Mechanism Of Insulin Secretion Research Articles

The PLC/PKC/Ras/MEK/Kv channel pathway is involved in uncarboxylated osteocalcin-regulated insulin secretion in rats

Uncarboxylated osteocalcin, a bone matrix protein, has been proposed to regulate glucose metabolism by increasing insulin secretion, improving insulin sensitivity and stimulating β cell proliferation. Our previous study also indicated that uncarboxylated osteocalcin stimulates insulin secretion by inhibiting voltage-gated potassium (KV) channels. The goal of this study is to further investigate the underlying mechanisms for the regulation of Kv channels and insulin secretion by uncarboxylated osteocalcin. Insulin secretion and Kv channel currents were examined by radioimmunoassay and patch-clamp technique, respectively. Calcium imaging system was applied to measure intracellular Ca2+ concentration ([Ca2+]i). The protein levels were detected by western blot. The results showed that uncarboxylated osteocalcin potentiated insulin secretion, inhibited Kv channels and increased [Ca2+]i compared to control. These effects were suppressed by phospholipase-C (PLC)/protein kinase C (PKC)/Ras/MAPK-ERK kinase (MEK) signaling pathway, indicating that this signaling pathway plays an important role in uncarboxylated osteocalcin-regulated insulinotropic effect. In addition, the results also showed that adenylyl cyclase (AC) did not influence the effect of uncarboxylated osteocalcin on insulin secretion and Kv channels, suggesting that AC is not involved in uncarboxylated osteocalcin-stimulated insulin secretion. These findings provide new insight into the mechanism of uncarboxylated osteocalcin-regulated insulin secretion.

Urea impairs β cell glycolysis and insulin secretion in chronic kidney disease.

Disorders of glucose homeostasis are common in chronic kidney disease (CKD) and are associated with increased mortality, but the mechanisms of impaired insulin secretion in this disease remain unclear. Here, we tested the hypothesis that defective insulin secretion in CKD is caused by a direct effect of urea on pancreatic β cells. In a murine model in which CKD is induced by 5/6 nephrectomy (CKD mice), we observed defects in glucose-stimulated insulin secretion in vivo and in isolated islets. Similarly, insulin secretion was impaired in normal mouse and human islets that were cultured with disease-relevant concentrations of urea and in islets from normal mice treated orally with urea for 3 weeks. In CKD mouse islets as well as urea-exposed normal islets, we observed an increase in oxidative stress and protein O-GlcNAcylation. Protein O-GlcNAcylation was also observed in pancreatic sections from CKD patients. Impairment of insulin secretion in both CKD mouse and urea-exposed islets was associated with reduced glucose utilization and activity of phosphofructokinase 1 (PFK-1), which could be reversed by inhibiting O-GlcNAcylation. Inhibition of O-GlcNAcylation also restored insulin secretion in both mouse models. These results suggest that insulin secretory defects associated with CKD arise from elevated circulating levels of urea that increase islet protein O-GlcNAcylation and impair glycolysis.

Modulation of alkaloid content, growth and productivity of Trigonella foenum-graecum L. using irradiated sodium alginate in combination with soil applied phosphorus

Trigonella foenum-graecum L. (fenugreek) belongs to family Fabaceae and has been used as medicinal herb since times immemorial. Fenugreek seed-alkaloid, known as trigonelline, controls diabetes through operation of the mechanism of insulin secretion, modulation of β cell regeneration and stimulation of activity of glucose metabolism related enzymes. Gamma-irradiated sodium alginate (ISA) elicits plant defence responses and biological activities in plants. A pot experiment was conducted to explore the effect of foliage-applied ISA (0, 40, 80 and 120mgL−1) alone and in combination with soil-applied phosphorus (40kgPha−1) on growth, yield and quality attributes of fenugreek employing the soil deficient in phosphorus. Un-irradiated sodium alginate (UN 40mgL−1) and de-ionized water were used as control. Of the treatments employed, 80mgL−1 of ISA applied with 40kgPha−1 (P40) proved the best. It increased the chlorophyll content by 24.85 and 27.40%, carotenoids content by 15.00 and 23.52% at 60 and 90days after sowing, respectively. Besides, it increased the seed yield by 131.0%, trigonelline content by 17.84%, trigonelline yield by 174.0%, seed alkaloid content by 32.98% and seed alkaloid yield by 208.64% over the control. Gel permeation chromatography of ISA revealed the formation of low molecular weight fractions which might be responsible for plant growth promotion in this study. Trigonelline content was determined by isocratic HPLC equipment (Model, LC-20AD).

