The Role of ICL1 and H8 in Class B1 GPCRs; Implications for Receptor Activation.

The first intracellular loop (ICL1) of G protein-coupled receptors (GPCRs) has received little attention, although there is evidence that, with the 8th helix (H8), it is involved in early conformational changes following receptor activation as well as contacting the G protein ß subunit. In class B1 GPCRs, the distal part of ICL1 contains a conserved R12.48KLRCxR2.46b motif that extends into the base of the second transmembrane helix; this is weakly conserved as a [R/H]12.48KL[R/H] motif in class A GPCRs. In the current study, the role of ICL1 and H8 in signaling through cAMP, iCa2+ and ERK1/2 has been examined in two class B1 GPCRs, using mutagenesis and molecular dynamics. Mutations throughout ICL1 can either enhance or disrupt cAMP production by CGRP at the CGRP receptor. Alanine mutagenesis identified subtle differences with regard elevation of iCa2+, with the distal end of the loop being particularly sensitive. ERK1/2 activation displayed little sensitivity to ICL1 mutation. A broadly similar pattern was observed with the glucagon receptor, although there were differences in significance of individual residues. Extending the study revealed that at the CRF1 receptor, an insertion in ICL1 switched signaling bias between iCa2+ and cAMP. Molecular dynamics suggested that changes in ICL1 altered the conformation of ICL2 and the H8/TM7 junction (ICL4). For H8, alanine mutagenesis showed the importance of E3908.49b for all three signal transduction pathways, for the CGRP receptor, but mutations of other residues largely just altered ERK1/2 activation. Thus, ICL1 may modulate GPCR bias via interactions with ICL2, ICL4 and the Gß subunit.

A GLP-1/GIP/Gcg receptor triagonist improves memory behavior, as well as synaptic transmission, neuronal excitability and Ca2+ homeostasis in 3xTg-AD mice.

Alzheimer's disease (AD) is a progressively neurodegenerative disorder, which seriously affects human health and cannot be stopped by current treatments. Type 2 diabetes mellitus (T2DM) is a risk factor for AD. Our recent studies reported the neuroprotective effects of a GLP-1/GIP/Glucagon receptor triagonist (Triagonist), a novel unimolecular anti-diabetic drug, in cognitive and pathological improvements of 3xTg-AD mice. However, the detailed electrophysiological and molecular mechanisms underlying neuroprotection remain unexplored. The present study investigated the underlying electrophysiological and molecular mechanisms further by using whole-cell patch clamp techniques. Our results revealed that chronic Triagonist treatment effectively reduced working memory and reference memory errors of 3xTg-AD mice in a radial maze test. In addition, the Triagonist increased spontaneous excitatory synaptic activities, differentially modulated voltage- and chemically-gated Ca2+ flux, and reduced the over-excitation of pyramidal neurons in hippocampal slices of 3xTg-AD mice. In addition, chronic Triagonist treatment also up-regulated the expression levels of synaptophysin and PSD-95 in the hippocampus of 3xTg-AD mice. These results indicate that the Triagonist could improve memory formation, as well as synaptic transmission, Ca2+ balance, and neuronal excitability in 3xTg-AD mice. These neuroprotective effects of Triagonist may be involved in the up-regulation of synaptophysin and PSD-95. Therefore, the study suggests that multi-receptor agonists might be a novel therapeutic strategy for the treatment of AD.

Glucagon-Like Peptide-1 Receptor Activation Does not Affect Re-Endothelialization but Reduces Intimal Hyperplasia via Direct Effects on Smooth Muscle Cells in a Nondiabetic Model of Arterial Injury.

