All positive inotropic agents that are available for clinical use exercise their effect predominately via an increase in cell calcium loading. However once the myocardial preparation has reached@ @its peak contractile response a further increase in cell calcium loading is associated initially with a plateau and then with a decline in the inotropic state of the muscle, an increase in diastolic tone, aftercontractions and arrhythmias. This condition which has been defined as """"""""calcium overload"""""""" represents the limiting factor in the clinical use of positive inotropic agents. Thus, it is desirable to develop drugs that increase the contractility of the heart via an enhancement in myofilament responsiveness to calcium rather than by increasing the extent of cell calcium loading. We tested the effect of novel thiadiazinone derivatives (designed by E. Merck, Darmstadt, FRG) that vary in potency (1) to sensitize skinned myocardial Tibers to Ca 2+: EMD 54622 > 53998 > 54650,-and (2) to inhibit phosphodiesterase Ill in cell homogenates (54650 > 53998 > 54622). We determined whether differential Ca 2+-myofilament effects are expressed in intact guinea pig cells, bathed in Hepes buffer (23 degree C) and loaded with the fluorescent Ca 21 probe, indo-1. Our results show that the steepness of relationship among twitches and indo-1 fluorescence transients measured across drug dose varies as 54622 > 53998 > 54650. Thus, differential modifications of the myofilament Ca 2+ response in intact cells can be effected via molecular modifications of thiadiazinone that enhance its potency to sensitize myofilaments to Ca 2+.