J Mol Cell Cardiol23, 325-331 (1991) A Novel Positive Inotropic Substance Enhances Contractility Without Increasing the C3+ Transient in Rat Myocardium Claudia Ferroni, Osamu Hano, Carlo Vcptlllp, Fdward G. Lakatta, Michael Klockow’, Harold Spurgeon and Maurizio C. Capogrossi* Laboratory of Cardiovascular Science, Gerontology, Research Center, National Instutue on Aging, National Institutes of Health, Baltimore, MD 21224, USA and ‘E. Merck, Postfach 41 19, 6100 Darmstadt I, Frankfurter Strasse 2.50, Germany (Received 9 May 1990, accepted in revised form I October 1990) C. FERRONI, 0. HANO, C. VENTURA, E. G. LAKA-ITA, M. KL~CKOW, H. SPURCEON AND M. C. CAPOGROSSI. A Novel Positive Inotropic Substance Enhances Contractility Without Increasing the Ca2+ Transient in Rat Myocardium. Journal of Moleculur uad Cellular Curdiofogy (1991) 23, 325-331. We have investigated the mechanism of action of a novel positive inotropic agent, the thiadiasinone derivative 5-( 1-(3,4- dimethoxybenzoyl)-l,2,3,4-tetrahydrochinolin-6-yl)-6methyl-3,6-dihydro-2H-l,3,4-thiadiazin-2-on (EMD 53998). This subsance inhibits phosphodiesterase III and, in skinned myocardial fibers, it increases myofila- ment sensitivity to Ca *+ However the effects of EMD 53998 on intact myocardial preparations are still . undefined. In isolated rat hearts EMD 53998 (0.5 to 5 m) had a dose-dependent effect to increase left ventricular systolic pressure. In isolated left ventricular myocytes loaded with the ester derivative of the Gas+ probe indo-1, EMD 53998 (0.5 to 5 war) enhanced twitch amplitude without increasing the associated indo-l transient. The myofilament responsiveness to Ca’+ was assessed as the relationship between twitch and the indo-l transient amplitudes as the latter is varied by altering the bathing [Ca’+], or stimulation pattern. EMD 53998 reversibly shifted this relationship to the left which indicates that for indo-l transients of the same amplitude in the absence and presence of the drug, twitch amplitude was enhanched by EMD 53998. In isolated myocytes studied in the absence of electrical stimulation, EMD 53998. (1.5 to 5 w) had a concentration-dependent effect to markedly and reversibly decrease cell length without increasing indo-l fluorescence ratio. Thus, the cellular basis for the positive inotropic action of EMD 53998 in rat myocardium is related to the unique effect of this substance to enhance myo6Iament responsiveness to Gas+ and not to an increase in the indo-l transient amplitude. KEY WORDS: Cardiotonic drugs; Thiadiazinone; EMD 53998; Indo-l; Excitation-ontraction coupling. llltrOdUCti0~ Two general mechanisms through which the contractility of the heart can be increased are an enhancement in the amplitude of the cyto- solic [Ca2+], (Cq) transient associated with each action potential and an increase in myo- filament responsiveness to Ca2+ (Blink and Endoh, 1986). Most positive inotropic agents, through different mechanisms, increase cell and sarcoplasmic reticulum (SR) Ca2+ load- ing and lead to an increase in magnitude of the Car transient. However, when cell Ca2+ exceeds an optimum a further increase in Ca2+ decreases contractile amplitude (Capo- grossi et al., 1988), increases diastolic tonus and is associated with the development of arrhythmias (Vassalle et al., 1962; Ferrier, 1976; Capogrossi et al., 1987, 1988). This condition of “Ca2+ overload” represents a major limitation of most positive inotropic drugs. Pharmacologic agents that act pre- dominantly via an increase in the myofila- ment responsiveness to Ca2+, i.e. without enhancing cell Ca2+ loading, are not cur- rently available. For example, although sul- mazole (Riiegg, 1986; van Meel, 1987), isomazole (Lues et al., 1988) and pimobendan (Riiegg, 1986; van Meel, 1987; Fujino et al., 1988a,b) shift leftward the force pCa rela- tionship of chemically skinned cardiac fibers, * Please address all correspondence to; Maurizio C. Capogrossi, Laboratory of Cardiovascular Science, Geronto- logy Research Center, National Institute on Aging, National Institutes of Health, 4940 Eastern Avenue, Baltimore, MD 21224, USA. 0022-2828/91/030325 + 07 303.00/O 0 1991 Academic Press Limited