A monitor of secretion from single pancreatic beta-cells.
ATP is known to be coreleased with insulin from pancreatic beta-cells. To monitor insulin secretion from single beta-cells, a single beta-cell was surrounded in culture by Fura 2-loaded calf pulmonary artery endothelium (CPAE) cells, which can detect the ATP. CPAE cells did not respond with an elevation in cytoplasmic calcium concentration ([Ca2+]i) to either tolbutamide (100 mumol/l) or kainate (1 mmol/l) but did respond with an elevation in [Ca2+]i to ATP (0.1-10 mumol/l) without desensitization and in a dose-dependent manner. A brief application of tolbutamide (10 mumol/l) increased [Ca2+]i in both the beta-cell and the adjacent CPAE cells in co-culture. Suramin (100 mumol/l), an ATP-receptor blocker, inhibited the tolbutamide-induced elevation in [Ca2+]i in the CPAE cells but did not inhibit the elevation in [Ca2+]i in the beta-cell, confirming that the insulin secretagogue-induced Ca2+ response in CPAE cells in co-culture is mediated by ATP released from the beta-cell. When co-culture of the beta-cell and CPAE cells was stimulated by kainate (1 mmol/l) and then tolbutamide (10 mumol/l), the CPAE cells showed elevations in [Ca2+]i in response to kainate and tolbutamide during elevation in [Ca2+]i in the beta-cell. This strongly suggests that insulin secretion as well as an increase in [Ca2+]i in response to different agents, i.e., kainate and tolbutamide, occurs in a single beta-cell. A long exposure of tolbutamide (100 mumol/l, 4 min) resulted in a long-lasting elevation in [Ca2+]i in the beta-cell.(ABSTRACT TRUNCATED AT 250 WORDS)
Kuromi
H
H
Division of Molecular Medicine, Chiba University School of Medicine, Japan.
Mizuno
N
N
Seino
S
S
D000255:Adenosine Triphosphate / Q000032:analysis / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D018920:Coculture Techniques; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D018846:Excitatory Amino Acids / Q000494:pharmacology; D005456:Fluorescent Dyes; D016257:Fura-2; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007328:Insulin / Q000032:analysis / Q000494:pharmacology*; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D007608:Kainic Acid / Q000494:pharmacology; D007700:Kinetics; D011651:Pulmonary Artery; D013997:Time Factors; D014044:Tolbutamide / Q000494:pharmacology
D018846:Excitatory Amino Acids; D005456:Fluorescent Dyes; D007004:Hypoglycemic Agents; D007328:Insulin; D000255:Adenosine Triphosphate; D014044:Tolbutamide; D007608:Kainic Acid; D002118:Calcium; D016257:Fura-2