Inhibition of voltage-gated potassium channels mediates uncarboxylated osteocalcin-regulated insulin secretion in rat pancreatic β cells

Insulin secretion from pancreatic β cells is important to maintain glucose homeostasis and is regulated by electrical activities. Uncarboxylated osteocalcin, a bone-derived protein, has been reported to regulate glucose metabolism by increasing insulin secretion, stimulating β cell proliferation and improving insulin sensitivity. But the underlying mechanisms of uncarboxylated osteocalcin-modulated insulin secretion remain unclear. In the present study, we investigated the relationship of uncarboxylated osteocalcin-regulated insulin secretion and voltage-gated potassium (KV) channels, voltage-gated calcium channels in rat β cells. Insulin secretion was measured by radioimmunoassay. Channel currents and membrane action potentials were recorded using the conventional whole-cell patch-clamp technique. Calcium imaging system was used to analyze intracellular Ca2+ concentration ([Ca2+]i). The data show that under 16.7mmol/l glucose conditions uncarboxylated osteocalcin alone increased insulin secretion and [Ca2+]i, but with no such effects on insulin secretion and [Ca2+]i in the presence of a KV channel blocker, tetraethylammonium chloride. In the patch-clamp experiments, uncarboxylated osteocalcin lengthened action potential duration and significantly inhibited KV currents, but had no influence on the characteristics of voltage-gated calcium channels. These results indicate that KV channels are involved in uncarboxylated osteocalcin-regulated insulin secretion in rat pancreatic β cells. By inhibiting KV channels, uncarboxylated osteocalcin prolongs action potential duration, increases intracellular Ca2+ concentration and finally promotes insulin secretion. This finding provides new insight into the mechanisms of osteocalcin-modulated insulin secretion.

Microfluidic Device for the Measurement of Amino Acid Secretion Dynamics from Murine and Human Islets of Langerhans.

Islets of Langerhans are the regulators of in vivo blood glucose levels through the secretion of endocrine hormones. Amino acids, released from various cells within islets or from intrapancreatic neurons, are hypothesized to further adjust hormone secretions. In contrast to the well-accepted mechanism of glucose-stimulated insulin secretion, several questions remain as to the function of amino acids in the regulation of hormone release from islets. To understand the autocrine and paracrine roles that amino acids play in islet physiology, a microfluidic system was developed to perform online monitoring of the secretion profiles of amino acids from 2-5 islets. The device contained an islet chamber with the ability to perfuse stimulants and an amino acid measurement system with derivatization and electrophoretic separation integrated on a single microchip. The setup was optimized to allow -15 kV to be applied to the device for high efficiency and rapid separations of derivatized amino acids. The compositions of the derivatization and separation buffers were optimized to prevent precipitations in the channels, which allowed continuous monitoring of secretion for over 2 h. With this method, 10 amino acids were resolved with limits of detection ranging from 1 to 20 nM. When murine islets were perfused with 3 mM glucose, the secretion rates of 9 amino acids were measured and ranged from 30 to 400 fmol islet(-1) min(-1). As the glucose concentration was increased to 20 mM, the dynamic changes of amino acids were monitored. The biological relevance of the amino acid secretions was verified using 2,4-dinitrophenol as an inhibitor of the proton motive force. The microfluidic system was also used to measure dynamic changes of amino acid release from human islets, which showed different release profiles compared to their murine counterparts.

Antihyperglycemic and Insulin Secretagogue Activities of Abrus precatorius Leaf Extract.

