32P-Labeling Patterns in Rat Pancreatic Islets: Tissue Source of the Radiophosphate Released after Glucose Stimulation*

Abstract

Immediately after stimulation with glucose in vitro, isolated rat pancreatic islets prelabeled with [32P]orthophosphate release a pulse of [32P]orthophosphate into the media (the "phosphate flush"). Islets have been rapidly frozen before, during, and after this pulse to assess the concurrent changes in the distribution of tissue radioactivity. Under the present experimental conditions, approximately 90% of the islet radioactivity was soluble in perchloric acid (PCA-soluble) immediately before stimulation and slightly more than half of that was present as [32P]orthophosphate. The tissue pool of [32P]orthophosphate declined 55% and 62% after 7 and 14 min of stimulation which, respectively, incorporated the peak and the end of the heightened efflux of radioactivity. The net decrementa in tissue orthophosphate could account for all of the radioactivity which was released during the "phosphate flush." During the 14-min period of stimulation, labeled ATP and GTP (which had accounted for 13% and 4% of total PCA-soluble radioactivity before stimulation) increased 51% and 35%, respectively, and labeled ADP and AMP (which had accounted for 5.4% and 1.5% of PCA-soluble counts) fell 36% and 77%, respectively. Certain other PCA-soluble components, such as phosphorylcholine and phosphorylethanolamine, and total PCA-insoluble radioactivity were not demonstrably altered. The findings indicate that the "phosphate flush" originates from a labile pool of tissue orthophosphate. It remains to be established whether the simultaneous changes in the turnover of selected nucleotides are coupled to the translocation of orthophosphate or are mediated separately.