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 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 beta-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 KATP+ channel closure but it is not associated with or a consequence of events occurring subsequent to KATP+ channel closure. These results indicate that AK-catalyzed phosphotransfer may be a determinant of ATP to ADP conversion rates in the KATP+ 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.
Olson
L K
LK
Department of Pharmacology, Division of Diabetes, Endocrinology, and Metabolism, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Schroeder
W
W
Robertson
R P
RP
Goldberg
N D
ND
Walseth
T F
TF
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000263:Adenylate Kinase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002384:Catalysis; D002460:Cell Line; D006224:Cricetinae; D005110:Extracellular Space / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism; D007773:Lactates / Q000096:biosynthesis; D019344:Lactic Acid; D008564:Membrane Potentials; D008647:Mesocricetus; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D015221:Potassium Channels / Q000378:metabolism
D007328:Insulin; D007773:Lactates; D010761:Phosphorus Radioisotopes; D015221:Potassium Channels; D019344:Lactic Acid; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000263:Adenylate Kinase; D005947:Glucose; D002118:Calcium