Aim:Abrus precatorius leaves methanolic extract (APME) was evaluated for in vivo antihyperglycemic activity and in vitro insulinotropic effect.Materials and Methods:In vivo antihyperglycemic and insulin secretagogue activities were assessed in streptozotocin-induced diabetic rats by oral administration of APME (200 mg/kg body weight [bw]) for 28 days. In vitro insulin secretion mechanisms were studied using mouse insulinoma beta cells (MIN6-β). In vivo body weight and blood glucose and in vivo and in vitro insulin levels were estimated.Results:In diabetic rats, APME treatment significantly restored body weight (26.39%), blood glucose (32.39%), and insulin levels (73.95%) in comparison to diabetic control rats. In MIN6-β cells, APME potentiated insulin secretion in a dependent manner of glucose (3–16.7 mM) and extract (5–500 μg/mL) concentration. Insulin secretagogue effect was demonstrated in the presence of 3-isobutyl-1-methyl xanthine, glibenclamide, elevated extracellular calcium, and K+ depolarized media. Insulin release was reduced in the presence of nifedipine, ethylene glycol tetra acetic acid (calcium blocking agents), and diazoxide (potassium channel opener).Conclusion:The study suggests that APME antihyperglycemic activity might involve the insulin secretagogue effect by pancreatic beta cells physiological pathways via K+-ATP channel dependent and independently, along with an effect on Ca2+ channels.SUMMARY Abrus precatorius leaves methanolic extract (APME) showed a significant anti hyperglycemic and insulin secretagogue activities in streptozotocin induced diabetic rats. Also demonstrated a potent In vitro insulin secretagogue effect in mouse insulinoma beta cells (MIN6-β)APME treatment significantly restored body weight (26.39%), reduced blood glucose (32.39%) and enhanced circulatory insulin levels (73.95%) in diabetic ratsAPME demonstrated glucose and extract dose dependent insulin secretionInsulin secretagogue effect was demonstrated in the presence of 3-isobutyl-1-methyl xanthine, glibenclamide, elevated extracellular calcium and K+ depolarized media. Insulin release was reduced in the presence of nifedipine and ethylene glycol tetra acetic acid (Calcium blocking agents), diazoxide (potassium channel opener)The insulinotropic effect of APME involves a physiologic effect on K+-ATP channel and Ca2+ channels Abbreviations Used: ANOVA: Analysis of variance, CMC: Carboxy methyl cellulose, cAMP: Cyclic adenosine monophosphate, CaCl: Calcium chloride, AP: Abrus precatorius, APME: Abrus precatorius methanolic extract, DMEM: Dulbecco's modified Eagle's medium, DMSO: Dimethyl sulphoxide, EGTA: Ethylene glycol tetra acetic acid, FCS: Fetal calf serum, IBMX: 3-Isobutyl-1-methylxanthine, KCl: Potassium chloride, KRB: Kreb's Ringer buffer, MIN6: Mouse insulinoma cell line, MTT: 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-Diphenyltetrazolium Bromide, STZ: Streptozotocin.

KCl -Permeabilized Pancreatic Islets: An Experimental Model to Explore the Messenger Role of ATP in the Mechanism of Insulin Secretion.

Our previous work has demonstrated that islet depolarization with KCl opens connexin36 hemichannels in β-cells of mouse pancreatic islets allowing the exchange of small metabolites with the extracellular medium. In this study, the opening of these hemichannels has been further characterized in rat islets and INS–1 cells. Taking advantage of hemicannels’opening, the uptake of extracellular ATP and its effect on insulin release were investigated. 70 mM KCl stimulated light emission by luciferin in dispersed rat islets cells transduced with the fire-fly luciferase gene: it was suppressed by 20 mM glucose and 50 μM mefloquine, a specific connexin36 inhibitor. Extracellular ATP was taken up or released by islets depolarized with 70 mM KCl at 5 mM glucose, depending on the external ATP concentration. 1 mM ATP restored the loss of ATP induced by the depolarization itself. ATP concentrations above 5 mM increased islet ATP content and the ATP/ADP ratio. No ATP uptake occurred in non-depolarized or KCl-depolarized islets simultaneously incubated with 50 μM mefloquine or 20 mM glucose. Extracellular ATP potentiated the secretory response induced by 70 mM KCl at 5 mM glucose in perifused rat islets: 5 mM ATP triggered a second phase of insulin release after the initial peak triggered by KCl-depolarization itself; at 10 mM, it increased both the initial, KCl-dependent, peak and stimulated a greater second phase of secretion than at 5 mM. These stimulatory effects of extracellular ATP were almost completely suppressed by 50 μM mefloquine. The magnitude of the second phase of insulin release due to 5 mM extracellular ATP was decreased by addition of 5 mM ADP (extracellular ATP/ADP ratio = 1). ATP acts independently of KATP channels closure and its intracellular concentration and its ATP/ADP ratio seems to regulate the magnitude of both the first (triggering) and second (amplifying) phases of glucose-induced insulin secretion.

