Suppression of adenylate kinase catalyzed phosphotransfer precedes and is associated with glucose-induced insulin secretion in intact HIT-T15 cells

Adenine nucleotide metabolism was characterized in intact insulin secreting HIT-T15 cells during the transition from non-stimulated (i.e. 0.2 mM glucose) to the glucose-stimulated secretory state. Metabolic dynamics were monitored by assessing rates of appearance of O-18-labeled phosphoryls of endogenous nucleotides in cells incubated in medium enriched in [O-18]water. Most prominent of the metabolic alterations associated with stimulated insulin secretion was the suppression in the rate of adenylate kinase (AK)-catalyzed phosphorylation of AMP by ATP, This was manifest as a graded decrease of up to 50% in the rate of appearance of beta-O-18-labeled species of ADP and ATP and corresponded to the magnitude of the secretory response elicited over a range of stimulatory glucose concentrations, The only nucleotide exhibiting a significant concentration change associated with suppression of AK activity was AMP, which decreased by about 50%, irrespective of the glucose concentration, Leucine-stimulated secretion also decreased the rate of AK-catalyzed phosphotransfer. This secretory stimulus-related suppression of AK-catalyzed phosphotransfer occurs within 45 s of glucose addition, precedes insulin secretion, depends on the internalization and metabolism of glucose, and is independent of membrane depolarization and the influx of extracellular calcium. The secretory stimulus-induced decrease in AK-catalyzed phosphotransfer, therefore occurs prior to or at the time of K-ATP(+) channel closure but it is not associated with or a consequence of events occurring subsequent to K-ATP(+) channel closure. These results indicate that AK-catalyzed phosphotransfer may be a determinant of ATP to ADP conversion rates in the K-ATP(+) channel microenvironment; secretory stimuli-linked decreased rates of AK-catalyzed ADP generation from ATP (and AMP) would translate into an increased probability of ATP-liganded and, therefore, closed state of the channel.