Glucagon-like Peptide-1 Signaling Research Articles

New ways in which GLP-1 can regulate glucose homeostasis

Glucagon-like peptide-1 (GLP-1) has a diverse set of peripheral actions which all serve to promote enhanced glucose tolerance, and for this reason it has become the basis for new treatments for type 2 diabetes. In this issue of the JCI, Knauf et al. provide clear evidence that GLP-1 signaling in the CNS is also linked to the control of peripheral glucose homeostasis by inhibiting non-insulin-mediated glucose uptake by muscle and increasing insulin secretion from the pancreas. The authors' work points to an important need to integrate diverse GLP-1 signaling actions and peripheral GLP-1 function in order to better understand both normal and abnormal glucose homeostasis.

Glucagon-like peptide 1 (GLP-1) and metabolic diseases

Glucagon-like peptide-1 (GLP-1) is a gastrointestinal hormone, mainly secreted after meals, which enhances glucose-induced insulin secretion and induces satiety. It has been reported that GLP-1 levels after a mixed meal and after an oral glucose load are reduced in patients with Type 2 diabetes. The reduction of oral glucose-stimulated active GLP-1 levels in patients with Type 2 diabetes has also been observed during euglycemic iperinsulinemic clamp. The reduction of post-prandial circulating active GLP-1 in Type 2 diabetic subjects, as a consequence of chronic hyperglycemia, could contribute to the reduction of early post-prandial insulin secretion; in fact, the administration of GLP-1 receptor antagonists to healthy volunteers elicits both an impairment of meal-induced insulin secretion and an increase of post-prandial glycemia similar to that observed in Type 2 diabetes. GLP-1 is rapidly inactivated by dipeptidyl peptidase IV (DPP-IV), an enzyme produced by endothelial cells in different districts and that circulates in plasma. It is still not clear whether the reduction of mealor oral-glucose stimulated GLP-1 levels in Type 2 diabetic patients is due to impairment of secretion, increase of degradation, or both. The major limitation of using GLP-1 to treat diabetic patients is the short half-life of the native compound. There are now several compounds in various stages of pre-clinical or clinical development for the treatment of Type 2 diabetes that utilize the GLP-1 signaling pathway; these include GLP-1 receptor agonists with extended half-lives, and inhibitors of DPP-IV that increase circulating levels of endogenous, intact and bioactive GLP-1.

Utilizing the GLP-1 signaling system to treat diabetes: Sorting through the pharmacologic approaches

Glucagon-like peptide-1 (GLP-1) is an intestinal hormone that promotes glucose homeostasis through the regulation of insulin and glucagon secretion, gastric emptying, and food intake. This spectrum of effects makes GLP-1 an attractive candidate for drug development. However, because GLP-1 is a small peptide with rapid metabolism in the circulation, its usefulness to treat patients is limited. However, GLP-1 mimetics that are resistant to degradation have been developed and are effective in lowering blood glucose in diabetic patients. A second strategy for harnessing GLP-1 therapeutically is to inhibit the metabolism of endogenous GLP-1; several orally available compounds are in clinical trials. These two new classes of drugs both enhance GLP-1 signaling but differ in several key characteristics that may lead to distinct roles in the treatment of diabetic patients.

Alpha cell function in health and disease: influence of glucagon-like peptide-1

Although there is abundant evidence that hyperglucagonaemia plays a key role in the development of hyperglycaemia in type 2 diabetes, efforts to understand and correct this abnormality have been overshadowed by the emphasis on insulin secretion and action. However, recognition that the incretin hormone glucagon-like peptide-1 (GLP-1) exerts opposing effects on glucagon and insulin secretion has revived interest in glucagon, the neglected partner of insulin, in the bihormonal hypothesis. In healthy subjects, glucagon secretion is regulated by a variety of nutrient, neural and hormonal factors, the most important of which is glucose. The defect in alpha cell function that occurs in type 2 diabetes reflects impaired glucose sensing. GLP-1 inhibits glucagon secretion in vitro and in vivo in experimental animals, and suppresses glucagon release in a glucose-dependent manner in healthy subjects. This effect is also evident in diabetic patients, but GLP-1 does not inhibit glucagon release in response to hypoglycaemia, and may even enhance it. Early clinical studies with agents acting through GLP-1 signalling mechanisms (e.g. exenatide, liraglutide and vildagliptin) suggest that GLP-1 can improve alpha cell glucose sensing in patients with type 2 diabetes. Therapeutic approaches based around GLP-1 have the potential to improve both alpha cell and beta cell function, and could be of benefit in patients with a broad range of metabolic disorders.

