Studies of the Ca²⁺ requirements for glucose- and carbachol-induced augmentation of inositol trisphosphate and inositol tetrakisphosphate accumulation in digitonin-permeabilized islets. Evidence for a glucose recognition site in insulin secretion

The metabolism of D-glucose is believed to initiate and regulate insulin secretion by islet β-cells, although the identity of the metabolite which couples glucose metabolism to the cellular events involved in insulin secretion is unknown. An alternative hypothesis involves the presence of a glucoreceptor for which there has been no biochemical evidence. We have investigated whether glucose recognition by the β-cell is coupled to phospholipase C. We have used digitonin-permeabilized, [³H]inositol-prelabeled islets to study glucose and carbachol activation of phospholipase C. In this model, carbachol recognition by its muscarinic receptor was coupled to phospholipase C activation. D-Glucose (but not L-glucose) also stimulated phospholipase C activity in these permeabilized islets. This effect was not due to glucose metabolism since glucose 6-phosphate did not affect phospholipase C activity and since phosphorylation of [³H]glucose was not detectable in digitonin-permeabilized islets. Glucose had no effect on the myo-inositol-1,4,5-trisphosphate-5-phosphatase or 3-kinase activities. In the absence of agonist, free Ca²⁺ concentrations between 0.1 and 1 μM (as determined with a Ca²⁺-specific electrode) did not influence phospholipase C activity. Stimulation of phospholipase C activity by either carbachol or glucose required Ca²⁺ in the submicromolar range and was optimal at 0.5 μM free Ca²⁺.myo-Inositol-1,3,4,5-tetrakisphosphate production from permeabilized islets was synergistically augmented by Ca²⁺ (0.5-10 μM) and glucose. Phospholipase C activity in islets is therefore not directly activated by free Ca²⁺ concentrations in the submicromolar range. Furthermore, glucose per se activates phospholipase C activity independently of glucose metabolism. A working hypothesis based on these findings is that glucose is recognized by a site which is coupled to phospholipase C in islets.