Emptying of intracellular Ca2+ stores stimulates Ca2+ entry in mouse pancreatic beta-cells by both direct and indirect mechanisms
(1997) The Journal of physiology — Vol. 503, n° Pt 2, p. 387-398 (1997)
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- 1. In non-excitable cells, the depletion of intracellular Ca2+ stores triggers Ca2+ influx by a process called capacitative Ca2+ entry. In the present study, we have investigated how the emptying of these stores by thapsigargin (1 microM) influences Ca2+ influx in electrically excitable pancreatic beta-cells. The cytoplasmic Ca2+ concentration ([Ca2+]i) was monitored in clusters of mouse beta-cells or in whole islets loaded with fura-2. 2. The membrane was first held hyperpolarized by diazoxide, an opener of ATP-sensitive K+ (KATP) channels, in the presence of 4.8 mM K+. Alternating between Ca(2+)-free medium and medium containing 2.5 mM Ca2+ caused a minor rise in [Ca2+]i (approximately 14 nM) in clusters of beta-cells. A larger rise (approximately 65 nM), resistant to the blockade of voltage-dependent Ca2+ channels by D600, occurred when extracellular Ca2+ was readmitted after emptying intracellular Ca2+ stores with thapsigargin or acetylcholine. Thus there exists a small capacitative Ca2+ entry in beta-cells. 3. When the membrane potential was clamped at depolarized levels with 10, 20 or 45 mM K+ in the presence of diazoxide, [Ca2+]i increased to different plateau levels ranging between 100 and 900 nM. Thapsigargin consistently caused a further transient rise in [Ca2+]i, but had little (at 10 mM K+) or no effect on the plateau level. This confirms that the capacitative Ca2+ entry is small. 4. In clusters of cells whose membrane potential was not clamped with diazoxide, 15 mM glucose (in 4.8 mM K+) induced [Ca2+]i oscillations by promoting Ca2+ influx through voltage-dependent Ca2+ channels. The application of thapsigargin accelerated these oscillations and increased their amplitude, sometimes causing a sustained elevation of [Ca2+]i. Similar results were obtained from whole islets perifused with a medium containing > or = 6 mM glucose. The effect of thapsigargin was always much larger than expected from the capacitative Ca2+ entry, probably because of a potentiation of Ca2+ influx through voltage-dependent Ca2+ channels. 5. This potentiating effect of thapsigargin did not result from an acceleration of cell metabolism since the drug did not affect glucose-induced changes in NAD(P)H fluorescence. It is also unlikely to involve the inhibition of KATP channels because thapsigargin steadily elevated [Ca2+]i in cells in which [Ca2+]i oscillations persisted in the presence of a maximally effective concentration of tolbutamide. 6. In conclusion, the emptying of intracellular Ca2+ stores in beta-cells induces a small capacitative Ca2+ entry and activates a depolarizing current which potentiates glucose-induced Ca2+ influx through voltage-dependent Ca2+ channels.
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Miura, Y., Henquin, J.-C., & Gilon, P. (1997). Emptying of intracellular Ca2+ stores stimulates Ca2+ entry in mouse pancreatic beta-cells by both direct and indirect mechanisms. The Journal of physiology, 503(Pt 2), 387-398. https://doi.org/10.1111/j.1469-7793.1997.387bh.x (Original work published 1997)