporting importance of purkinje firing in maintenance of long duration VF. Atropine suppress VF by underlying mechanism of reduction of purkinje firing, and may be a potential target as antiarrhythmic agent suppressing VF.