CARDIAC CELL VOLUME REGULATION IN HEALTH AND DISEASE

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DESCRIPTION (adapted from the applicant's abstract): The proposed studies focus on one aspect of myocardial injury, cardiac cell edema, and the underlying mechanisms of cell volume regulation. Lipid metabolites, including lysolipids derived from diacyl and plasmalogen species (e.g., lysophosphatidylcholine, lysoplasmenylcholine), long chain acylcarnitine, and arachidonic acid are reported to rapidly accumulate during ischemia and reflow and have been implicated in the ensuing electrophysiological disturbances. The central question to be evaluated here is: Do ischemic lipid metabolites disrupt cardiac cell volume regulation under isosmotic and anisosmotic conditions? It is postulated that lipid metabolites: (1) Cause cell swelling in the absence of an osmotic gradient by altering passive ion permeation and ion transport; (2) Exacerbate cell swelling in the presence of an osmotic gradient by increasing the permeability of cardiac cell membranes to water (i.e., hydraulic conductivity); and (3) Alter the response to swelling by modulating the function of stretch-activated channels and other ion permeation processes. The proposed experiments combine novel techniques to address the following aims: (1) The effects of lipid metabolites on cell volume under isosmotic conditions will be measured by digital video microscopy; (2) Hydraulic conductivity (water permeability) will be calculated from the kinetics of volume changes in response to osmotic stress; (3) Transport processes involved in lipid metabolite-induced volume changes will be identified by ion substitution studies and specific blockers; (4) Ion selective microelectrodes will be used to measure changes in the intracellular ion activities of K+, Na+, and Cl-; (5) The effect of metabolites on stretch- activated currents will be determined by simultaneous perforated-patch voltage clamp and cell volume measurements, and the magnitude of volume changes and currents will be related; and (6) The effects of lipid metabolites on sarcolemmal fluidity will be determined by fluorescence recovery after photobleaching (FRAP) in intact cells.