UNLABELLED: Diabetic patients have an increased risk of restenosis and late stent thrombosis after angioplasty, i.e. complications that are related to a defective re-endothelialization. Exendin-4, a stable glucagon-like peptide (GLP)-1 receptor agonist, has been suggested to influence the formation of intimal hyperplasia and to increase endothelial cell proliferation in vitro. Thus, the aim of this study was to investigate the mechanisms by which treatment with exendin-4 could influence re-endothelialization and intimal hyperplasia after vascular injury. METHODS: Sprague-Dawley rats were subjected to balloon injury of the left common carotid artery and treated for 4 weeks with exendin-4 or vehicle. Intimal hyperplasia and vessel wall elasticity were monitored noninvasively by high-frequency ultrasound, and re-endothelialization was evaluated upon sacrifice using Evans blue dye. RESULTS AND CONCLUSION: Exendin-4 selectively reduced the proliferation of smooth muscle cells (SMCs) and intimal hyperplasia in vivo without affecting the re-endothelialization process, but treatment with exendin-4 improved arterial wall elasticity. Our data also show that exendin-4 significantly decreased the proliferation and increased the apoptosis of SMCs in vitro, effects that appear to be mediated through cAMP signaling and endothelial nitric oxide synthase following GLP-1 receptor activation. Together, these effects of exendin-4 are highly desirable and may lead to an improved outcome for patients undergoing vascular interventions.

GLP-1R Agonists Modulate Enteric Immune Responses Through the Intestinal Intraepithelial Lymphocyte GLP-1R.

Obesity and diabetes are characterized by increased inflammation reflecting disordered control of innate immunity. We reveal a local intestinal intraepithelial lymphocyte (IEL)-GLP-1 receptor (GLP-1R) signaling network that controls mucosal immune responses. Glp1r expression was enriched in intestinal IEL preparations and copurified with markers of Tαß and Tγδ IELs, the two main subsets of intestinal IELs. Exendin-4 increased cAMP accumulation in purified IELs and reduced the production of cytokines from activated IELs but not from splenocytes ex vivo. These actions were mimicked by forskolin, absent in IELs from Glp1r(-/-) mice, and attenuated by the GLP-1R agonist exendin (9-39) consistent with a GLP-1R-dependent mechanism of action. Furthermore, Glp1r(-/-) mice exhibited dysregulated intestinal gene expression, an abnormal representation of microbial species in feces, and enhanced sensitivity to intestinal injury following administration of dextran sodium sulfate. Bone marrow transplantation using wild-type C57BL/6 donors normalized expression of multiple genes regulating immune function and epithelial integrity in Glp1r(-/-) recipient mice, whereas acute exendin-4 administration robustly induced the expression of genes encoding cytokines and chemokines in normal and injured intestine. Taken together, these findings define a local enteroendocrine-IEL axis linking energy availability, host microbial responses, and mucosal integrity to the control of innate immunity.

Exenatide Protects Against Glucose- and Lipid-Induced Endothelial Dysfunction: Evidence for Direct Vasodilation Effect of GLP-1 Receptor Agonists in Humans.

GLP-1 receptor (GLP-1R) agonists may improve endothelial function (EF) via metabolic improvement and direct vascular action. The current study determined the effect of GLP-1R agonist exenatide on postprandial EF in type 2 diabetes and the mechanisms underlying GLP-1R agonist-mediated vasodilation. Two crossover studies were conducted: 36 participants with type 2 diabetes received subcutaneous exenatide or placebo for 11 days and EF, and glucose and lipid responses to breakfast and lunch were determined; and 32 participants with impaired glucose tolerance (IGT) or diet-controlled type 2 diabetes had EF measured before and after intravenous exenatide, with or without the GLP-1R antagonist exendin-9. Mechanisms of GLP-1R agonist action were studied ex vivo on human subcutaneous adipose tissue arterioles and endothelial cells. Subcutaneous exenatide increased postprandial EF independent of reductions in plasma glucose and triglycerides. Intravenous exenatide increased fasting EF, and exendin-9 abolished this effect. Exenatide elicited eNOS activation and NO production in endothelial cells, and induced dose-dependent vasorelaxation and reduced high-glucose or lipid-induced endothelial dysfunction in arterioles ex vivo. These effects were reduced with AMPK inhibition. In conclusion, exenatide augmented postprandial EF in subjects with diabetes and prevented high-glucose and lipid-induced endothelial dysfunction in human arterioles. These effects were largely direct, via GLP-1R and AMPK activation.

Evidence That in Uncontrolled Diabetes, Hyperglucagonemia Is Required for Ketosis but Not for Increased Hepatic Glucose Production or Hyperglycemia.