Glucagon‐like peptide‐1, corticotropin‐releasing hormone, and hypothalamic neuronal histamine interact in the leptin‐signaling pathway to regulate feeding behavior

Glucagon-like peptide-1 (GLP-1), corticotropin-releasing hormone (CRH), and hypothalamic neuronal histamine suppress food intake, a target of leptin action in the brain. This study examined the interactions of GLP-1, CRH, and histamine downstream from the leptin-signaling pathway in regulating feeding behavior. Infusion of GLP-1 into the third cerebral ventricle (i3vt) at a dose of 1 mug significantly decreased the initial 1 h cumulative food intake in rats as compared with phosphate-buffered saline (PBS) controls. The GLP-1-induced suppression of feeding was partially attenuated by intraperitoneal pretreatment with alpha-fluoromethylhistidine (FMH), a specific suicide inhibitor of histidine decarboxylase, which depletes hypothalamic neuronal histamine. Pretreatment with alpha-helical CRH (10 microg/rat, i3vt), a nonselective CRH antagonist, abolished the GLP-1-induced suppression of feeding completely. I3vt infusion of GLP-1 increased the CRH content and histamine turnover assessed using the pargyline-induced accumulation of tele-methyl histamine (t-MH), a major metabolite of neuronal histamine, in the hypothalamus. The central infusion of CRH also induced the increase of histamine turnover and CRH receptor type 1 was localized on the cell body of histamine neuron. Pretreatment with exendin(9-39), a GLP-1 receptor antagonist, attenuated the leptin-induced increase in CRH content of the hypothalamus. Finally, i3vt infusion of leptin also increased histamine turnover in the hypothalamus. Pretreatment with exendin(9-39), alpha-helical CRH or both antagonists attenuated the leptin-induced responses of t-MH levels in the hypothalamus. These results suggest that CRH or hypothalamic neuronal histamine mediates the GLP-1-induced suppression of feeding behavior, that CRH mediates GLP-1 signaling to neuronal histamine and that a functional link from GLP-1 to neuronal histamine via CRH constitutes the leptin-signaling pathway regulating feeding behavior.

Distinct Effects of Glucose-Dependent Insulinotropic Polypeptide and Glucagon-Like Peptide-1 on Insulin Secretion and Gut Motility

Glucose-induced insulin secretion from pancreatic beta-cells depends critically on ATP-sensitive K(+) channel (K(ATP) channel) activity, but it is not known whether K(ATP) channels are involved in the potentiation of insulin secretion by glucose-dependent insulinotropic polypeptide (GIP). In mice lacking K(ATP) channels (Kir6.2(-/-) mice), we found that pretreatment with GIP in vivo failed to blunt the rise in blood glucose levels after oral glucose load. In Kir6.2(-/-) mice, potentiation of insulin secretion by GIP in vivo was markedly attenuated, indicating that K(ATP) channels are essential in the insulinotropic effect of GIP. In contrast, pretreatment with glucagon-like peptide-1 (GLP-1) in Kir6.2(-/-) mice potentiated insulin secretion and blunted the rise in blood glucose levels. We also found that GLP-1 inhibited gut motility whereas GIP did not. Perfusion experiments of Kir6.2(-/-) mice revealed severely impaired potentiation of insulin secretion by 1 nmol/l GIP and substantial potentiation by 1 nmol/l GLP-1. Although both GIP and GLP-1 increase the intracellular cAMP concentration and potentiate insulin secretion, these results demonstrate that the GLP-1 and GIP signaling pathways involve the K(ATP) channel differently.

