MECHANISMS OF SLOWED MYOCARDIAL RELAXATION

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DESCRIPTION (adapted from the applicant's abstract): The overall goal of this research is to determine the roles of calcium and cross-bridge kinetics in the slowing of myocardial relaxation during 4 physiological conditions relevant to ischemia/reperfusion: hypoxia, reoxygenation, acidosis and inotropic stimulation with Ca-sensitizers. The rationale is that the mechanisms of impaired relaxation are unresolved. Previous studies emphasized the importance of slowing of the decline of cytosolic [calcium] ([Ca]c) in slowed relaxation. However [Ca]c decline does not correlate well with relaxation because [Ca]c influences force indirectly through several intervening steps. Relaxation is determined by the decline in the state of activation in response to [Ca]c and by cross-bridge kinetics, but these have not been measured during conditions relevant to ischemia/reperfusion. The proposed experiments will determine the relative contributions of [Ca]c, Ca-activation, and cross-bridge kinetics to slowed relaxation. The experimental strategy will be to use isolated rat cardiac trabeculae. [Ca]c will be measured with indo-1 fluorescence. Ca-activation will be determined by applying a brief muscle stretch to detach all cross-bridges and reduce force to zero; Ca-activation (the ability of cross-bridges to attach and develop force) will be assessed from the initial rate of force redevelopment after the stretch. The time course of Ca-activation will be defined by applying stretches at different times during twitches. Cross-bridge kinetics will be assessed from the maximum shortening velocity. The specific aim will be to test 4 hypotheses: 1. With hypoxia, slowed [Ca]c decline causes slowed Ca-activation decline which causes slowed relaxation. 2. With reoxygenation, slowed Ca-activation decline and slowed cross-bridge kinetics causes slowed relaxation. 3. With hypoxia, acidosis causes slowed [Ca]c decline, slowed Ca-activation decline, and relaxation. 4. The myofilament Ca-sensitizer Levosimendan does not slow Ca-activation decline, and relaxation.