Several lines of evidence implicate excess glucagon secretion in the elevated rates of hepatic glucose production (HGP), hyperglycemia, and ketosis characteristic of uncontrolled insulin-deficient diabetes (uDM), but whether hyperglucagonemia is required for hyperglycemia in this setting is unknown. To address this question, adult male Wistar rats received either streptozotocin (STZ) to induce uDM (STZ-DM) or vehicle and remained nondiabetic. Four days later, animals received daily subcutaneous injections of either the synthetic GLP-1 receptor agonist liraglutide in a dose-escalating regimen to reverse hyperglucagonemia or its vehicle for 10 days. As expected, plasma glucagon levels were elevated in STZ-DM rats, and although liraglutide treatment lowered glucagon levels to those of nondiabetic controls, it failed to attenuate diabetic hyperglycemia, elevated rates of glucose appearance (Ra), or increased hepatic gluconeogenic gene expression. In contrast, it markedly reduced levels of both plasma ketone bodies and hepatic expression of the rate-limiting enzyme involved in ketone body production. To independently confirm this finding, in a separate study, treatment of STZ-DM rats with a glucagon-neutralizing antibody was sufficient to potently lower plasma ketone bodies but failed to normalize elevated levels of either blood glucose or Ra. These data suggest that in rats with uDM, hyperglucagonemia is required for ketosis but not for increased HGP or hyperglycemia.

GLP-1 and exendin-4 transiently enhance GABAA receptor-mediated synaptic and tonic currents in rat hippocampal CA3 pyramidal neurons.

Glucagon-like peptide-1 (GLP-1) is a hormone that stimulates insulin secretion. Receptors for GLP-1 are also found in the brain, including the hippocampus, the center for memory and learning. Diabetes is a risk factor for decreased memory functions. We studied effects of GLP-1 and exendin-4, a GLP-1 receptor agonist, on γ-aminobutyric acid (GABA) signaling in hippocampal CA3 pyramidal neurons. GABA is the main inhibitory neurotransmitter and decreases neuronal excitability. GLP-1 (0.01-1 nmol/L) transiently enhanced synaptic and tonic currents, and the effects were blocked by exendin (9-39). Ten pmol/L GLP-1 increased both the spontaneous inhibitory postsynaptic current (sIPSC) amplitudes and frequency by a factor of 1.8. In 0.1, 1 nmol/L GLP-1 or 10, 50, or 100 nmol/L exendin-4, only the sIPSC frequency increased. The tonic current was enhanced by 0.01-1 nmol/L GLP-1 and by 0.5-100 nmol/L exendin-4. When action potentials were inhibited by tetrodotoxin (TTX), inhibitory postsynaptic currents decreased and currents were no longer potentiated by GLP-1 or exendin-4. In contrast, although the tonic current decreased in TTX, it was still enhanced by GLP-1 or exendin-4. The results demonstrate GLP-1 receptor regulation of hippocampal function and are consistent with GLP-1 receptor agonists enhancing GABAA signaling by pre- and postsynaptic mechanisms.

[The physiology of glucagon-like peptide-1 and its role in the pathophysiology of type 2 diabetes mellitus]. / Fisiología del GLP-1 y su papel en la fisiopatología de la diabetes mellitus tipo 2.

The hormone glucagon-like peptide-1 (GLP-1) is synthesized and secreted by L cells in the small intestine in response to food ingestion. After reaching the general circulation it has a half-life of 2-3 minutes due to degradation by the enzyme dipeptidyl peptidase-4. Its physiological role is directed to control plasma glucose concentration, though GLP-1 also plays other different metabolic functions following nutrient absorption. Biological activities of GLP-1 include stimulation of insulin biosynthesis and glucose-dependent insulin secretion by pancreatic beta cell, inhibition of glucagon secretion, delay of gastric emptying and inhibition of food intake. GLP-1 is able to reduce plasma glucose levels in patients with type 2 diabetes and also can restore beta cell sensitivity to exogenous secretagogues, suggesting that the increasing GLP-1 concentration may be an useful therapeutic strategy for the treatment of patients with type 2 diabetes.

[Glucagon-like peptide-1 (GLP-1) mimetics: a new treatment for Alzheimer's disease?]. / Análogos del glucagon-like peptide-1 (GLP-1): ¿una nueva estrategia de tratamiento para la enfermedad de Alzheimer?

