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In the present study we have investigated the reorganization of the sarcolemmal phospholipids during the first 60 minutes of simulated ischemia ("ischemia") as induced by anoxia, volume restriction, and nutrient deprivation. Experiments were carried out on [3H]acetate-labeled neonatal rat cardiomyocytes and isolated (nonradiolabeled) sarcolemmal membranes obtained from the same culture system. After 60 minutes of "ischemia," cellular high-energy phosphate (ATP) levels had decreased to approximately 40% of the control values, but no significant phospholipid hydrolysis was detected. Labeling experiments using the nonpermeant (primary amine-containing phospholipid) probe trinitrobenzenesulfonic acid and nonlytic treatment with (different) exogenous phospholipases A2 were both indicative of a shifted transbilayer distribution of the hexagonalII phase-preferring and fusion-promoting sarcolemmal phosphatidylethanolamine in favor of the outer membrane leaflet. This specific change in sarcolemmal phospholipid asymmetry preceded the loss of integrity of the sarcolemma, monitored by the release of lactate dehydrogenase as well as by scanning electron microscopy. It is proposed that, in addition to the previously reported lateral phospholipid reorganization, uncontrolled transbilayer movement of the non-bilayer-preferring phosphatidylethanolamine from the inner to the outer leaflet of the sarcolemma is an additional factor in destabilizing the lipid bilayer, eventually leading to the irreversible membrane damage seen after a prolonged period of ischemia.

Loss of asymmetric distribution of sarcolemmal phosphatidylethanolamine during simulated ischemia in the isolated neonatal rat cardiomyocyte.