Membrane potential dependent modulations of calcium oscillations in insulin-secreting INS-1 cells

Abstract

This study was undertaken to examine the role of K+ channels on cytosolic Ca2+ ([Ca2+]i) in insulin secreting cells. [Ca2+]i was measured in single glucose-responsive INS-1 cells using the fluorescent Ca2+ indicator Fura-2. Glucose, tolbutamide and forskolin elevated [Ca2+]i and induced [Ca2+] oscillations. Whereas the glucose effect was delayed and observed in 60% and 93% of the cells, in a poorly and a highly glucose-responsive INS-1 cell clone, respectively, tolbutamide and forskolin increased [Ca2+]i in all cells tested. In the latter clone, glucose induced [Ca2+]i oscillations in 77% of the cells. In 16% of the cells a sustained rise of [Ca2+]i was observed. The increase in [Ca2+]i was reversed by verapamil, an L-type Ca2+ channel inhibitor. Adrenaline decreased [Ca2+]i in oscillating cells in the presence of low glucose and in cells stimulated by glucose alone or in combination with tolbutamide and forskolin. Adrenaline did not lower [Ca2+]i in the presence of 30mM extracellular K+, indicating that adrenaline does not exert a direct effect on Ca2+ channels but increases K+ channel activity. As for primary b-cells, [Ca2+]i oscillations persisted in the presence of closed KATP channels; these also persisted in the presence of thapsigargin, which blocks Ca2+ uptake into Ca2+ stores. In contrast, in voltage-clamped cells and in the presence of diazoxide (50mM), which hyperpolarizes the cells by opening KATP channels, [Ca2+]i oscillations were abolished. These results support the hypothesis that [Ca2+]i oscillations depend on functional voltage-dependent Ca2+ and K+ channels and are interrupted by a hyperpolarization in insulin-secreting cells.