Role of inositol trisphosphate in preconditioning

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DESCRIPTION (provided by applicant): Brief, nonlethal episodes of myocardial ischemia paradoxically protect the heart and profoundly limit infarct size caused by a subsequent sustained ischemic insult. There is general agreement that this phenomenon, termed "ischemic preconditioning," is initiated by stimulation of G-protein coupled receptors during the brief ischemic stimulus. However, despite intensive study, the second messenger pathway(s) initiated by receptor stimulation and culminating in this profound endogenous cardiac protection remain unresolved. Evidence from our laboratory has implicated inositol (1,4,5)-trisphosphate (InsP3) signaling - i.e., release of the second messenger InsP3; stimulation of InsP3 receptors; and resultant alterations in intracellular calcium concentration - as a potential "trigger" for infarct size reduction with preconditioning in rabbit heart. Moreover, the cardioprotection achieved with preconditioning is mimicked by the synthetic, membrane-impermeable analog, + ____________________ D-myo-InsP3, presumably via stimulation of as-yet poorly characterized 'external' InsP3 receptors. Our current aims are to: (1) document the existence of 'external' InsP3 receptors and elucidate their contribution to the reduction in infarct size seen with brief preconditioning ischemia and exogenous administration of D-myo-InsP3; (2) determine whether ischemic preconditioning and D-myo-InsP3 achieve cardioprotection via common downstream signaling pathways (with emphasis on the possible role(s) of protein kinase C, mitogen-activated protein kinases, and ATP-sensitive potassium channels) and (3) determine whether Insp3 represents a common cellular mediator among species for infarct size reduction with preconditioning. Aims 1 and 2 will be addressed in the isolated buffer-perfused rabbit heart model of regional ischemia, with infarct size delineated by tetrazolium staining and the involvement of protein kinase C, etc assessed using selective pharmacologic antagonists and quantitative biochemical assays. Aim 3 will involve the measurement of infarct size, and integrated tissue sampling for biochemical analysis, in the in vivo canine model of coronary artery occlusion/reperfusion. If InsP3 proves to be an important cellular mediator contributing to infarct size reduction with preconditioning, this insight into the signal transduction pathways triggered by brief antecedent ischemia may ultimately assist in the design of novel and benign therapeutic strategies to prophylactically render human myocardium resistant to ischemia and infarction.