The skeleton in the closet: actin cytoskeletal remodeling in β-cell function.

Over the last few decades, biomedical research has considered not only the function of single cells but also the importance of the physical environment within a whole tissue, including cell-cell and cell-extracellular matrix interactions. Cytoskeleton organization and focal adhesions are crucial sensors for cells that enable them to rapidly communicate with the physical extracellular environment in response to extracellular stimuli, ensuring proper function and adaptation. The involvement of the microtubular-microfilamentous cytoskeleton in secretion mechanisms was proposed almost 50 years ago, since when the evolution of ever more sensitive and sophisticated methods in microscopy and in cell and molecular biology have led us to become aware of the importance of cytoskeleton remodeling for cell shape regulation and its crucial link with signaling pathways leading to β-cell function. Emerging evidence suggests that dysfunction of cytoskeletal components or extracellular matrix modification influences a number of disorders through potential actin cytoskeleton disruption that could be involved in the initiation of multiple cellular functions. Perturbation of β-cell actin cytoskeleton remodeling could arise secondarily to islet inflammation and fibrosis, possibly accounting in part for impaired β-cell function in type 2 diabetes. This review focuses on the role of actin remodeling in insulin secretion mechanisms and its close relationship with focal adhesions and myosin II.

Elucidation of genetic factors in diabetes based on studies of animal models

Although genetic factors are involved in the development of both type 1 (T1D) and type 2 (T2D) diabetes, a complete picture of those factors remains to be elucidated. T1D is an autoimmune disease in which the major histocompatibility complex (MHC) serves as a major genetic factor. By genetic analysis of an animal model of T1D, the Komeda diabetes-prone (KDP) rat, we identified the casitas B-lineage lymphoma b (Cblb) gene as another major genetic factor in this model. The Cblb mutation and MHC u haplotype determine autoimmune reaction and tissue specificity to pancreatic β cells, respectively. By mutation screening of the CBLB gene in Japanese patients with T1D, six missense mutations were found, among which Phe328Leu showed impaired suppression of T-cell activation. T2D is a metabolic disorder associated with insulin resistance and/or impaired insulin secretion. By genetic analysis of the spontaneously diabetic Torii (SDT) rat, an animal model of nonobese T2D, we identified several quantitative trait loci, among which a major locus, designated Dmsdt1, was involved in islet inflammation and fibrosis. Identification of the genes responsible should provide insight into the pathogenesis of diabetes. We recently established a novel model of obese T2D, the Zucker fatty diabetes mellitus (ZFDM) rat. In addition to severe insulin resistance and diminished insulin response to incretin, intrinsic fragility of islets in ZFDM rats is involved in the development of T2D. The ZFDM strain should be useful for studying the pathogenesis and pathophysiology of T2D, especially the mechanisms of incretin-induced insulin secretion and islet fragility.

Effect of Co-60 gamma irradiated chitosan and phosphorus fertilizer on growth, yield and trigonelline content of Trigonella foenum-graecum L.

Natural marine polysaccharides such as chitosan, sodium alginate and carrageenan; after depolymerization by different techniques, have shown promising plant growth promoting and other biological activities. Chitosan after irradiation with Co-60 gamma-rays have proven to be a miraculous plant growth promoter in different plants. Trigonelline; an anti-diabetic compound is obtained mainly from the seeds of Fenugreek (Trigonella foenum-graecum L.). Trigonelline counteracts diabetes mellitus (DM) through the operation of the mechanisms of insulin secretion, modulation of β-cell regeneration and stimulation of activity of glucose metabolism related enzymes. Considering the medicinal importance a pot experiment was conducted to investigate the effect of foliar application of Co-60 gamma irradiated chitosan (IC) with soil applied phosphorus supplement on growth, biochemical and quality attributes of fenugreek. Four concentrations of irradiated chitosan were used (0, 40, 80 and 120 mg L−1) individually as well as in combination with single dose of phosphorus 40 kg ha−1. Un-irradiated chitosan (UN) 40 mg L−1 and de-ionized water were used as control. IC significantly affected almost all the parameters including seed yield, trigonelline content and trigonelline yield. There were 10 treatments and among all, P40 + IC 40 mg L−1 proved to be the best, which increased total alkaloid content by 34.9%, seed yield by 125.4%, and trigonelline content by 17.8%. Contents of photosynthetic pigments and activity of nitrate reductase, carbonic anhydrase enzymes were also significantly enhanced.

