NMR VISIBILITY OF P(I) IN PERFUSED RAT HEARTS IS AFFECTED BY CHANGES IN SUBSTRATE AND CONTRACTILITY

Nuclear magnetic resonance (NMR) spectroscopy does not always detect the total metabolite content in isolated perfused rat hearts. Alterations in NMR peak areas therefore could be caused by a change either in the metabolite content per se or in its NMR visibility. We have therefore compared the ATP, phosphocreatine (PCr), and P(i) content of hearts, as determined by P-31-NMR spectroscopy, with the total, chemically determined ATP, PCr, and P(i) contents of the same hearts to determine the fractions, if any, that are NMR invisible under different perfusion conditions. The three perfusion buffers used contained 1) glucose, 2) glucose plus high K+, and 3) pyruvate. Fully relaxed P-31-NMR spectra were collected during the final 10 min of perfusion in each group, and the ATP, PCr, and P(i) contents were quantified using methylene diphosphonate as an external standard. The hearts were then freeze-clamped and chemically assayed for ATP, PCr, and P(i). Under all three conditions, the NMR-determined ATP and PCr contents were almost identical to the chemically determined values. However, only a portion of the chemically determined P(i) was NMR visible. During perfusion with glucose-containing buffer, 39 +/- 8% of the total P(i) was NMR visible, and this decreased to 12 +/- 2% (P < 0.01) during K+ arrest and to 9 +/- 5% (P < 0.01) during perfusion with pyruvate-containing buffer. In conclusion, whereas the total cellular content of ATP and PCr is always NMR visible during normoxic perfusion, alterations in substrate and contractile status can change the fraction of NMR-visible P(i).