The Recombinant Rat Glucagon-Like Peptide-1 Receptor, Expressed in an α-Cell Line, Is Coupled to Adenylyl Cyclase Activation and Intracellular Calcium Release

The glucagon-like peptide-1 (GLP-1) receptor is expressed on alpha-cells, though its functional significance is unknown. The endogenous beta-cell GLP-1 receptor is coupled to adenylyl cyclase, cell depolarization, activation of voltage-dependent Ca2+ channels (VDCC) and extracellular Ca2+ influx (Lu et al., 1993 b). In contrast, the signaling pathways of the GLP-1 receptor in alpha-cells are poorly understood. To determine the signaling mechanisms of the alpha-cell GLP-1 receptor, we established a stable pancreatic islet alpha-cell line expressing the recombinant rat GLP-1 receptor (INR1-SF2), using INRl-G9 cells. These INRl-G9 cells do not express endogenous GLP-1 receptor. In INR1-SF2 cells, GLP-1 bound to the recombinant receptor (Kd = 0.9 nM) and increased cAMP (ED50 = 0.6 nM). GLP-1 increased the free cytosolic Ca2+ ([Ca2+]i) (ED50 = 50 nM) by release from intracellular stores, but did not affect INR1-SF2 cell phosphoinositol turnover. Despite expressing VDCC, the INR1-SF2 cells were not depolarized by GLP-1, even in the presence of glucose. This contrasts with the depolarizing action of GLP-1 in beta-cells in the presence of glucose (Lu et al., 1993 b). This study establishes that a single GLP-1 receptor species can mediate the effects of GLP-1 through multiple signaling pathways, including the adenylyl cyclase system and intracellular Ca2+ release, in an alpha-cell type. Furthermore, since GLP-1 is unable to cause cellular depolarization or activate VDCC in INR1-SF2 cells, these data suggest that glucose-induced membrane depolarization may be crucial for GLP-1 to further activate VDCC and potentiate glucose-stimulated insulin release in beta-cells. Finally this study describes a cell line that can be used as a model system for evaluation of GLP-1 signaling in alpha-cells.

Glucagon-like peptide 1: evolution of an incretin into a treatment for diabetes.

Glucagon-like peptide 1 (GLP-1) is a product of proglucagon that is secreted by specialized intestinal endocrine cells after meals. GLP-1 is insulinotropic and plays a role in the incretin effect, the augmented insulin response observed when glucose is absorbed through the gut. GLP-1 also appears to regulate a number of processes that reduce fluctuations in blood glucose, such as gastric emptying, glucagon secretion, food intake, and possibly glucose production and glucose uptake. These effects, in addition to the stimulation of insulin secretion, suggest a broad role for GLP-1 as a mediator of postprandial glucose homeostasis. Consistent with this role, the most prominent effect of experimental blockade of GLP-1 signaling is an increase in blood glucose. Recent data also suggest that GLP-1 is involved in the regulation of beta-cell mass. Whereas other insulinotropic gastrointestinal hormones are relatively ineffective in stimulating insulin secretion in persons with type 2 diabetes, GLP-1 retains this action and is very effective in lowering blood glucose levels in these patients. There are currently a number of products in development that utilize the GLP-1-signaling system as a mechanism for the treatment of diabetes. These compounds, GLP-1 receptor agonists and agents that retard the metabolism of native GLP-1, have shown promising results in clinical trials. The application of GLP-1 to clinical use fulfills a long-standing interest in adapting endogenous insulinotropic hormones to the treatment of diabetes.

Glucagon-like peptide-1 regulates proliferation and apoptosis via activation of protein kinase B in pancreatic INS-1 beta cells.

The incretin hormone glucagon-like peptide-1 augments islet cell mass in vivo by increasing proliferation and decreasing apoptosis of the beta cells. However, the signalling pathways that mediate these effects are mostly unknown. Using a clonal rat pancreatic beta cell line (INS-1), we examined the role of protein kinase B in mediating beta-cell growth and survival stimulated by glucagon-like peptide-1. Immunoblot analysis was used to detect active (phospho-) and total protein kinase B. Proliferation was assessed using (3)H-thymidine incorporation, while apoptosis was quantitated using 4'-6-diamidino-2-phenylindole staining and APO percentage apoptosis assay. Kinase-dead and wild-type protein kinase B was introduced into cells using adenoviral vectors. Glucagon-like peptide-1 rapidly activated protein kinase B in INS-1 cells (by 2.7+/-0.7-fold, p<0.05). This effect was completely abrogated by inhibition, with wortmannin, of the upstream activator of protein kinase B, phosphatidylinositol-3-kinase. Glucagon-like peptide-1 also stimulated INS-1 cell proliferation in a dose-dependent manner (by 1.8+/-0.5-fold at 10(-7) mol/l, p<0.01), and inhibited staurosporine-induced apoptosis (by 69+/-12%, p<0.05). Both of these effects were also prevented by wortmannin treatment. Ablation of protein kinase B by adenovirus-mediated overexpression of the kinase-dead form of protein kinase Balpha prevented protein kinase B phosphorylation and completely abrogated both cellular proliferation ( p<0.05) and protection from drug-induced cellular death ( p<0.01) induced by glucagon-like peptide-1. These results identify protein kinase B as an essential mediator linking the glucagon-like peptide-1 signal to the intracellular machinery that modulates beta-cell growth and survival.

