Cellular contractility can be altered by two general mechanisms: a change in the amplitude of the calcium signal that triggers contraction or a change in the contractile proteins that alters their response to a given signal. The goal of this project is to elucidate changes in the contractile proteins that modify the contraction. In order to study regulation of the contractile proteins without interference from the electromechanical coupling steps, we use rat cardiac cells in which the surface membrane has been made hyperpermeable and the contractile proteins can be directed activated by Ca++ added to the bath. With this preparation, a physiologically relevant, Ca-independent mechanism of regulating contractility has been demonstrated. This regulatory system is controlled by Beta adrenergic system, and it appears to determine how much of the myosin isozyme with the greatest ATPase activity (V1) will participate in a Ca activated contraction. It should therefore influence not only maximum force but also velocity of shortening.
The specific aims i n the next period of study are to: 1) determine whether the other major myosin isozyme (V3) is regulated by the Alpha adrenergic system; 2) determine whether this type of regulation exists in other mammals; 3) determine whether the cardiac cell can regulate velocity of shortening at a given load and degree of activation by varying the relative amounts of V1 and V3 that participate in the contraction; 4) isolate the active substance in the regulatory system. Hyperpermeable cardiac cells will be studied using standard biophysical techniques for measuring force and velocity in unloaded and afterloaded contractions. The myosin composition of each tissue will be determined with gel electrophoresis in non dissociating media. Dialysis, gel filtration and ion exchange chromatography will be used to isolate the active component of the regulatory system. The control systems that are being studied appear not only to play a major part in the normal regulation of cardiac contractility but also to change with physical conditioning, aging and disturbances in hemodynamics. It is important therefore to understand their normal function as well as their response to various disease states of the cardiovascular system.
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