Physiological Regulation Of Insulin Secretion Research Articles

295-LB: Pancreatic Peroxisomes Are Required for Physiological Regulation of Insulin Secretion and Maintenance of Normal Glucose Tolerance in Male Mice

The regulation of glucose-stimulated insulin secretion (GSIS) is central to maintenance of glucose disposal. The role of lipid storage and lipid oxidation to regulate pancreatic islet β-cell insulin secretion is not completely understood. Peroxisomes are enriched in insulin-producing β-cells, but their contribution to β-cell function and maturity is lacking. Therefore, we generated a novel model of peroxisomal dysfunction to test the hypothesis that peroxisomal lipid metabolism is required to prevent a hyperlipidemic environment that interferes with β-cell function and maturity. Pex5 plays a critical role in importing the majority of peroxisomal proteins into the peroxisomal matrix; thus, deletion of the Pex5 gene creates a cellular environment with reductions in function of this organelle. In this study, we bred Pex5 mice on a C57BL/6J background containing ‘floxed’ alleles to Pdx1-cre mice to generate offspring with a pancreas-wide deficiency in Pex5 expression and abundance. Glucose tolerance was impaired by 12 weeks in male mice (n=20; AUC p<0.05). Surprisingly, GSIS was elevated in Pex5Pdx1-/- mice compared to littermate controls both in vivo (n=15; AUC p<0.05) and ex vivo in isolated islets as measured by perifusion analysis (n=12; AUC p<0.01). We further found that a subset of β-cells from Pex5Pdx1-/- mice expressed the de-differentiation marker Aldh1a3, while a subset of β-cells showed reductions in MafA and Nkx6.1 staining compared to controls. Perilipin-1 staining showed increased presence of lipid droplets in pancreatic tissue of Pex5Pdx1-/- mice versus controls. Thus, pancreatic deletion of peroxisomal function results in increased lipid storage in pancreatic tissue that is correlated with exaggerated insulin secretion. Collectively, these data are in support of a model whereby peroxisomal function supports β-cell insulin secretion and maintenance of the fully differentiated state. Disclosure S. Burke: None. S. Ghosh: None. D. Burk: None. R. Noland: None. J. Collier: None.

Membrane protease prostasin promotes insulin secretion by regulating the epidermal growth factor receptor pathway

Prostasin (PRSS8) is a serine protease that metabolizes and moderates the effect of specific substrates. Epidermal growth factor receptor (EGFR), which modulates insulin secretion and pancreatic β-cell proliferation, is regulated via proteolytic shedding by PRSS8. We first detected PRSS8 expression in β-cells of pancreatic islets of mice. To better understand the molecular processes involved in PRSS8-associated insulin secretion, pancreatic β-cell-specific PRSS8 knockout (βKO) and PRSS8-overexpressing (βTG) male mice were generated. We found that glucose intolerance and reduction in glucose-stimulated insulin secretion developed in βKO mice compared with the control subjects. A higher response to glucose was noted in islets retrieved from βTG mice. Erlotinib, a specific blocker of EGFR, blocks EGF- and glucose-stimulated secretion of insulin among MIN6 cells, and glucose improves EGF release from β-cells. After silencing PRSS8 in MIN6 cells, glucose-stimulated insulin secretion decreased, and EGFR signaling was impaired. Conversely, overexpression of PRSS8 in MIN6 cells induced higher concentrations of both basal and glucose-stimulated insulin secretion and increased phospho-EGFR concentrations. Furthermore, short-term exposure to glucose improved the concentration of endogenous PRSS8 in MIN6 cells through inhibition of intracellular degradation. These findings suggest that PRSS8 is involved in glucose-dependent physiological regulation of insulin secretion via the EGF–EGFR signaling pathway in pancreatic β-cells.

