Aims: An inwardly rectifying K⁺ current is present in atrial cardiac myocytes that is characteristically activated by acetylcholine I_KACh. Physiologically activation of the current in the SA node is important in slowing the heart rate with increased parasympathetic tone. It forms a now canonical signalling pathway and is a paradigm for the direct regulation of signalling effectors by the GbgG-protein subunit. Many of the molecular questions have been addressed in heterologous expression systems with less focus on the behaviour in native myocytes. This is partly because of the technical difficulties in undertaking comparable studies in these cells. In this study we undertake the characterisation of potassium current in the atrial-derived cell line HL-1. Methods: By using an electrophysiological approach, we compare the characteristics of the potassium current with those in native atrial cells and in a HEK cell line expressing the cloned Kir3.1/3.4 channel. Results: The potassium current recorded in HL-1 is inwardly rectifying, inhibited by Tertiapin-Q (P<0.004) and activated by the muscarinic agonist carbachol. Carbachol-activated currents were inhibited by Pertussis toxin (P<0.003). The basal current was time-dependently increased (P<0.025) when GTP was substituted in the patch-clamp pipette by the non-hydrolysable analogue GTPtgS. Using antibody staining we look at the distribution of GIRK4 in HL-1 where a strong membrane signal was detected. RT-PCR confirms the presence of mRNA for the main G- protein signalling protein, as well as for M2 muscarinic and A1 adenosine receptors. We compared the kinetics of current modulation in HL- 1 with those of freshly isolated atrial mouse cardiomyocytes. The overall characteristics of the current activation and deactivation kinetics of HL-1 cells is comparable to the one measured in atrial cardiomyocytes. Conclusion: The data suggest HL-1 cells are a good model to study I_KACh.