Continuous monitoring of ATP levels in living insulin secreting cells expressing cytosolic firefly luciferase.
The second messenger role of ATP in insulin secretion was investigated in living INS-1 insulinoma cells. ATP-dependent luminescence was monitored in cells expressing high levels of firefly luciferase under the control of the tetracycline-dependent transactivator. The calibration of luminescence in permeabilized cells yielded similar ATP levels as those obtained in cell extracts with a conventional ATP assay. Stimulation of insulin secretion by glucose or methyl-succinate was correlated with rises of cellular ATP in simultaneous measurements. ATP generation was decreased by inhibition of the ADP-ATP translocase. This approach demonstrates the feasibility of defining the dynamic relationship between ATP and other parameters involved in metabolism-secretion coupling.
Maechler
P
P
Department of Internal Medicine, University Medical Centre, Geneva, Switzerland.
Wang
H
H
Wollheim
C B
CB
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D001517:Coleoptera; D003600:Cytosol / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000378:metabolism*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured
D007328:Insulin; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D008156:Luciferases