The role of acetylcholine (ACh) as a signalling molecule in plants was investigated using a model system of Characeae cells. The effect of ACh on conductance of K+ channels in Nitella flexilis cells and on the action potential generation in Nitellopsis obtusa cells after H+-ATPase inhibition, where repolarization occurs after the opening of outward rectifying K+ channels, was investigated. Voltage-clamp method based on only one electrode impalement was used to evaluate the activity of separate potassium ion transport system at rest. We found that ACh at high concentrations (1 mM and 5 mM) activates K+ channels as the main membrane transport system at the resting state involved in electrogenesis of Characeaen membrane potential. We observed that ACh caused an increase in duration of AP repolarization of cells in K+ state when plasmalemma electrical characteristics are determined by large conductance K+ channels irrespective of whether AP were spontaneous or electrically evoked. These results indicate interference of ACh with electrical cellular signalling pathway in plants.