Pseudoislet formation enhances gene expression, insulin secretion and cytoprotective mechanisms of clonal human insulin-secreting 1.1B4 cells.

We have studied the effects of cell communication on human beta cell function and resistance to cytotoxicity using the novel human insulin-secreting cell line 1.1B4 configured as monolayers and pseudoislets. Incubation with the incretin gut hormones GLP-1 and GIP caused dose-dependent stimulation of insulin secretion from 1.1B4 cell monolayers and pseudoislets. The secretory responses were 1.5-2.7-fold greater than monolayers. Cell viability (MTT), DNA damage (comet assay) and apoptosis (acridine orange/ethidium bromide staining) were investigated following 2-h exposure of 1.1B4 monolayers and pseudoislets to ninhydrin, H2O2, streptozotocin, glucose, palmitate or cocktails of proinflammatory cytokines. All agents tested decreased viability and increased DNA damage and apoptosis in both 1.1B4 monolayers and pseudoislets. However, pseudoislets exhibited significantly greater resistance to cytotoxicity (1.5-2.7-fold increases in LD50) and lower levels of DNA damage (1.3-3.4-fold differences in percentage tail DNA and olive tail moment) and apoptosis (1.3-1.5-fold difference) compared to monolayers. Measurement of gene expression by reverse-transcription, real-time PCR showed that genes involved with insulin secretion (INS, PDX1, PCSK1, PCSK2, GLP1R and GIPR), cell-cell communication (GJD2, GJA1 and CDH1) and antioxidant defence (SOD1, SOD2, GPX1 and CAT) were significantly upregulated in pseudoislets compared to monolayers, whilst the expression of proapoptotic genes (NOS2, MAPK8, MAPK10 and NFKB1) showed no significant differences. In summary, these data indicate cell-communication associated with three-dimensional islet architecture is important both for effective insulin secretion and for protection of human beta cells against cytotoxicity.

Pancreatic Changes in Nerve Growth Factor/TrkA Associated with Insulin Secretion in Cerebral Ischemia.

Regulation of blood glucose levels as a therapeutic strategy for cerebral ischemia plays an important role in suppressing neuronal damage. In particular, suppression of post-ischemic glucose intolerance improves cerebral ischemia. We have reported that cerebral ischemia induces glucose intolerance and an increase in plasma insulin levels. However, the mechanism of insulin secretion after cerebral ischemia is unclear. Nerve growth factor (NGF), a member of the neurotrophin family, has high affinity for tropomyosin-related kinase A (TrkA). NGF/TrkA signaling is associated with neuronal survival, differentiation, and function. Recently, NGF/TrkA signaling has been reported to be associated with insulin synthesis and secretion. In the present study, we evaluated the insulin content and expression of NGF/TrkA by immunofluorescence and Western blotting after middle cerebral artery occlusion (MCAO) as a cerebral ischemia model. At 6, 12, and 24 h after MCAO, insulin contents were increased in MCAO mice. The expression of NGF was increased at 6, 12, and 24 h, whereas the expression of TrkA tended to decrease in pancreas after MCAO. These results suggest that NGF/TrkA signaling is an important factor in cerebral ischemia-induced insulin synthesis and secretion in the pancreas.

Curcumin improves high glucose-induced INS-1 cell insulin resistance via activation of insulin signaling.

