Effect of the insulin mimetic L-783,281 on intracellular [Ca2+] and insulin secretion from pancreatic β-cells

L-783,281, an antidiabetic fungal metabolite than has previously been shown to activate insulin signaling in CHO cells, was tested for its effect on intracellular Ca2+ ([Ca2+](i)) and insulin secretion in single mouse pancreatic beta-cells. Application of 10 mumol/l L-783,281 for 40 s to isolated beta-cells in the presence of 3 mmol/l glucose increased [Ca2+](i) to 178 +/- 10% of basal levels (n = 18) as measured by fluo-4 fluorescence. L-767,827, an inactive structural analog of the insulin mimetic, had no effect on beta-cell [Ca2+](i). The L-783,281-evoked [Ca2+](i) increase was reduced by 82 +/- 4% (n = 6, P < 0.001) in cells incubated with 1 mumol/l of the SERCA (sarco/ endoplasmic reticulum calcium ATPase) pump inhibitor thapsigargin and reduced by 33 +/- 6% (n = 6, P < 0.05) in cells incubated with 20 mumol/l of the L-type Ca2+-channel blocker nifedipine. L-783,281-stimulated [Ca2+](i) increases were reduced to 31 +/- 3% (n = 9, P <: 0.05) and 48 +/- 10% (n = 6, P < 0.05) of control values by the phosphatidylinositol 3-kinase (PI3-K) inhibitors LY294002 (25 mumol/l) and wortmannin (100 nmol/l), respectively. In beta-cells from IRS-1(-/-) mice, 10 mumol/l L-783,281 had no significant effect on [Ca2+](i) (n = 5). L-783,281 also resulted in insulin secretion at single beta-cells. Application of 10 mumol/l L-783,281 for 40 s resulted in 12.2 2.1 (n = 14) exocytotic events as measured by amperometry, whereas the inactive structural analog had no stimulatory effect on secretion. Virtually no secretion was evoked by L-783,281 in IRS-1(-/-) beta-cells. LY294002 (25 mumol/l) significantly reduced the effect of the insulin mimetic on beta-cell exocytosis. It is concluded that L-783,281 evokes [Ca2+](i) increases and exocytosis in beta-cells via an IRS-1/PI3-K-dependent pathway and that the [Ca2+](i) increase involves release of Ca2+ from intracellular stores.