A critical role of the mechanosensor PIEZO1 in glucose-induced insulin secretion in pancreatic β-cells

Glucose-induced insulin secretion depends on β-cell electrical activity. Inhibition of ATP-regulated potassium (KATP) channels is a key event in this process. However, KATP channel closure alone is not sufficient to induce β-cell electrical activity; activation of a depolarizing membrane current is also required. Here we examine the role of the mechanosensor ion channel PIEZO1 in this process. Yoda1, a specific PIEZO1 agonist, activates a small membrane current and thereby triggers β-cell electrical activity with resultant stimulation of Ca2+-influx and insulin secretion. Conversely, the PIEZO1 antagonist GsMTx4 reduces glucose-induced Ca2+-signaling, electrical activity and insulin secretion. Yet, PIEZO1 expression is elevated in islets from human donors with type-2 diabetes (T2D) and a rodent T2D model (db/db mouse), in which insulin secretion is reduced. This paradox is resolved by our finding that PIEZO1 translocates from the plasmalemma into the nucleus (where it cannot influence the membrane potential of the β-cell) under experimental conditions emulating T2D (high glucose culture). β-cell-specific Piezo1-knockout mice show impaired glucose tolerance in vivo and reduced glucose-induced insulin secretion, β-cell electrical activity and Ca2+ elevation in vitro. These results implicate mechanotransduction and activation of PIEZO1, via intracellular accumulation of glucose metabolites, as an important physiological regulator of insulin secretion.

Preserving Insulin Secretion in Diabetes by Inhibiting VDAC1 Overexpression and Surface Translocation in β Cells

SummaryType 2 diabetes (T2D) develops after years of prediabetes during which high glucose (glucotoxicity) impairs insulin secretion. We report that the ATP-conducting mitochondrial outer membrane voltage-dependent anion channel-1 (VDAC1) is upregulated in islets from T2D and non-diabetic organ donors under glucotoxic conditions. This is caused by a glucotoxicity-induced transcriptional program, triggered during years of prediabetes with suboptimal blood glucose control. Metformin counteracts VDAC1 induction. VDAC1 overexpression causes its mistargeting to the plasma membrane of the insulin-secreting β cells with loss of the crucial metabolic coupling factor ATP. VDAC1 antibodies and inhibitors prevent ATP loss. Through direct inhibition of VDAC1 conductance, metformin, like specific VDAC1 inhibitors and antibodies, restores the impaired generation of ATP and glucose-stimulated insulin secretion in T2D islets. Treatment of db/db mice with VDAC1 inhibitor prevents hyperglycemia, and maintains normal glucose tolerance and physiological regulation of insulin secretion. Thus, β cell function is preserved by targeting the novel diabetes executer protein VDAC1.

RD Lawrence Lecture 2017 Incretins: the intelligent hormones in diabetes.

The incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) have attracted considerable scientific and clinical interest due largely to their insulin-releasing and glucose-lowering properties. Indeed, GLP-1-based therapies are now key treatment options for many people with diabetes worldwide. In contrast, GIP-based agents have yet to reach the clinic based primarily on the impaired insulinotropic action of GIP observed in people with diabetes. Nevertheless, GIP is a key physiological regulator of insulin secretion and stable forms of GIP show much promise in rodent models to alleviate diabetes-obesity. Recent studies suggest that GIP may have an important role to play in a combination therapeutic approach or bioengineered with other gut peptides. Moreover, recent experimental studies indicate that incretins also exert pleiotropic effects in regions of the brain associated with learning and memory, thereby supporting preclinical data demonstrating that incretin-based drugs improve cognitive function. This review article, based on the RD Lawrence Lecture presented at Diabetes UK Annual Professional Conference (2017), provides a brief overview of incretins with a major focus on GIP, the development of designer GIP analogues, and how these molecules can improve cognition. Thus, incretins can be considered as 'the intelligent hormones' and may hold the key to successfully treating the alarming rise in neurodegenerative disorders.

GLP-1 could improve the similarity of IPCs and pancreatic beta cells in cellular ultrastructure and function

Transplantation of functional insulin-producing cells (IPCs) provides a novel mode for insulin replacement, but is often accompanied by many undesirable side effects. Our previous studies suggested that IPCs could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells. To obtain a better method through which to acquire more similar IPCs, we compared the difference between IPCs of the GLP-1 group and IPCs of the non-GLP-1 group in the morphological features in cellular level and physiological function. The levels of insulin secretion were measured by ELISA. The insulin and glucagon-like peptide-1 (GLP-1) mRNA gene expression was determined by real-time quantitative PCR. The morphological features were detected by atomic force microscopy (AFM) and laser confocal scanning microscopy (LCSM). Intracellular Ca(2+) levels and Glucagon-like peptide-1 receptor (GLP-1R) levels were determined by flow cytometer (FCM). We found that IPCs of the GLP-1 group had bigger membrane particle size and average roughness (Ra ) than IPCs of the non-GLP-1 group but still smaller than normal human pancreatic beta cells. The physiology function of IPCs of the GLP-1 group were much closer to normal human pancreatic beta cells than IPCs of the non-GLP-1 group. GLP-1 could improve the similarity of IPCs from human adipose tissue-derived mesenchymal stem cells and pancreatic beta cells in cellular ultrastructure and function.