Curcumin is a yellow pigment isolated from Corcuma longan. This research investigates the improvement of curcumin on INS-1 cells with insulin resistance induced by high glucose. INS-1 cells were treated with high glucose (30 mmol L(-1)) for 48 h. Subsequently, the medium was replaced with curcumin for 24 h. Curcumin effectively increased insulin gene expression and glucose stimulated insulin secretion (GSIS) in a dose-dependent manner. Furthermore, the molecular mechanism of curcumin-induced insulin expression and secretion in high glucose-induced INS-1 cells was investigated in this study. Curcumin increased the expression of glucose transporter 2 (GLUT2) and phosphorylation of insulin receptor (IR), insulin receptor substrate-1 (IRS1), phosphatidylinositol-3-kinase (PI3K) and AKT in the INS-1 cells. Moreover, curcumin stimulation increased the expression of PDX-1 and GCK. This investigation suggests that curcumin prevented high glucose-reduced insulin expression and secretion through activation of the PI3K/Akt/GLUT2 pathway in INS-1 cells.

Glucose-Induced Cyclic-AMP Oscillations: Modeling Incretin Impact on Pancreatic Beta Cell Secretion

Pancreatic beta cells modulate insulin secretion via incretin-induced amplification pathways of cytosolic calcium (Ca2+)-induced exocytosis, which may be help compensate for reduced insulin sensitivity in Type II Diabetes Mellitus. One modulator is cAMP, which is activated by an incretin-triggered Gs protein cascade and sensitive to glucose levels. Interestingly, cAMP is found to be oscillatory, even in cases where Ca2+ is not, providing a possible mechanism for pulstile insulin secretion in the absence of Ca2+ oscillations. When Ca2+ and cAMP both oscillate, the two may be either in-phase or out-of-phase with one another. To understand the cAMP oscillations and their relationship to Ca2+ oscillations we use mathematical modeling. We model the cAMP production sensitivity to glucose metabolism with AMP sensitive adenylyl cyclases and degradation with Ca2+ sensitive phosphodiesterases. These modest additions to the Dual Oscillator Model for beta cells allow us to capture observed relationships between cAMP and Ca2+. This provides further evidence for metabolic oscillations in beta-cells that are affected by, but that do not require, oscillations in Ca2+.

Controlling insulin secretion: an exciting TASK.

The -cells of the pancreatic islets of Langerhans secrete insulin in response to metabolic fuels and neurohormonal inputs after a meal. The proper regulation of insulin secretion is critical for the maintenance of glucose homeostasis. An absolute or relative impairment of insulin secretion in the face of insulin resistance underlies the pathophysiology of type 2 diabetes. Accordingly, some of the key antidiabetes agents promote increased insulin secretion from islets, including the newer incretin-based therapeutics and the sulfonylurea class of drugs. The consensus mechanism for glucose-stimulated insulin secretion was elucidated almost 30 years ago and has remained largely unchanged since. A glucose-dependent increase in the intracellular ATP to ADP ratio closes ATP-sensitive potassium (KATP) ion channels (1, 2), themselves the target of the sulfonylurea drugs (3). This results in depolarization of the -cell membrane and the firing of action potentials, which allows the activation of voltage-gated Ca channels and a rise in intracellular Ca that triggers insulin release (4). It is well-recognized, however, that a diverse array of additional ion channels contribute to -cell excitability, and only recently have we begun to understand the roles and molecular identities of the many key channels that shape the electrical response in these cells. Although the outward flux of K mediated by KATP channels dominates the electrical conductance of the -cell membrane in the resting condition,holding themembranepotential of these cells near the equilibrium potential for K ( 70 mV), closure of KATP channels upon glucose stimulation is not in itself sufficient to depolarize the cell. For the -cell to depolarize, an inward depolarizing leak current is required to push the membrane potential toward a more positive equilibrium (or plateau, Figure 1) sufficient to allow the activation of the voltage-gated Na , Ca , and K channels that mediate regenerative action potential firing (5). This depolarizing force is likely mediated by channels that mainly allow the inward leak of Na (and possibly Ca ) such as the transient receptor potential (TRP) family of channels (6) and/or the muscarinic-activated nonselective Na -leak channel (NALNC) (7). Additional currents, such as those mediated by the smallconductance Ca -sensitive K (SK) channels (8) and hyperpolarization-activated cyclic nucleotide-gated (HCN) channels (9), contribute to the oscillatory nature of the electrical activity that characterizes -cell electrical function. Although required for membrane depolarization after glucose stimulation, these inward leak currents pose a problem of their own: the equilibrium potential for Na and Ca is much too positive ( 50 mV) to allow the rhythmic opening and closing of voltage-dependent ion channels required for action potential firing. What this means is that upon KATP channel closure in response to elevated glucose or sulfonylurea, the inward leak currents could depolarize the -cell too much, unless a counterbalancing outward leak is also present. In the present issue of Endocrinology, Dadi et al (10) present evidence to suggest that this key outward leak current is mediated by TASK-1, a member of the 2-poredomain K (K2P) channel family shown to be expressed at the mRNA level in human -cells only very recently (11, 12) and suggested previously as a target of inhaled anesthetics (13) and for the treatment of multiple sclerosis (14), T cell-mediated autoimmunity (15), and cardiac arrhythmia (16) among other conditions. Using cell-type–specific knockout mice and a selective TASK-1 inhibitor, the authors demonstrate that this channel mediates a back-