Exendin-4, a GLP-1 Receptor Agonist, Interacts with Proteins and Their Products of Digestion to Suppress Food Intake in Rats

This study investigated the hypotheses that dietary proteins suppress food intake partly through the glucagon-like peptide-1 (GLP-1) signaling pathway, and that this effect is mediated by products of protein digestion. The GLP-1 receptor agonist, Exendin-4 (Ex-4) (0.5 micro g/rat), was given intraperitoneally to male Wistar rats, and food intake was measured when Ex-4 was given alone or with preloads of intact whey and casein proteins, their hydrolysates and amino acid mixtures (0.5 g x 4 mL(-1) x rat(-1)). Both Ex-4 and the preloads suppressed food intake (P < 0.05), but the effect of Ex-4 on food intake was reduced when coadministered with the preloads (P < 0.05). Because the effect of Ex-4 was reduced by the protein hydrolysates and by the amino acid preloads, the results support a role for the end products of protein digestion and GLP-1 release in the suppression of food intake in response to protein ingestion. We concluded that the GLP-1 signaling pathway, activated by the release of products of protein digestion, is another mechanism accounting for the reduction of food intake after protein ingestion.

Glucagon-like peptide 1 receptor signaling influences topography of islet cells in mice.

Glucagon-like peptide 1 (GLP-1) amplifies glucose-induced insulin release in vivo and in vitro. Activation of GLP-1 receptor (GLP-1R) signaling leads to differentiation of exocrine cells towards a beta-cell phenotype in vitro and stimulation of islet cell proliferation in vitro and in vivo, suggesting a potential role for GLP-1 in the modulation of islet growth and differentiation. To determine whether basal levels of GLP-1R signaling are essential for islet development, we examined islet cell composition and topography in GLP-1R-/- mice. Total beta-cell volume and number are not altered, but the topography of beta cells is markedly different in GLP-1R-/- mice compared with GLP-1R+/+ controls. The distribution of beta cells is shifted from large to small and medium-sized islets in the absence of GLP-1R signaling (large islets: 50 +/- 3% in GLP-1R+/+ vs 28 +/- 4% in GLP- 1R-/-, P < 0.01 and medium islets: 32 +/- 2% in GLP- 1R+/+ vs 48 +/- 3% in GLP-1R-/-, P < 0.001). Furthermore, GLP-1R-/- islets exhibit abnormalities in cell topography, with two to threefold more centrally located alpha cells detected in GLP-1R-/- islets. These alterations in alpha- and beta-cell topography indicate that basal levels of GLP-1 signaling in the normal rodent are involved in the normal cellular organization of the endocrine pancreas.

Elimination of glucagon-like peptide 1R signaling does not modify weight gain and islet adaptation in mice with combined disruption of leptin and GLP-1 action.