Insulin-producing cells could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells.

ObjectiveThe aim of this study was to compare the difference between insulin-producing cells (IPCs) and normal human pancreatic beta cells both in physiological function and morphological features in cellular level.MethodsThe levels of insulin secretion were measured by enzyme-linked immunosorbent assay. The insulin gene expression was determined by real-time quantitative polymerase chain reaction. The morphological features were detected by atomic force microscopy (AFM) and laser confocal scanning microscopy.ResultsIPCs and normal human pancreatic beta cells were similar to each other under the observation in AFM with the porous structure features in the cytoplasm. Both number of membrane particle size and average roughness of normal human beta cells were higher than those of IPCs.ConclusionsOur results firstly revealed that the cellular ultrastructure of IPCs was closer to that of normal human pancreatic beta cells, but they still could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells.

Nitric oxide (NO) — Production and regulation of insulin secretion in islets of freely fed and fasted mice

Production of nitric oxide through the action of nitric oxide synthase (NOS) has been detected in the islets of Langerhans. The inducible isoform of NOS (iNOS) is induced by cytokines and might contribute to the development of type-1 diabetes, while the constitutive isoform (cNOS) is thought to be implicated in the physiological regulation of insulin secretion. In the present study we have detected and quantified islet cNOS- and iNOS-derived NO production concomitant with measuring its influence on insulin secretion in the presence of different secretagogues: glucose, l-arginine, l-leucine and α-ketoisocaproic acid (KIC) both during fasting and freely fed conditions. In intact islets from freely fed mice both cNOS- and iNOS-activity was greatly increased by glucose (20mmol/l). Fasting induced islet iNOS activity at both physiological (7mmol/l) and high (20mmol/l) glucose concentrations. NOS blockade increased insulin secretion both during freely fed conditions and after fasting. l-arginine stimulated islet cNOS activity and did not affect islet iNOS activity. l-leucine or KIC, known to enter the TCA cycle without affecting glycolysis, did not affect either islet cNOS- or iNOS activity. Accordingly, insulin secretion stimulated by l-leucine or KIC was unaffected by addition of l-NAME both during feeding and fasting. We conclude that both high glucose concentrations and fasting increase islet total NO production (mostly iNOS derived) which inhibit insulin secretion. The insulin secretagogues l-leucine and KIC, which do not affect glycolysis, do not interfere with the islet NO–NOS system.

Polymorphism and Ligand Dependent Changes in Human Glucagon-Like Peptide-1 Receptor (GLP-1R) Function: Allosteric Rescue of Loss of Function Mutation

The glucagon-like peptide-1 receptor (GLP-1R) is a key physiological regulator of insulin secretion and a major therapeutic target for the treatment of type II diabetes. However, regulation of GLP-1R function is complex with multiple endogenous peptides that interact with the receptor, including full-length (1-37) and truncated (7-37) forms of GLP-1 that can exist in an amidated form (GLP-1(1-36)NH₂ and GLP-1(7-36)NH₂) and the related peptide oxyntomodulin. In addition, the GLP-1R possesses exogenous agonists, including exendin-4, and the allosteric modulator, compound 2 (6,7-dichloro-2-methylsulfonyl-3-tert-butylaminoquinoxaline). The complexity of this ligand-receptor system is further increased by the presence of several single nucleotide polymorphisms (SNPs) that are distributed across the receptor. We have investigated 10 GLP-1R SNPs, which were characterized in three physiologically relevant signaling pathways (cAMP accumulation, extracellular signal-regulated kinase 1/2 phosphorylation, and intracellular Ca²⁺ mobilization); ligand binding and cell surface receptor expression were also determined. We demonstrate both ligand- and pathway-specific effects for multiple SNPs, with the most dramatic effect observed for the Met¹⁴⁹ receptor variant. At the Met¹⁴⁹ variant, there was selective loss of peptide-induced responses across all pathways examined, but preservation of response to the small molecule compound 2. In contrast, at the Cys³³³ variant, peptide responses were preserved but there was attenuated response to compound 2. Strikingly, the loss of peptide function at the Met¹⁴⁹ receptor variant could be allosterically rescued by compound 2, providing proof-of-principle evidence that allosteric drugs could be used to treat patients with this loss of function variant.

