The objective of the proposal is to study the principal ionic derangements of actue myocardial ischemia -- extracellular K+ accumulation, cytoplasmic acidification and """"""""calcium overload"""""""" -- and to explore the role which these abnormalities play in the electrophysiologic manifestations of ischemia. The proposal is organized around four recent technical innovations: 1. Ability to measure intracellular calcium activity with the fluorescent indicator indo-1. This indicator can be introduced into isolated cells or intact tissue as a cell permeant ester, and permits cancellation of movement artifact by comparison of simultaneous signals at different emission wavelengths. 2. Ability to measure intracellular pH with the cell permeant indicator 6-carboxy-fluorescein diacetate. 3. Ability to measure local extracellular K+ accumulation with miniaturized valinomycin electrodes. 4. Ability to monitor local myocardial depolarization and conduction velocity in intact hearts with an improved monophasic action potential electrode. Principal scientific goals include the following: To establish whether conduction impairment during the first minutes of ischemia is adequately explained by reduction of the resting membrane potential under a variety of experimental conditions. To establish whether early ischemic depolarization is accompanied by an increase in cytoplasmic calcium activity, and whether the time course of the calcium increase depends upon experimental conditions: e.g. heart rate, extracellular calcium, and the presence of calcium channel blockers. To characterize changes in intracellular calcium activity during reperfusion, and the mechanism of postischemic calcium uptake. To determine whether cytoplasmic acidification during ischemia is sufficient to explain resulting physiological abnormalities (e.g. cellular depolarization and contractile failure), and whether these abnormalities are prevented by cytoplasmic alkalinization with NH4C1. To determine whether changes in cytoplasmic calcium or production of organic acids has the correct time course to explain the efflux of K+ ions from ischemic myocardial cells. Interrelated experiments will be performed concurrently in isolated rabbit hearts, isolated canine hearts and tissue cultured ventricular cells exposed to metabolic inhibitors. The latter two preparations permit more reliable assessment of changes in transmembrane potential. Parallel findings in isolated cells and ischemic hearts would facilitate investigation of these phenomena at the subcellular level.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL032093-07
Application #
3343337
Study Section
Cardiovascular and Renal Study Section (CVB)
Project Start
1984-04-01
Project End
1991-03-31
Budget Start
1990-04-01
Budget End
1991-03-31
Support Year
7
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Stanford University
Department
Type
Schools of Medicine
DUNS #
800771545
City
Stanford
State
CA
Country
United States
Zip Code
94305
Wu, Y; Clusin, W T (1997) Calcium transient alternans in blood-perfused ischemic hearts: observations with fluorescent indicator fura red. Am J Physiol 273:H2161-9
Binah, O (1994) Immune effector mechanisms in heart transplant rejection. Cardiovasc Res 28:1748-57
Anderson, M E; Braun, A P; Schulman, H et al. (1994) Multifunctional Ca2+/calmodulin-dependent protein kinase mediates Ca(2+)-induced enhancement of the L-type Ca2+ current in rabbit ventricular myocytes. Circ Res 75:854-61
Lauer, M R; Gunn, M D; Clusin, W T (1992) Endothelin activates voltage-dependent Ca2+ current by a G protein-dependent mechanism in rabbit cardiac myocytes. J Physiol 448:729-47
Mohabir, R; Lee, H C; Kurz, R W et al. (1991) Effects of ischemia and hypercarbic acidosis on myocyte calcium transients, contraction, and pHi in perfused rabbit hearts. Circ Res 69:1525-37
McDonald, T V; Courtney, K R; Clusin, W T (1989) Use-dependent block of single sodium channels by lidocaine in guinea pig ventricular myocytes. Biophys J 55:1261-6
Lee, H C; Clusin, W T (1989) Effect of Bay K8644 on cytosolic calcium transients and contraction in embryonic cardiac ventricular myocytes. Pflugers Arch 413:225-33
Lee, H C; Mohabir, R; Smith, N et al. (1988) Effect of ischemia on calcium-dependent fluorescence transients in rabbit hearts containing indo 1. Correlation with monophasic action potentials and contraction. Circulation 78:1047-59
Blake, K; Clusin, W T; Franz, M R et al. (1988) Mechanism of depolarization in the ischaemic dog heart: discrepancy between T-Q potentials and potassium accumulation. J Physiol 397:307-30
Lee, H C; Smith, N; Mohabir, R et al. (1987) Cytosolic calcium transients from the beating mammalian heart. Proc Natl Acad Sci U S A 84:7793-7

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