Regulation of steady-state free Ca2+ levels by the ATP/ADP ratio and orthophosphate in permeabilized RINm5F insulinoma cells.
Stimulation of insulin secretion in the pancreatic beta-cell by a fuel such as glucose requires the metabolism of the fuel and is accompanied by increases in oxygen consumption and intracellular free Ca2+. A very early signal for these events could be a decrease in the cytosolic ATP/ADP ratio due to fuel phosphorylation. To test this hypothesis the regulation of free Ca2+ was evaluated in permeabilized RINm5F insulinoma cells that sequester Ca2+ and maintain a low medium free Ca2+ concentration (set point), between 100 and 200 nM, in the presence of Mg2+ and ATP. ATP, creatine, creatine phosphate, and creatine phosphokinase were added to the media to achieve various constant ratios of ATP/ADP. Free Ca2 was monitored using fura-2. The results demonstrated that the steady-state free Ca2+ concentration varied inversely with the ATP/ADP ratio and orthophosphate (Pi) levels. In contrast, no correlation between free Ca2+ and the phosphorylation potential (ATP/ADP.Pi) was found. Regulation of the Ca2+ set point by the ATP/ADP ratio was observed at ratios between 5 and 50 and at Pi concentrations between 1 and 7 mM, irrespective of whether mitochondria were participating in Ca2+ sequestration or were inhibited. Increasing the ATP/ADP ratio stimulated Ca2+ uptake by the nonmitochondrial pool but did not modify Ca2+ efflux. Glucose 6-phosphate (1 mM) had no effect on the Ca2+ set point. The data suggest that variations in the cytosolic ATP/ADP ratio induced by fuel stimuli may regulate Ca2+ cycling across nonmitochondrial compartments and the plasma membrane by modulating the activity of Ca2+ -ATPases. A mechanism linking fuel metabolism and cytosolic ATP/ADP ratio to activation of the Ca2+ messenger system in pancreatic beta-cells is proposed.
Corkey
B E
BE
Department of Biochemistry, Boston University School of Medicine, Massachusetts 02118.
Deeney
J T
JT
Glennon
M C
MC
Matschinsky
F M
FM
Prentki
M
M
D007516:Adenoma, Islet Cell / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D000968:Antimycin A / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002463:Cell Membrane Permeability; D007340:Insulinoma / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D009840:Oligomycins / Q000494:pharmacology; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism* / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000378:metabolism
D009840:Oligomycins; D010710:Phosphates; D000244:Adenosine Diphosphate; D000968:Antimycin A; D000255:Adenosine Triphosphate; D008274:Magnesium; D002118:Calcium