CGMP-independent anti-apoptotic effect of nitric oxide on thapsigargin-induced apoptosis in the pancreatic β-cell line INS-1

Aims Low concentrations of nitric oxide (NO) produced by constitutive NO synthase (cNOS) in pancreatic β-cells have been suggested to be a physiological regulator of insulin secretion. In contrast, excessive NO produced by inducible NO synthase is known to mediate β-cell apoptosis. The aim of the present study was to investigate the effect of low concentrations of NO on β-cell apoptosis. Main methods Apoptosis of the pancreatic β-cell line INS-1 was quantitatively determined by Annexin V flow cytometry. Key findings The 24-h incubation with 1 mM DETA/NO, a long half-life NO donor, induced β-cell apoptosis, which was insensitive to the soluble guanylate cyclase (sGC) inhibitor ODQ. In contrast, DETA/NO at lower concentrations until 300 μM concentration-dependently decreased the apoptosis induced by thapsigargin, an inhibitor of endoplasmic reticulum Ca 2+-ATPase. ODQ did not affect the anti-apoptotic effect of DETA/NO. Moreover, neither the cGMP analogue 8-Br-cGMP nor the sGC activator YC-1 mimicked the anti-apoptotic effect of DETA/NO. Significance These results suggest that low levels of NO protect β-cells from thapsigargin-induced apoptosis in a cGMP-independent manner.

The effect of dietary taurine supplementation on plasma and urinary free amino acid concentrations in diabetic rats.

Taurine (2-aminoethanesulfonic acid) is a normal constituent of the human diet and is found in animal food sources. Taurine has many biological functions including bile acid conjugation, osmoregulation, membrane protection, antioxidant defense, and regulation of cellular calcium homeostasis1–3. It has been suggested that taurine might serve as a physiological regulator of insulin secretion and plasma glucose levels4. It has been reported that taurine acts as a potent antioxidant and exhibits anti-diabetic activity in streptozotocin-induced diabetes mellitus5–6. Recently an immunohistochemical study showed that taurine protects pancreatic β-cells of rats against destruction by streptozotocin injection in a dose-dependent way7. Also, it has been reported that the plasma and platelet taurine content are reduced in subjects with insulin-dependent diabetes mellitus, although this defect can be reversed by taurine supplementation8. Therefore, the purpose of this study was to investigate the effect of dietary taurine supplementation on the plasma and urinary free amino acid content in diabetic rats.KeywordsControl DietAmino Acid ConcentrationAtherogenic IndexFree Amino Acid ConcentrationAnimal Food SourceThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

A Functional Link between Glucokinase Binding to Insulin Granules and Conformational Alterations in Response to Glucose and Insulin

Glucokinase (GK) activity is essential for the physiological regulation of insulin secretion by glucose. Because the enzyme exerts nearly total control over glucose metabolism in the beta-cell, even small changes in GK activity exert effects on glucose-stimulated insulin secretion and, consequently, the blood glucose concentration. Using quantitative imaging of multicolor fluorescent proteins fused to GK, we found that the association of GK with insulin granules is regulated by glucose in the beta-cell. Glucose stimulation increased the rate of fluorescence recovery after photobleaching of GK to insulin granules, indicating that GK is released into the cytoplasm after glucose stimulation. Changes in fluorescence resonance energy transfer between two different fluorescent protein variants inserted on opposing ends of GK were observed after glucose stimulation and correlated with increased enzyme activity. Furthermore, glucose-stimulated changes in GK regulation were blocked by two inhibitors of insulin secretion. Insulin treatment restored GK regulation in inhibited cells and stimulated GK translocation and activation by itself. Together, these data support a model for post-translational regulation of GK whereby insulin regulates both the association of GK with secretory granules and the activity of the enzyme within the pancreatic beta-cell.

