To establish the temporal relation of the second heart sound to left ventricular ejection during various conditions of preload, afterload and ventricular function, the electromagnetic aortic flow, aortic pressure, left atrial pressure, external phonocardiogram and electrocardiogram were recorded in 15 open chest chloralose-anesthetized dogs. Propranolol and atropine were infused to hold heart rate and contractility relatively constant. Preload was varied by volume changes, afterload by aortic constriction or angiotensin or methoxamine infusion, and ventricular function by citrate or barbiturate infusion. Fifteen-hundred cycles were analyzed using photographic recordings at 200 mm/sec with 0.04 or 0.004 second time lines. The onset of the second heart sound occurred an average of 5.2 ± 2.6 msec before the nadir of flow reversal and 23.2 ± 5.7 msec after the onset of rapid deceleration of flow. The latter interval could be varied by preload, afterload or changes in ventricular function without changes in the relation of the second sound to flow reversal. Recordings of the first derivative of aortic flow revealed the onset of the second heart sound to occur simultaneously with peak deceleration of aortic flow (0.3 ± 2 msec) under all hemodynamic conditions. External and internal systolic intervals correlated well during all maneuvers ( r = 0.90, P <0.001). These data indicate that the second sound is temporally related to flow reversal, not the onset of rapid deceleration, and support the hypothesis that the second sound results from the peak deceleration of forward flow and from the acceleration of backflow in the aorta.