Studies on oxygen and extracellular fluid restrictions in cultured heart cells

Although cultured heart cells are increasingly used for the study of cardiac metabolism, relatively little is known about their energy turnover. We studied the effects of anoxia with simultaneous restrictions of the volume of the extracellular medium ("ischaemia") on high energy phosphate catabolism in cells from neonatal rat ventricles, cultured for 5 days. The cells were incubated for up to 4 h in Ham-F10 medium either in the presence or in the absence of glucose. High energy phosphates in cell extracts and AMP catabolites in the incubation medium were measured by high pressure liquid chromatography. ATP and creatine phosphate content in normoxic cells did not change significantly, either in the presence or absence of glucose, and the values were similar to those found in the heart in vivo. Energy rich phosphates decreased during anoxia, and were more rapidly depleted during simultaneous oxygen deprivation and volume restriction. Glucose delayed the decline in high energy phosphates. In the presence of glucose, hypoxanthine uptake was higher during normoxia than in anoxia, whereas in "ischaemic" conditions some hypoxanthine was produced. In the absence of glucose, only minor changes were observed in hypoxanthine levels during anoxia, but hypoxanthine production was marked when anoxia was coupled with extracellular volume restriction. Adenosine levels were below the limit of detection. Inosine release was relatively low under all conditions. Xanthine release did not show variation, and anoxia suppressed urate production. Oxygen and glucose deprivation thus led to various degrees of ATP and creatine phosphate breakdown in cultured neonatal heart cells both during anoxia and in simulated "ischaemia".

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