During very long underwater excursions, diving mammals experience hypoxic conditions (arterial PO2+15 torr) that would be fatal to humans and other terrestrial animals. Previous work from this laboratory has shown a number adaptations to hypoxia by muskrat hearts. The adaptations involve an increase in the energy supply and a reduction in energy demand of the heart during hypoxia. Recent studies from this laboratory have shown that mitochondria isolated from muskrat hearts have a greater calcium uptake capability than guinea pig heart mitochondria and this may be of benefit in the calcium overload which may occur during hypoxia, ischemia, acidosis and the calcium paradox. Isolated muskrat hearts demonstrate a negative relationship between force of contraction and heart rate. This finding also suggests that calcium dynamics in muskrat hearts may differ from that seen in the hearts of terrestrial animals, as most hearts have a positive force-frequency relationship. The proposed study would test the hypothesis that control of intracellular calcium is different in muskrats hearts compared with guinea pig hearts and this difference relates to the hypoxia resistance seen in the muskrat heart. The proposed study would use NMR techniques to follow changes in cytosolic calcium, pH, ATP and CrP in muskrat and rabbit hearts which are subjected to 60 min of substrate-free hypoxia followed by 15 min of reoxygenation. 5-F-BAPTA, a calcium binding dye, would be loaded into a myocytes suspension and changes in free calcium followed using 19F NMR. The various phosphorous containing compounds would be followed using 31P NMR. The effect of the Na-Ca exchanger would be evaluated by subjecting myocytes to hypoxia and reoxygenation in a medium containing a Na-Ca exchange inhibitor and measuring the NMR spectra during this procedure.A second part of the study involves calcium transport in cell organelles. Purified sarcolemmal and both junctional and longitudinal sarcoplasmic reticulum vesicles would be prepared from muskrat and rabbit hearts. Millipore filtration techniques would be used to evaluate the calcium transport properties of the two Ca-ATPases, the Na-Ca exchanger and the calcium release channel. In the event that the first part of the study cannot be completed using NMR, then changes in calcium concentration would be followed by using the calcium indicator Indo-1 and fluorometric techniques. ATP and CrP would be measured using standard chemical techniques.
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| McKean, T; Scherzer, A; Park, H (1997) Hypoxia and ischaemia in buffer-perfused toad hearts. J Exp Biol 200:2575-81 |
| McKean, T; Mendenhall, W (1996) Comparison of the responses to hypoxia, ischaemia and ischaemic preconditioning in wild marmot and laboratory rabbit hearts. J Exp Biol 199:693-7 |
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| McKean, T A; Rocklage, A; Schneider, R J (1993) Glibenclamide binding in vertebrate cardiac membranes. J Exp Biol 182:275-81 |
