Glucose stimulation of insulin release in the absence of extracellular Ca2+ and in the absence of any increase in intracellular Ca2+ in rat pancreatic islets.

Abstract

Insulin secretion has been studied in isolated rat pancreatic islets under stringent Ca2l-depleted, Ca2+-free conditions. Under these conditions, the effect of 16.7 mM glucose to stimulate insulin release was abolished. Forskolin, which activates adenylyl cyclase, also failed to stimulate release in the presence of either low or high glucose concen- (6, 7). Another important effect of glucose with an enhancing effect on insulin release is time-dependent potentiation. This pathway too is Ca2+ dependent (14). In contrast to these Ca2+-dependent effects, we report here the existence of an interesting Ca2+-independent effect of glucose to augment insulin secretion when protein kinase C (PKC) is activated in the f3 cell. Furthermore, when protein kinase A (PKA) and PKC are activated simultaneously, this Ca2+-independent augmentation of insulin release is of large magnitude. MATERIALS AND METHODSIsolation of Pancreatic Islets and Insulin Secretion. Pancreatic islets were isolated from adult male Sprague-Dawley rats by collagenase dispersion as described (6, 15). Insulin release was measured under both static incubation and perifusion conditions. In static incubations, batches of five size-matched islets were incubated in 1 ml of Krebs-Ringer bicarbonate (KRB) buffer containing 129 mM NaCl, 5 mM NaHCO3, 4.8 mM KCl, 1.2 mM KH2PO4, 2.5 mM CaCl2, 1.2 mM MgSO4, 2.8 mM glucose, 0.1% bovine serum albumin, and 10 mM Hepes (pH 7.4) for 60 min at 37°C (preincubation). Then the incubation medium was removed by aspiration and 1 ml of fresh KRB buffer containing test substances was introduced. Incubation under the test conditions was then continued for 30 or 60 min at 37°C. At the end of the incubations, the medium was aspirated and kept at -200C until radioimmunoassayed for insulin. Rat insulin was used as a standard for radioimmunoassay. When Ca2+-depleted conditions were needed, KRB buffer devoid of Ca2+ with different concentrations of EGTA was used throughout the experiments (for washing the islets and both the 60-min preincubation and the 60-min experimental incubation). When nitrendipine and thapsigargin were used, they were present throughout the preincubation and incubation periods. When norepinephrine was used, it was present during experimental incubation periods only. In perifusion experiments, 25 size-matched islets were placed in each 0.7-ml perifusion chamber (16,17) The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Adiponectin induces insulin secretion in vitro and in vivo at a low glucose concentration

“…However, a Ca 2+ -independent pathway has also been proposed. Thus, Komatsu et al have suggested that both Ca 2+ -dependent and Ca 2+ -independent augmentation occurs via a pathway dependent on glucose metabolism [32,33]. In this context, we confirmed that adiponectin is able to stimulate insulin secretion when the influx of Ca 2+ through voltage-dependent Ca 2+ channels is blocked by nitrendipine; in other words, adiponectin stimulates insulin secretion without requiring influx of Ca 2+ through voltagedependent Ca 2+ channels.…”

Oscillatory control of insulin secretion

“…This K ATP channel-independent process is mostly considered as an amplification mechanism, which remains functionally silent unless [Ca 2+ ] i is elevated. Yet other studies have provided evidence that the K ATP channel-independent pathway may also exhibit triggering properties, since glucose under some conditions stimulates insulin release in a Ca 2+ -independent manner (Komatsu et al, 1995;Komatsu et al, 1996;Komatsu et al, 2001). The K ATP channel-independent pathway has remained elusive and many mechanisms have been proposed (reviewed in (Komatsu et al, 2001)).…”

Intracellular pH Plays a Critical Role in Glucose-Induced Time-Dependent Potentiation of Insulin Release in Rat Islets

“…The basal glucose concentration used was 2.8 mmol/l, whereas the priming glucose concentration was 11.1 mmol/l in Ca 2ϩ -free conditions and 16.7 mmol/l in the presence of Ca 2ϩ . These are the optimal concentrations of glucose for the stimulation of insulin secretion in the absence and presence of Ca 2ϩ , respectively (22). Stimulation of islets in the test period, as distinct from the priming period, was always done with 16.7 mmol/l glucose in the presence of Ca 2ϩ .…”