Phosphorus-31 nuclear magnetic resonance studies of pig adrenal glands

We have used phosphorus-31 nuclear magnetic resonance to study the secretion of adenosine 5'-triphosphate (ATP) from the medulla of perfused pig adrenal glands. The resonances of the nucleotide pools in the chromaffin granules and cytoplasm are clearly resolved and therefore the intragranular and cytoplasmic processes involving ATP can be monitored simultaneously in the gland during secretion. Secretion of nucleotide during a 3 h continuous stimulation by infusion of acetylcholine chloride was monitored by the decrease in intensity of the intragranular ATP resonances. Up to 40% of the total intragranular nucleotide was released under these conditions. The rate of secretion decreased with duration of stimulation. No significant changes in the steady-state levels of cytoplasmic ATP or in oxygen consumption were observed. The intragranular pH in ischaemic glands was 5.52 + 0.15, while in glands which had been perfused until their cytoplasmic nucleotide levels had recovered, the intragranular pH was 5.76 +/- 0.16. These results provide the first estimate of the internal pH of the chromaffin granules in intact perfused adrenal glands and show that no net acidification occurs in the presence of cytoplasmic ATP. However, the isolated chromaffin granule possesses a proton-pumping adenosine 5'-triphosphatase which, in the presence of a permeant counter-ion, such as chloride, acidifies the granule interior. It is, therefore, suggested that in the intact cell, the cytoplasmic concentration of permeant counter-ions is too low to allow electrically neutral proton accumulation in the granules.