Pancreatic Duodenal Homeobox (PDX-1) in health and disease.

Insulin is expressed exclusively in the adult beta-cells of the islets of Langerhans. Pancreatic Duodenum Homeobox-1 (PDX-1) is a major regulator of transcription in these cells. It transactivates the insulin gene by binding to a specific DNA motif in its promoter region. Glucose, the main physiological regulator of insulin secretion, also regulates insulin gene transcription through PDX-1. While acute exposure to high glucose concentrations causes an increase in PDX-1 binding, and consequently in insulin mRNA levels, chronic hyperglycemia (toxic to the beta-cell) leads to a decrease in PDX-1 and insulin levels. PDX-1 is absolutely required for pancreas development. In view of the selective expression in adult beta-cells, pancreatic agenesis in both the pdx-1 null mouse and a human carrying a homozygous mutation of PDX-1 was an unexpected and remarkable finding. The homozygous clinical phenotype was neonatal diabetes mellitus (DM) and exocrine insufficiency. Heterozygosity for PDX-1 mutations was found in some individuals with a newly characterized subtype of maturity-onset diabetes of the young (MODY4) and in others with type 2 DM. This review underlines the unique role of PDX-1 in maintaining adult beta-cell-specific functions in normal and disease-related states.

Are 5-Hydroxytryptamine-preloaded β-Cells an Appropriate Physiologic Model System for Establishing That Insulin Stimulates Insulin Secretion?

The release and oxidation of 5-hydroxytryptamine from 5-hydroxytryptamine-preloaded beta-cells has been used as a surrogate marker for insulin secretion. Findings made using this methodology have been used to support the concept that insulin stimulates its own release. In the present studies, the effects of 5-hydroxytryptamine on stimulated insulin secretion from isolated perifused rat islets was determined. When added together with stimulatory glucose, 5-hydroxytryptamine (0.5 mm) significantly reduced both phases of 8 mm glucose-induced secretion and reduced the first phase of 15 mm glucose-induced release by 60% without any effect on sustained insulin release rates. Preloading of beta-cells with 0.5 mm 5-hydroxytryptamine for 3 h resulted in a more severe impairment of 15 mm glucose-induced secretion. First and second phase release rates were reduced by 70 and 55%, respectively. In addition, this pretreatment protocol also abolished 200 microm tolbutamide-induced insulin secretion from perifused islets. These findings confirm that 5-hydroxytryptamine is a powerful inhibitor of stimulated insulin secretion. The responses of 5-hydroxytryptamine-preloaded beta-cells may not accurately reflect the biochemical events occurring during the physiologic regulation of insulin secretion. The suggestion that insulin stimulates its own secretion based exclusively on amperometric measurements should be reconsidered.

Characterization of Low-Affinity Binding Sites for Glibenclamide on the Kir6.2 Subunit of the β-Cell KATPChannel

The ATP-sensitive K+channel, an octameric complex of two structurally unrelated types of subunits, SUR1 and Kir6.2, plays a central role in the physiological regulation of insulin secretion. The sulfonylurea glibenclamide, which trigger insulin secretion by blocking the ATP-sensitive K+channel, interacts with both high and low affinity binding sites present on β-cells. The high affinity binding site has been localized on SUR1 but the molecular nature of the low affinity site is still uncertain. In this study, we analyzed the pharmacology of glibenclamide in a transformed COS-7 cell line expressing the rat Kir6.2 cDNA and compared with that of the MIN6 β cell line expressing natively both the Kir6.2 and the SUR1 subunits. Binding studies and Scatchard analysis revealed the presence of a single class of low affinity binding sites for glibenclamide on the COS/Kir6.2 cells with characteristics similar to that observed for the low affinity site of the MIN6 β cells.

Signal transduction in pancreatic beta-cells: regulation of insulin secretion by information flow in the phospholipase C/protein kinase C pathway.

