In vivo NMR studies are carried out at the level of cell suspensions, perfused organs, and intact experimental animals in order to determine the mechanisms by which chemicals and other types of stress produce irreversible cell injury. Physiological, biochemical, and magnetic resonance measurements are carried out in parallel when possible, both to validate the techniques used, and more importantly, to determine differences which arise as a consequence of the selectivity of each type of measurement for different cellular pools. During the past year, this parallel approach has been used to demonstrate significant differences between the response of cytosolic and mitochondrial high energy phosphate pools resulting from ischemic stress. In order to establish cause/effect relationships, it is desirable to correlate measurements of many metabolic parameters, some of which are directly accessible to NMR observation, and some of which can be made accessible via the introduction of NMR active indicators into the cells of interest. Most effort has been focused on the use of intracellular fluorinated indicators for cytosolic calcium which can be detected by F-19 NMR. Basal free calcium levels in beating and arrested perfused rat hearts have been measured, and the response to ischemia and hypoxia studied. Cytosolic calcium levels are initially unchanged in response to ischemia, but gradually increase to micromolar levels, consistent with a role for cytosolic calcium in the mediation of irreversible cell injury. More recent studies have shown that cardioplegic arrest, which is known to retard ischemic injury, also delays the increase in cytosolic calcium relative to simple ischemia.
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