MUSCARINIC SIGNLING: REGULATION OF VENTRICULAR FUNCTION

Rhode Island Hospital, Providence RI

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Abstract

DESCRIPTION (provided by applicant): The mechanical performance of the heart is determined by its intrinsic contractile properties and is subject to counterbalancing regulation by the autonomic nervous system. While a-adrenergic stimulation increases pump function, cholinergic stimulation decreases it by reducing the conductance of the L-type Ca2+ channel (ICa-L) previously elevated by cAMP ("accentuated antagonism"). Two different G proteins, Go and Gi-2, have been implicated to be absolutely required for this effect, but their exact role and interplay as well as the signaling mechanisms involved are still poorly defined. We have preliminary data suggesting that Go protein may also regulate Ca2+ cycling and force generation by altering excitation contraction (E-C) coupling and the responsiveness of the myofilaments to Ca2+. The overall goal of this investigation is to define the signaling mechanisms that link Go and/or Gi2 protein activation to the muscarinic regulation of ventricular Ca2+ fluxes and contractile function and to test the hypothesis that selective interference with this pathway can be utilized to enhance contractile function in vivo. The Specific Aims are: (1) to test the hypothesis that expression of activated Gao regulates Ca2+ cycling and cell shortening in the ventricular myocardium by increasing (rather than decreasing) E-C coupling gain and myofilament responsiveness to Ca2+ in concert to its blunting effect on ICa-L, (2) to test whether cell-permeable peptide import can be adapted for introduction of dominant negative inhibitory peptides into adult ventricular cardiocytes as an alternative and possible improvement to adenoviral gene transfer, (3) to test the hypotheses that Go and Gi2 proteins act in concert as mediators of M2-receptor effects on ICa-L, Ca2+ cycling and contractility and that both Galpha and Gbetagamma contribute to this effect (4) to test the hypothesis that interference with Go and/or Gi-mediated signal transduction in the ventricular myocardium can be utilized to enhance ventricular contractile function. Muscarinic "accentuated antagonism" is of physiological importance both under normal and pathophysiological conditions. A better understanding of the processes involved in muscarinic receptor-mediated opposition of beta-adrenergic stimulation and blunting of contractile function may form the basis for therapeutic interventions aimed at the long-term autonomic input to the heart through a so far underexplored mechanism.