Functional insulin receptors are known to occur in pancreatic beta cells; however, except for a positive feedback on insulin synthesis, their physiological effects are unknown. Amperometric measurements at single, primary pancreatic beta cells reveal that application of exogenous insulin in the presence or absence of nonstimulatory concentrations of glucose evokes exocytosis mediated by the beta cell insulin receptor. Insulin also elicits increases in intracellular Ca 2؉ concentration in beta cells but has minimal effects on membrane potential. Conditions where the insulin receptor is blocked or cell surface concentration of free insulin is reduced during exocytosis diminishes secretion induced by other secretagogues, providing evidence for direct autocrine action of insulin upon secretion from the same cell. These results indicate that the beta cell insulin receptor can mediate positive feedback for insulin secretion. The presence of a positive feedback mechanism for insulin secretion mediated by the insulin receptor provides a potential link between impaired insulin secretion and insulin resistance.

The signaling pathway by which insulin stimulates insulin secretion and increases in intracellular freeInsulin secreted by pancreatic ␤-cells is the primary regulator of serum glucose concentrations in mammals. Although substantial progress has been made in elucidating the mechanisms responsible for normal regulation of insulin secretion from the ␤-cell, many aspects of this process remain unclear. In particular, chemical and physiological interactions between cells within the islet exert an important level of control in the physiological regulation of insulin secretion that is not entirely understood. Both hormonal and neuronal influences within islets may modulate ␤-cell activity and insulin secretion in vitro and in vivo (1-3). Although such influences have been demonstrated, the existence of significant autocrine effects of insulin on ␤-cells remained controversial for many years because a variety of studies yielded conflicting evidence on the modulation of insulin secretion by insulin in whole islets or in vivo. Recently, however, a variety of new methods have been utilized that demonstrate potent and possibly clinically important autocrine actions of insulin.Several recent studies have indicated that ␤-cells express components of insulin signaling systems including insulin receptors (4 -6), insulin receptor substrates (IRS-1 and IRS-2) 1 (7-9), phosphatidylinositol 3-kinase (PI3-K) (10, 11), and protein kinase B (12). Evidence has also been obtained indicating that insulin released by glucose can activate these components in addition to other proteins in the cells. Insulin binds to receptors on the surface of ␤-cells (4, 13) and activates tyrosine phosphorylation of insulin receptors (6), insulin receptor substrates (8), and PHAS-I (an inhibitor of mRNA cap-binding protein) (14). Furthermore, maximal glucose-stimulated production of phosphatidylinositol 3,4,5-triphosphate (PIP 3 ), a major product of PI3-K activity, coincides with the early peak phase insulin secretion in islets and clonal ␤-cells (10). Thus, autocrine activation of the ␤-cell insulin receptors and several downstream proteins has been demonstrated.Some of the physiological consequences of insulin receptor activation at ␤-cells have recently been revealed. Activation of the insulin signaling pathway in ␤-cells leads to initiation of insulin synthesis at both transcriptional and translational levels, increasing the cellular content of releasable hormone in primary and clonal ␤-cell cultures (14 -16). In ␤TC6-F7 cells transfected to overexpress the insulin receptor, basal and glucose-stimulated insulin secretion was enhanced compared with kinase negative controls (15). In another report, clonal cells lacking the IRS-1 protein showed both decreased insulin content and glucose-stimulated secretion (17). These latter studies suggest that insulin can exert positive control over synthesis and/or secretion. Direct evidence for the effects of insulin on insulin secretion has been obtained by application of exogenous insulin to isolated ␤-cells and d...

The effects of extracellular Zn 2؉ and pH and intravesicular pH on insulin and 5-hydroxytryptamine (5-HT) secretion from pancreatic beta cells were investigated. Insulin and 5-HT secretion from single cells was detected by amperometry as a series of current spikes corresponding to detection of multimolecular packets secreted by exocytosis. Spike width was used as a measure of the kinetics of clearance from the cell and the area of spikes as a measure of amount released. Changes in extracellular pH from 6.9 to 7.9 caused insulin spikes to become narrower with no change in area, whereas the same treatments had no effect on 5-HT secretion. Treatment of cells with Bafilomycin A 1 or N-ethylmaleimide, both of which are expected to increase intravesicular pH by inhibiting V-type H ؉ -ATPase, had no effect on 5-HT secretion but caused insulin spikes to become more narrow. These results indicate that exposure to high pH, whether intravesicular or extracellular, accelerates release of insulin during exocytosis without affecting the amount of insulin released. Increasing extracellular Zn 2؉ concentration from 0 to 25 M increased the width and decreased the area of insulin spikes without affecting 5-HT secretion. Zn 2؉ effects were likely exerted through a common-ion effect on Zn 2؉ -insulin dissociation. It was concluded that intravesicular storage conditions and extracellular ions can affect free insulin concentration in the vicinity of beta cells during secretion.

An oscillatory increase in pancreatic beta cell cytoplasmic free Ca2+ concentration, [Ca2+]i, is a key feature in glucose-induced insulin release. The role of the voltage-gated Ca2+ channel beta3 subunit in the molecular regulation of these [Ca2+]i oscillations has now been clarified by using beta3 subunit-deficient beta cells. beta3 knockout mice showed a more efficient glucose homeostasis compared to wild-type mice due to increased glucose-stimulated insulin secretion. This resulted from an increased glucose-induced [Ca2+]i oscillation frequency in beta cells lacking the beta3 subunit, an effect accounted for by enhanced formation of inositol 1,4,5-trisphosphate (InsP3) and increased Ca2+ mobilization from intracellular stores. Hence, the beta3 subunit negatively modulated InsP3-induced Ca2+ release, which is not paralleled by any effect on the voltage-gated L type Ca2+ channel. Since the increase in insulin release was manifested only at high glucose concentrations, blocking the beta3 subunit in the beta cell may constitute the basis for a novel diabetes therapy.