INTRODUCTION: The glucagon-like peptide-1 (GLP-1) mimetics are an established therapeutic option for patients with type 2 diabetes. However, the properties of the GLP-1 mimetics go beyond the strict metabolic control of the patients with diabetes. The neuroprotective effects of GLP-1 have been shown in recent studies opening new areas of research in neurodegenerative diseases such as Alzheimer's disease (AD), among others. AIM. Systematic review including experimental studies and human clinical trials demonstrating the neuroprotective properties of GLP-1 mimetics in AD. DEVELOPMENT: The experimental studies that have been conducted in rodent models of AD have demonstrated the neuroprotective properties of GLP-1 in the central nervous system reducing beta-amyloid plaques, the oxidative stress and the inflammatory brain response. Clinical trials in patients with cognitive impairment and AD testing the effects of GLP-1 analogs have recently started. CONCLUSION: The GLP-1 analogs have neuroprotective properties. Considering that type 2 diabetes is a risk factor for cognitive impairment and dementia, the benefits of GLP-1 mimetics on cognition must be considered. Likewise, the GLP-1 mimetics represent a promising treatment for neurodegenerative diseases such as AD.

TITLE: Analogos del glucagon-like peptide-1 (GLP-1): una nueva estrategia de tratamiento para la enfermedad de Alzheimer?Introduccion. Los analogos del glucagon-like peptide-1 (GLP-1) son una opcion terapeutica establecida en los pacientes con diabetes tipo 2. Sin embargo, las propiedades de los analogos del GLP-1 van mas alla del control estrictamente metabolico del paciente diabetico. Los efectos neuroprotectores de los analogos del GLP-1 se han puesto de manifiesto en estudios recientes y han abierto nuevos campos de investigacion en trastornos neurodegenerativos como la enfermedad de Alzheimer (EA), entre otros. Objetivo. Revision sistematica de los estudios experimentales y ensayos clinicos en humanos que demuestran las propiedades neuroprotectoras de los analogos del GLP-1 en la EA. Desarrollo. Los estudios experimentales que se han llevado a cabo en modelos de roedores con EA demuestran las propiedades neuroprotectoras de los analogos del GLP-1 sobre el sistema nervioso central que reducen las placas de beta-amiloide, el estres oxidativo y la respuesta inflamatoria cerebral. Recientemente se han puesto en marcha estudios con analogos del GLP-1 en humanos con deterioro cognitivo y EA. Conclusiones. Los analogos del GLP-1 presentan propiedades neuroprotectoras. Al considerarse la diabetes tipo 2 un factor de riesgo para el deterioro cognitivo y la demencia, deben considerarse los beneficios de los analogos del GLP-1 sobre la cognicion. Del mismo modo, los analogos del GLP-1 suponen un tratamiento prometedor en la EA.

GLP-1 receptor activation modulates appetite- and reward-related brain areas in humans.

Gut-derived hormones, such as GLP-1, have been proposed to relay information to the brain to regulate appetite. GLP-1 receptor agonists, currently used for the treatment of type 2 diabetes (T2DM), improve glycemic control and stimulate satiety, leading to decreases in food intake and body weight. We hypothesized that food intake reduction after GLP-1 receptor activation is mediated through appetite- and reward-related brain areas. Obese T2DM patients and normoglycemic obese and lean individuals (n = 48) were studied in a randomized, crossover, placebo-controlled trial. Using functional MRI, we determined the acute effects of intravenous administration of the GLP-1 receptor agonist exenatide, with or without prior GLP-1 receptor blockade using exendin 9-39, on brain responses to food pictures during a somatostatin pancreatic-pituitary clamp. Obese T2DM patients and normoglycemic obese versus lean subjects showed increased brain responses to food pictures in appetite- and reward-related brain regions (insula and amygdala). Exenatide versus placebo decreased food intake and food-related brain responses in T2DM patients and obese subjects (in insula, amygdala, putamen, and orbitofrontal cortex). These effects were largely blocked by prior GLP-1 receptor blockade using exendin 9-39. Our findings provide novel insights into the mechanisms by which GLP-1 regulates food intake and how GLP-1 receptor agonists cause weight loss.