The physiologic regulation of glucose-induced insulin secretion is dependent upon the activation of information flow in the phospholipase C (PLC)/protein kinase C (PKC) signal transduction system. In both rat and human pancreatic beta-cells, glucose has several time-dependent effects on secretory responsiveness including the regulation of biphasic insulin secretion, time-dependent potentiation and time-dependent suppression. PLC/PKC activation has been implicated in all three response patterns. Islets of Langerhans contain the three major PLC isozyme classes (beta1, gamma1 and delta1) and the available evidence suggests that one class is activated by fuel secretagogues and another by neurohumoral agonists. The expression and activation of PLC is labile. When rat islet are cultured for short periods, the content and activation of PLC in response to glucose decreases and this biochemical defect in signal transduction is paralleled by significant reductions in glucose-induced insulin secretion. Similar defects are observed when human islets are cultured as well. Mouse islet responses to glucose stimulation differ in several major and dramatic ways from rat and human islet responses. When stimulated by 15mM glucose, mouse islets fail to develop a rising second phase secretory response, they fail to exhibit either time-dependent potentiation or time-dependent suppression to the hexose. Biphasic insulin secretion can be evoked and time-dependent potentiation can be induced in mouse islets by carbachol, an agonist that activates an isozyme of PLC distinct from that activated by glucose. These divergent response patterns are attributable to the underexpression in mouse islets, when compared to rat islets, of a fuel-sensitive PLC isozyme. When taken in their entirety, the experimental evidence suggests that the activation of PLC is an essential component in the physiologic regulation of insulin secretion and that disordered activation of the enzyme culminates in disordered insulin release.

Regulation of Ca2+ transport by insulin and taurine. Interaction at the level of the Na(+)-Ca2+ exchanger.

In 1983 Lampson et al. 16 and Tokunaga et al. 31 introduced the concept that an interaction must exist between the actions of taurine and those of insulin. The initial studies examining this question focused on the metabolic actions of insulin. Tokunaga et al. 31 found that taurine suppressed insulin secretion in nondiabetic pancreatic islets, suggesting that taurine might serve as a physiological regulator of insulin secretion and plasma glucose levels. They also found that taurine levels increased significantly in diabetic islets, implying that taurine might contribute to the state of hypoinsulinemia and hyperglycemia. Lampson et al. 16 also concluded that taurine influenced plasma glucose levels, but their studies linked the hypoglycemic actions of taurine to a potentiation of insulin-mediated glucose uptake. Lampson et al. 16 also reported that taurine promoted several other insulin sensitive reactions, implying that taurine either facilitated the interaction of insulin with its receptor or increased the production of a putative second messenger of insulin. Subsequent studies revealed that taurine could interact directly with the insulin receptor15, 20. This interaction appeared to be quite specific since insulin was capable of displacing taurine from its binding sites20. Based on these data, Maturo and Kulakowski20 suggested that taurine might exhibit insulin-mimetic activity. However, this notion is inconsistent with a recent observation by Schaffer et al. 30 that insulin, but not taurine, could stimulate Na+-Ca2+ exchanger activity. Yet, the studies by Schaffer et al. 30 did not address the possibility that taurine might function as a general modulator of insulin action.

Clinical profile of glimepiride

In order to achieve appropriate blood glucose control, the treatment of non-insulin dependent (NIDDM) Type II diabetes usually starts with diet and exercise. If this still results in insufficient metabolic control, oral hypoglycaemic drugs or insulin are added to the non-pharmacological measures. Sulphonylureas have been used successfully as oral hypoglycaemic agents since the 1950s but there are aspects where medication could be better adjusted to the patients' needs. Preclinical investigations on animals and in vitro studies with glimepiride (HOE490), a new sulphonylurea, suggested some benefit over sulphonylureas currently available, including lower dosage, rapid onset and long duration of action, lower insulin and C-peptide levels, possibly due to less stimulation of insulin secretion and more pronounced extrapancreatic effects. The clinical relevance of these findings were studied in clinical trials. 19 phase II and 4 phase III clinical studies, in a total of about 3750 Type II diabetic patients, established efficacy and safety of glimepiride in comparison to placebo and glibenclamide and showed its therapeutic value. 1 mg per day induced a marked blood glucose reduction (FPG 2.4 mmol/1; HbA lc 1.2%) which could be enhanced by increasing the dose to the maximum effective 4 and 8 mg daily. In patients, glimepiride had a more rapid onset of action than glibenclamide, with a long duration of action. Glimepiride achieved metabolic control with the lowest dose (1-8 mg daily) of all the sulphonylureas. In addition, it maintained a more physiological regulation of insulin secretion than glibenclamide during physical exercise, suggesting that there may be less risk of hypoglycaemia with glimepiride. Large phase III studies were designed to characterise the product under conditions which were to be as close as possible to every-day life oral therapy of Type II diabetes. These long-term, glibenclamide-controlled studies showed that equivalent metabolic control was achieved with a dose range of 1-8 mg glimepiride given once daily and 2.5-20 mg glibenclamide daily (given as divided dose at the higher dose levels). This equivalent metabolic control was achieved with lower insulin concentrations (median difference: −0.92 μU/ml; P = 0.04) and C-peptide (median difference: −0.14 ng/ml; P = 0.03) with glimepiride. Glimepiride was well tolerated and fewer episodes of hypoglycaemia were observed in the glimepiride group than in the glibenclamide group. In conclusion, glimepiride showed a number of improvements over currently available sulphonylureas that may provide clinical benefit to patients with NIDDM.

