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

L. Karl Olson, William Schroeder, R. Paul Robertson, Nelson D. Goldberg, Timothy F. Walseth

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44 Scopus citations

Abstract

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 18O-labeled phosphoryls of endogenous nucleotides in cells incubated in medium enriched in [18O]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 β-18O-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.

Original languageEnglish (US)
Pages (from-to)16544-16552
Number of pages9
JournalJournal of Biological Chemistry
Volume271
Issue number28
DOIs
StatePublished - 1996

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