Glucose regulation of free Ca2+ in the endoplasmic reticulum of mouse pancreatic beta cells

Abstract

Free Ca2+ was measured in organelles of individual mouse pancreatic beta cells loaded with the low affinity indicator furaptra. After removal of cytoplasmic indicator by controlled digitonin permeabilization the organelle Ca2+ was located essentially in the endoplasmic reticulum (ER), >90% being sensitive to inhibition of sarco(endo)plasmic reticulum Ca2+-ATPases. The Ca2+ accumulation in the ER of intact beta cells depended in a hyperbolic fashion on the glucose concentration with half-maximal and maximal filling at 5.5 and >20 mM, respectively. Also elevation of cytoplasmic Ca2+ by K+ depolarization significantly enhanced the Ca2+ accumulation. In permeabilized beta cells 1-3 mM ATP caused rapid Ca2+ filling of the ER reaching almost 500 μM. At 50 nM, Ca2+ ER became half-maximally filled at 45 μM ATP, whereas only 3.5 μM ATP was required at 200 nM Ca2+. Inositol 1,4,5-trisphosphate induced a rapid release of about 65% of the ER Ca2+, and its precursor phosphatidylinositol 4,5-bisphosphate was found to slowly mobilize 75% by another mechanism. It is concluded that glucose is an efficient stimulator of Ca2+ uptake in the ER of pancreatic beta cells both by increasing ATP and cytoplasmic Ca2+. Because physiological concentrations of cytoplasmic ATP are in the mM range, Ca2+ sequestration can be anticipated to be modulated by factors reducing its ATP sensitivity.