Islet NADPH oxidase activity modulates β-cell mass and endocrine function in rats with fructose-induced oxidative stress

BackgroundIslet NADPH oxidase activity is modulated by glucose and other insulin secretagogues and it might be part of the regulatory mechanism of insulin secretion. We studied its modulatory role of islet NADPH oxidase upon β-cell function in rats with fructose-induced oxidative stress. MethodsNormal rats were fed for 3weeks with a standard diet, a fructose-rich diet or both diets plus apocynin. We measured plasma glucose, insulin, triacylglycerol and lipid peroxidation levels and the homeostasis model assessment-insulin resistance (HOMA-IR) and HOMA-β indexes, and performed an oral glucose tolerance test. β-cell volume density and the number of islets per mm2 were determined by immunomorphometric analysis of the pancreas. Insulin secretion, glucose metabolism, glucokinase and NADPH oxidase activities were studied in islets isolated from each experimental group. ResultsFructose-fed rats had increased plasma triacylglycerol, insulin and lipid peroxidation levels associated with an insulin resistance state; the reactive higher secretion was unable to cope with the increased demand of insulin, leading to an impaired glucose tolerance. They also have a lower number of islets per area unit with a decreased β-cell volume density. All these alterations were prevented by blocking NADPH oxidase activity with apocynin. ConclusionFructose-induced changes are partly mediated by modulation of NADPH oxidase activity. General significanceThe metabolic dysfunctions and enhanced oxidative stress measured in fructose-fed rats resemble those recorded in human prediabetes; thus, successful strategies employed in this model could be later used to prevent the progression of this state towards type 2 diabetes in human beings.

Meal feeding improves oral glucose tolerance in male rats and causes adaptations in postprandial islet hormone secretion that are independent of plasma incretins or glycemia.

Meal-fed (MF) rats with access to food for only 4 consecutive hours during the light cycle learn to eat large meals to maintain energy balance. MF animals develop behavioral and endocrine changes that permit glucose tolerance despite increased meal size. We hypothesized that enhanced activity of the enteroinsular axis mediates glucose homeostasis during MF. Cohorts of rats were allocated to MF or ad libitum (AL) regimens for 2-4 wk. Insulin secretion and glucose tolerance were determined after oral carbohydrate and intraperitoneal (ip) and intravenous (iv) glucose. MF rats ate less than AL in the first week but maintained a comparable weight trajectory thereafter. MF rats had decreased glucose excursions after a liquid mixed meal (AUC: MF 75 ± 7, AL 461 ± 28 mmol·l⁻¹·min, P < 0.001), with left-shifted insulin secretion (AUC(0-15): MF 31.0 ± 4.9, AL 9.6 ± 4.4 pM·min, P < 0.02), which peaked before a significant rise in blood glucose. Both groups had comparable fasting glucagon levels, but postprandial responses were lower with MF. However, neither intestinal expression of proGIP and proglucagon mRNA nor plasma incretin levels differed between MF and AL groups. There were no differences in the insulin response to ip or iv glucose between MF and AL rats. These findings demonstrate that MF improves oral glucose tolerance and is associated with significant changes in postprandial islet hormone secretion. Because MF enhanced β-cell function during oral but not parenteral carbohydrate administration, and was not accounted for by changes in circulating incretins, these results support a neural mechanism of adaptive insulin secretion.