Leptin and glucagon-like peptide 1 (GLP-1) exhibit opposing actions in the endocrine pancreas. GLP-1 stimulates insulin biosynthesis, secretion, and islet growth, whereas leptin inhibits glucose-dependent insulin secretion and insulin gene transcription. In contrast, GLP-1 and leptin actions overlap in the central nervous system, where leptin has been shown to activate GLP-1 circuits that inhibit food intake. To determine the physiological importance of GLP-1 receptor (GLP-1R)-leptin interactions, we studied islet function and feeding behavior in ob/ob:GLP-1R(-/-) mice. Although GLP-1R actions are thought to be essential for glucose-dependent insulin secretion, the levels of fasting glucose, glycemic excursion after glucose loading, glucose-stimulated insulin, and pancreatic insulin RNA content were similar in ob/ob:GLP-1R(+/+) versus ob/ob:GLP-1R(-/-) mice. Despite evidence linking GLP-1R signaling to the regulation of islet neogenesis and proliferation, ob/ob:GLP-1R(-/-) mice exhibited significantly increased islet numbers and area and an increase in the number of large islets compared with GLP-1R(+/+) or (-/-) mice (P < -0.01 to 0.05). Similarly, growth rates and both shortand long-term control of food intake were comparable in ob/ob:GLP-1R(+/+) versus ob/ob:GLP-1R4(-/-) mice. Furthermore, leptin produced a similar inhibition of food intake in GLP-1R(-/-), ob/ob:GLP-1R(+/+), and ob/ob:GLP1R4(-/-) mice. These findings illustrate that although leptin and GLP-1 actions overlap in the brain and endocrine pancreas, disruption of GLP-1 signaling does not modify the response to leptin or the phenotype of leptin deficiency in the ob/ob mouse, as assessed by long-term control of body weight or the adaptive beta-cell response to insulin resistance in vivo.

Neuroendocrine function and response to stress in mice with complete disruption of glucagon-like peptide-1 receptor signaling.

Glucagon-like peptide-1 (GLP-1), a potent regulator of glucose homeostasis, is also produced in the central nervous system, where GLP-1 has been implicated in the neuroendocrine control of hypothalamic-pituitary function, food intake, and the response to stress. The finding that intracerebroventricular GLP-1 stimulates LH, TSH, corticosterone, and vasopressin secretion in rats prompted us to assess the neuroendocrine consequences of disrupting GLP-1 signaling in mice in vivo. Male GLP-1 receptor knockout (GLP-1R-/-) mice exhibit reduced gonadal weights, and females exhibit a slight delay in the onset of puberty; however, male and female GLP-1R-/- animals reproduce successfully and respond appropriately to fluid restriction. Although adrenal weights are reduced in GLP-1R-/- mice, hypothalamic CRH gene expression and circulating levels of corticosterone, thyroid hormone, testosterone, estradiol, and progesterone are normal in the absence of GLP-1R-/- signaling. Intriguingly, GLP-1R-/- mice exhibit paradoxically increased corticosterone responses to stress as well as abnormal responses to acoustic startle that are corrected by glucocorticoid treatment. These findings suggest that although GLP-1R signaling is not essential for development and basal function of the murine hypothalamic-pituitary-adrenal axis, abrogation of GLP-1 signaling is associated with impairment of the behavioral and neuroendocrine responses to stress.

Effects of aging and a high fat diet on body weight and glucose tolerance in glucagon-like peptide-1 receptor -/- mice.

Disruption of glucagon-like peptide-1 (GLP-1) receptor signaling in mice results in mild glucose intolerance, principally due to elimination of the incretin effect of GLP-1. Despite the inhibitory effects of GLP-1 on food intake, 6- to 8-week-old GLP-1 receptor -/-(GLP-1R-/-) mice were not obese and did not exhibit disturbances of feeding behavior. As both diabetes and obesity frequently become more phenotypically evident in older rodents, we studied the consequences of aging and a high fat diet on glucose control and body weight in GLP-1R-/- mice. No evidence of obesity or deterioration in glucose control was detected in 11- and 16-month-old GLP-1R-/- mice (mean weight, 34.7 +/- 2.0, 30.5 +/- 1.5, and 34.6 +/- 2.8 g in male and 25.3 +/-1.6, 28.4 +/-1.2, and 31.9 +/- 2.9 g in female GLP-1R+/+, GLP-1R+/-, and GLP-1R-/- mice, respectively; P = NS). After 18 weeks of high fat feeding, GLP-1R-/- mice gained similar (males) or less (females) weight than age- and sex-matched CD1 controls. No significant deterioration in glucose tolerance was observed after high fat feeding in GLP-1R-/- mice. These observations demonstrate that long term disruption of GLP-1 signaling in the central nervous system and peripheral tissues of older mice is not associated with the development of obesity or deterioration in glucose homeostasis.