RELATION BETWEEN GLUCOSE-STIMULATED INSULIN-SECRETION AND INTRACELLULAR CALCIUM ACCUMULATION STUDIED WITH A SUPERFUSION SYSTEM OF A GLUCOSE-RESPONSIVE PANCREATIC BETA-CELL LINE MIN6

The concept that cytosolic free calcium is the primary signal for insu lin secretion is generally accepted, but studies with intact pancreati c beta-cells of the cytosolic free calcium concentration-insulin secre tion relationship have produced contradictory and sometimes confusing data. We designed a superfusion system of a pancreatic beta-cell line, MIN6, loaded with fura-2, which allowed simultaneous measurement of c ytosolic free calcium concentration and insulin secretion. MIN6 cells released insulin in response to high glucose, thus resembling events i n normal islet cells. Cytosolic free calcium concentration and insulin secretion rapidly increased, and the increase was suppressed by manno heptulose or by sodium azide. This increase was suppressed by lowering the temperature of the medium. Cytosolic free calcium concentration a nd the insulin secretion induced by leucine were not influenced by man noheptulose but were inhibited by sodium azide. In RINm5F cells, cytos olic free calcium concentration and insulin release were slightly supp ressed by glucose but were increased by ionomycin. There was a close r elation between the rise in cytosolic free calcium concentration and i nsulin secretion in all cases. Our findings provide a direct evidence that a rise in cytosolic free calcium concentration depends on glucose metabolism and is a primary signal for insulin secretion.