Anchored phosphatases modulate glucose homeostasis.

Endocrine release of insulin principally controls glucose homeostasis. Nutrient-induced exocytosis of insulin granules from pancreatic β-cells involves ion channels and mobilization of Ca(2+) and cyclic AMP (cAMP) signalling pathways. Whole-animal physiology, islet studies and live-β-cell imaging approaches reveal that ablation of the kinase/phosphatase anchoring protein AKAP150 impairs insulin secretion in mice. Loss of AKAP150 impacts L-type Ca(2+) currents, and attenuates cytoplasmic accumulation of Ca(2+) and cAMP in β-cells. Yet surprisingly AKAP150 null animals display improved glucose handling and heightened insulin sensitivity in skeletal muscle. More refined analyses of AKAP150 knock-in mice unable to anchor protein kinase A or protein phosphatase 2B uncover an unexpected observation that tethering of phosphatases to a seven-residue sequence of the anchoring protein is the predominant molecular event underlying these metabolic phenotypes. Thus anchored signalling events that facilitate insulin secretion and glucose homeostasis may be set by AKAP150 associated phosphatase activity

Glucose- and Hormone-Induced cAMP Oscillations in α- and β-Cells Within Intact Pancreatic Islets

OBJECTIVEcAMP is a critical messenger for insulin and glucagon secretion from pancreatic beta- and alpha-cells, respectively. Dispersed beta-cells show cAMP oscillations, but the signaling kinetics in cells within intact islets of Langerhans is unknown.RESEARCH DESIGN AND METHODSThe subplasma-membrane cAMP concentration ([cAMP](pm)) was recorded in alpha-and beta-cells in the mantle of intact mouse pancreatic islets using total internal reflection microscopy and a fluorescent translocation biosensor. Cell identification was based on the opposite effects of adrenaline on cAMP in alpha- and beta-cells.RESULTSIn islets exposed to 3 mmol/L glucose, [cAMP](pm) was low and stable. Glucagon and glucagon-like peptide-1(7-36)-amide (GLP-1) induced dose-dependent elevation of [cAMP](pm), often with oscillations synchronized among beta-cells. Whereas glucagon also induced [cAMP](pm) oscillations in most alpha-cells, &lt; 20% of the alpha-cells responded to GLP-1. Elevation of the glucose concentration to 11-30 mmol/L in the absence of hormones induced slow [cAMP](pm) oscillations in both alpha- and beta-cells. These cAMP oscillations were coordinated with those of the cytoplasmic Ca2+ concentration ([Ca2+](i)) in the beta-cells but not caused by the changes in [Ca2+](i) . The transmembrane adenylyl cyclase (AC) inhibitor 2'5'-dideoxyadenosine suppressed the glucose- and hormone-induced [cAMP](pm) elevations, whereas the preferential inhibitors of soluble AC, KH7, and 1,3,5(10)-estratrien-2,3,17-beta-triol perturbed cell metabolism and lacked effect, respectively.CONCLUSIONSOscillatory [cAMP](pm) signaling in secretagogue-stimulated beta-cells is maintained within intact islets and depends on transmembrane AC activity. The discovery of glucose- and glucagon-induced [cAMP](pm) oscillations in alpha-cells indicates the involvement of cAMP in the regulation of pulsatile glucagon secretion.</p

A Novel Method for Simulating Insulin Mediated GLUT4 Translocation

Glucose transport in humans is a vital process which is tightly regulated by the endocrine system. Specifically, the insulin hormone triggers a cascade of intracellular signals in target cells mediating the uptake of glucose. Insulin signaling triggers cellular relocalization of the glucose transporter protein GLUT4 to the cell surface, which is primarily responsible for regulated glucose import. Pathology associated with the disruption of this pathway can lead to metabolic disorders, such as type II diabetes mellitus, characterized by the failure of cells to appropriately uptake glucose from the blood. We describe a novel simulation tool of the insulin intracellular response, incorporating the latest findings regarding As160 and GEF interactions. The simulation tool differs from previous computational approaches which employ algebraic or differential equations; instead, the tool incorporates statistical variations of kinetic constants and initial molecular concentrations which more accurately mimic the intracellular environment. Using this approach, we successfully recapitulate observed in vitro insulin responses, plus the effects of Wortmannin- like inhibition of the pathway. The developed tool provides insight into transient changes in molecule concentrations throughout the insulin signaling pathway, and may be employed to identify or evaluate potentially critical components of this pathway, including those associated with insulin resistance. In the future, this highly tractable platform may be useful for simulating other complex cell signaling pathways

Functional, metabolic and transcriptional maturation of human pancreatic islets derived from stem cells

Transplantation of pancreatic islet cells derived from human pluripotent stem cells is a promising treatment for diabetes. Despite progress in the generation of stem-cell-derived islets (SC-islets), no detailed characterization of their functional properties has been conducted. Here, we generated functionally mature SC-islets using an optimized protocol and benchmarked them comprehensively against primary adult islets. Biphasic glucose-stimulated insulin secretion developed during in vitro maturation, associated with cytoarchitectural reorganization and the increasing presence of alpha cells. Electrophysiology, signaling and exocytosis of SC-islets were similar to those of adult islets. Glucose-responsive insulin secretion was achieved despite differences in glycolytic and mitochondrial glucose metabolism. Single-cell transcriptomics of SC-islets in vitro and throughout 6 months of engraftment in mice revealed a continuous maturation trajectory culminating in a transcriptional landscape closely resembling that of primary islets. Our thorough evaluation of SC-islet maturation highlights their advanced degree of functionality and supports their use in further efforts to understand and combat diabetes. Pancreatic islets derived from stem cells are benchmarked against primary cells.Peer reviewe

Pulsatility of insulin release – a clinically important phenomenon

The mechanisms and clinical importance of pulsatile insulin release are presented against the background of more than half a century of companionship with the islets of Langerhans. The insulin-secreting β-cells are oscillators with intrinsic variations of cytoplasmic ATP and Ca2+. Within the islets the β-cells are mutually entrained into a common rhythm by gap junctions and diffusible factors (ATP). Synchronization of the different islets in the pancreas is supposed to be due to adjustment of the oscillations to the same phase by neural output of acetylcholine and ATP. Studies of hormone secretion from the perfused pancreas of rats and mice revealed that glucose induces pulses of glucagon anti-synchronous with pulses of insulin and somatostatin. The anti-synchrony may result from a paracrine action of somatostatin on the glucagon-producing α-cells. Purinoceptors have a key function for pulsatile release of islet hormones. It was possible to remove the glucagon and somatostatin pulses with maintenance of those of insulin with an inhibitor of the P2Y1 receptors. Knock-out of the adenosine A1 receptor prolonged the pulses of glucagon and somatostatin without affecting the duration of the insulin pulses. Studies of isolated human islets indicate similar relations between pulses of insulin, glucagon, and somatostatin as found during perfusion of the rodent pancreas. The observation of reversed cycles of insulin and glucagon adds to the understanding how the islets regulate hepatic glucose production. Current protocols for pulsatile intravenous infusion therapy (PIVIT) should be modified to mimic the anti-synchrony between insulin and glucagon normally seen in the portal blood