Phosphorus nuclear magnetic resonance of bovine platelets

31P nuclear magnetic resonance (NMR) spectra of bovine platelets at 28 degrees C display prominent peaks from adenine nucleotides in two different environments, the metabolic and storage pools. Addition of 20 mM 2-deoxy-D-glucose and 0.1 mM 2,4-dinitrophenol depletes metabolic ATP, resulting in loss of nucleotide signals at 5, 10, and 19 ppm and leaving peaks at 6.5, 10.5, and 19 ppm that are assigned primarily to the dense granule storage pool. The ATP/ADP concentration ratio of the remaining pools is 1.9 +/- 0.2, with chemical shifts which reflect a more acidic environment for the intragranular nucleotides than for the cytosol. Isolated bovine dense granules give spectra with similar peak positions to those ascribed to the storage pool after metabolic depletion of whole cells. Storage pool nucleotide spectra are highly temperature dependent. Below 20 degrees C, the beta-ATP peak broadens and decreases in area to the point that it is almost undetectable at 0 degrees C, probably reflecting formation of progressively higher molecular weight aggregates at the lower temperatures. The release reaction was followed by 31P NMR after addition of thrombin at 28 degrees C. Spectra of the cell suspension taken at 5-min intervals were compared to spectra of perchloric acid extracts produced in parallel. A new finding of these experiments is that the released nucleotides rapidly undergo a transition to a "NMR silent" form as part of the bovine platelet release reacton. Metabolic breadkdown and external paramagnetic ion interactions did not explain the loss of signal from the storage pool. During release reaction, the nucleotides may be broadened out beyond detection due to immobilization by divalent cation and/or platelet membrane binding.