LIM-homeodomain transcription factor Isl-1 mediates kisspeptin's effect on insulin secretion in mice.

Kisspeptin and the G protein-coupled receptor 54 (GPR54) are highly abundant in the pancreas. In addition, circulating kisspeptin directly influences insulin secretion through GPR54. However, the mechanisms by which kisspeptin affects insulin release are unclear. The LIM-homeodomain transcription factor, Isl-1, is expressed in all pancreatic islet cells and is involved in regulating both islet development and insulin secretion. We therefore investigated potential interactions between kisspeptin and Isl-1. Our results demonstrate that Isl-1 and GPR54 are coexpressed in mouse pancreatic islet β-cells and NIT cells. Both in vitro and in vivo results demonstrate that kisspeptin-54 (KISS-54) inhibits Isl-1 expression and insulin secretion and both the in vivo and in vitro effects of KISS-54 on insulin gene expression and secretion are abolished when an Isl-1-inducible knockout model is used. Moreover, our results demonstrate that the direct action of KISS-54 on insulin secretion is mediated by Isl-1. Our results further show that KISS-54 influences Isl-1 expression and insulin secretion through the protein kinase C-ERK1/2 pathway. Conversely, insulin has a feedback loop via the Janus kinase-phosphatidylinositol 3-kinase pathway regulating kisspeptin expression and secretion. These findings are important in understanding mechanisms of insulin secretion and metabolism in diabetes.

A Novel Pathway for Regulation of Insulin Secretion by Fractalkine and CX3CR1 System (LB772)

Fractalkine (FKN) (CX3CL1) and its receptor CX3CR1 mediate cell‐to‐cell interactions in different tissues. Here, we demonstrate that the FKN/CX3CR1 system represents a novel regulatory mechanism for pancreatic islet beta cell function and insulin secretion. CX3CR1 KO mice exhibit glucose intolerance with normal insulin sensitivity, due to a marked beta cell defect in glucose and GLP1‐stimulated insulin secretion. The defect in insulin secretion was also observed in vitro in isolated islets from CX3CR1 KO mice. In vivo administration of FKN improved glucose tolerance with an increase in insulin secretion. In vitro treatment of islets with FKN increased intracellular Ca2+ level and potentiated insulin secretion. The KO islets exhibited reduced expression of a set of genes which are necessary for the fully functional, differentiated beta cell state, whereas, treatment of WT islets with FKN leads to increased expression of these genes. Lastly, expression of FKN in islets was decreased by aging and HFD/obesity, suggesting that decreased fractalkine/CX3CR1 signaling could be a mechanism underlying beta cell dysfunction in type 2 diabetes.

Nutrient regulation of insulin secretion and action

Pancreatic β-cell function is of critical importance in the regulation of fuel homoeostasis, and metabolic dysregulation is a hallmark of diabetes mellitus (DM). The β-cell is an intricately designed cell type that couples metabolism of dietary sources of carbohydrates, amino acids and lipids to insulin secretory mechanisms, such that insulin release occurs at appropriate times to ensure efficient nutrient uptake and storage by target tissues. However, chronic exposure to high nutrient concentrations results in altered metabolism that impacts negatively on insulin exocytosis, insulin action and may ultimately lead to development of DM. Reduced action of insulin in target tissues is associated with impairment of insulin signalling and contributes to insulin resistance (IR), a condition often associated with obesity and a major risk factor for DM. The altered metabolism of nutrients by insulin-sensitive target tissues (muscle, adipose tissue and liver) can result in high circulating levels of glucose and various lipids, which further impact on pancreatic β-cell function, IR and progression of the metabolic syndrome. Here, we have considered the role played by the major nutrient groups, carbohydrates, amino acids and lipids, in mediating β-cell insulin secretion, while also exploring the interplay between amino acids and insulin action in muscle. We also focus on the effects of altered lipid metabolism in adipose tissue and liver resulting from activation of inflammatory processes commonly observed in DM pathophysiology. The aim of this review is to describe commonalities and differences in metabolism related to insulin secretion and action, pertinent to the development of DM.