The effect of atriopeptin III (ANF) was studied on K+, Ca2+ and Na+ currents of single heart cells of 10-day-old chick embryos and 17- to 20-week old human fetuses. ANF III (2 x 10(-9) M) greatly decreased the basal and cAMP prestimulated high-threshold (IL) Ca2+ current without affecting the low-threshold Ca2+ current (IT) or the TTX-sensitive fast inward Na+ current. ANF III was also found to increase the delayed outward K+ currents (IK) in a dose-dependent fashion (10(-10) to 10(-7) M). The effects of ANF III on IK and IL were reversible upon washout of this hormone. Increasing intracellular guanosine 3',5'-cyclic monophosphate (cGMP) blocked IL but had no effect on IK. These results suggested that ANF regulated one type of Ca2+ current and the delayed outward K+ current in single heart cells. The effects of ANF on IL (but not on IK) could be due in part to the increased [cGMP]i levels or to an unknown signal transduction that is stimulated by this hormone.