Correlation between Ca(2+)-ATPase activity of rat islet cells and insulin secretion.

Using medium with a low ionic strength, a low concentration of Ca2+ and Mg2+ and devoid of K+, we have measured Ca(2+)-ATPase activity in the homogenates of rat islets preincubated for 3 min with several hormones in the presence of 3.3 mmol glucose/l. Insulin secretion was also measured in islets incubated for 5 min under identical experimental conditions. Islets preincubated with glucose (3.3 mmol/l) and glucagon (1.4 mumol/l) plus theophylline (10 mmol/l), ACTH (0.11 nmol/l), bovine GH (0.46 mumol/l), prolactin (0.2 mumol/l) or tri-iodothyronine (1.0 nmol/l) have significantly lower Ca(2+)-ATPase activity than those preincubated with only 3.3 mmol glucose/l. All these hormones increased the release of insulin significantly. Dexamethasone (0.1 mumol/l) and somatostatin (1.2 mumol/l) enhanced the Ca(2+)-ATPase activity while adrenaline (10 mumol/l) did not produce any significant effect on the activity of the enzyme. These hormones decreased the release of insulin significantly. These results demonstrated that islet Ca(2+)-ATPase activity was modulated by the hormones tested. Their inhibitory or enhancing effect seemed to be related to their effect on insulin secretion; i.e. those which stimulated the secretion of insulin inhibited the activity of the enzyme and vice versa. Hence, their effect on insulin secretion may be due, in part, to their effect on enzyme activity and consequently on the concentration of cytosolic Ca2+. These results reinforce the assumption that Ca(2+)-ATPase activity participates in the physiological regulation of insulin secretion, being one of the cellular targets for several agents which affect this process.

Functional receptors for the insulinotropic hormone glucagon-like peptide-I(7–37) on a somatostatin secreting cell line

Glucagon-like peptide-I (7–37) [(GLP-I(7–37)] is an instestinal peptide hormone that has potent insulinotropic activities in vivo in response to oral nutrients. In the isolated perfused pancreas, and in vitro in cultured B cells, GLP-I(7–37) receptor binding and GLP-I(7–37)-induced cAMP generation and harmone secretion was studied using cell lines producing insulin/B cell (βTC-1), glucagon/A cell (INR1G9) and somatostatin/D cell (RIN 1027-B2). [ 125I]GLP-I(7–37) bound specifically to both B and D cells but not to A cells. GLP-I(7–37) induced cAMP-formation in B and D cells with a maximum response at 10 nmol/l(B cells) or at 100 nmol/l (D cells). Insulin secretion from perifused B cells was stimulated by GLP-I(7–37) (maximum at 10 nmol/l) and 10 nmol/l GLP-I(7–37) released somatostatin from perifused D cells, GLP-I(7–37) did not influence cAMP or glucagon secretion from A cells. These data indicate that pancreatic B and D cells, but not the A cells are influenced directly by GLP-I(7–37) via binding to specific receptors. Our findings support a model of physiologic regulation of insulin secretion whereby GLP-I(7–37) released from the intestine in response to oral nutrients potently stimulates insulin secretion via an endocrine mechanism that in turn may be dampened by a feed-back suppression by the release of somatostatin. In addition, suppression of the secretion of glucagon, a hormone whose actions are counter-regulatory to those of insulin, may occur by paracrine mechanisms involving GLP-I(7–37)-mediated stimulation of both insulin and somatostatin secretion.