Glucagon-like peptide-1 receptor agonist activation ameliorates venous thrombosis-induced arteriovenous fistula failure in chronic kidney disease.

High shear stress that develops in the arteriovenous fistula of chronic kidney diseases (CKD) may increase H2O2 and thromboxane A2 (TXA2) release, thereby exacerbating endothelial dysfunction, thrombosis, and neointimal hyperplasia. We investigated whether glucagon-like peptide-1 receptor agonist/exendin-4, a potentially cardiovascular protective agent, could improve TXA2-induced arteriovenous fistula injury in CKD. TXA2 administration to H2O2-exposed human umbilical vein endothelial cells increased apoptosis, senescence, and detachment; these phenotypes were associated with the downregulation of phosphorylated endothelial nitric oxide synthase/heme oxygenase-1 (eNOS/HO-1) signalling. Exendin-4 reduced H2O2/TXA2-induced endothelial injury via inhibition of apoptosis-related mechanisms and restoration of phosphorylated eNOS/HO-1 signalling. Male Wistar rats subjected to right common carotid artery-external jugular vein anastomosis were treated with exendin-4 via cervical implant osmotic pumps for 16-42 days. High shear stress induced by the arteriovenous fistula significantly increased venous haemodynamics, blood and tissue H2O2 and TXB2 levels, macrophage/monocyte infiltration, fibrosis, proliferation, and adhesion molecule-1 expression. Apoptosis was also increased due to NADPH oxidase gp91 activation and mitochondrial Bax translocation in the proximal end of the jugular vein of CKD rats. Exendin-4-treatment of rats with CKD led to the restoration of normal endothelial morphology and correction of arteriovenous fistula function. Exendin-4 treatment or thromboxane synthase gene deletion in CKD mice markedly reduced ADP-stimulated platelet adhesion to venous endothelium, and prevented venous occlusion in FeCl3-injured vessels by upregulation of HO-1. Together, these data reveal that the use of glucagon-like peptide-1 receptor agonists is an effective strategy for treatment of CKD-induced arteriovenous fistula failure.

Characterization of [¹²5I]GLP-1(9-36), a novel radiolabeled analog of the major metabolite of glucagon-like peptide 1 to a receptor distinct from GLP1-R and function of the peptide in murine aorta.

AIMS: Glucagon-like peptide 1 (GLP-1) is an insulin secretagogue, released in response to meal ingestion and efficiently lowers blood glucose in Type 2 diabetic patients. GLP-1(7-36) is rapidly metabolized by dipeptidyl peptidase IV to the major metabolite GLP-1(9-36)-amide, often thought to be inactive. Inhibitors of this enzyme are widely used to treat diabetes. Our aim was to characterize the binding of GLP-1(9-36) to native mouse tissues and to cells expressing GLP1-R as well as to measure functional responses in the mouse aorta compared with GLP-1(7-36). MAIN METHODS: The affinity of [(125)I]GLP-1(7-36) and [(125)I]GLP-1(9-36) was measured in mouse tissues by saturation binding and autoradiography used to determine receptor distribution. The affinity of both peptides was compared in binding to recombinant GLP-1 receptors using cAMP and scintillation proximity assays. Vasoactivity was determined in mouse aortae in vitro. KEY FINDINGS: In cells expressing GLP-1 receptors, GLP-1(7-36) bound with the expected high affinities (0.1 nM) and an EC50 of 0.07 nM in cAMP assays but GLP-1(9-36) bound with 70,000 and 100,000 fold lower affinities respectively. In contrast, in mouse brain, both labeled peptides bound with a single high affinity, with Hill slopes close to unity, although receptor density was an order of magnitude lower for [(125)I]GLP-1(9-36). In functional experiments both peptides had similar potencies, GLP-1(7-36), pD2=7.40 ± 0.24 and GLP-1(9-36), pD2=7.57 ± 0.64. SIGNIFICANCE: These results suggest that GLP-1(9-36) binds and has functional activity in the vasculature but these actions may be via a pathway that is distinct from the classical GLP-1 receptor and insulin secretagogue actions.

The protective roles of GLP-1R signaling in diabetic nephropathy: possible mechanism and therapeutic potential.

Glucagon-like peptide-1 (GLP-1) is a gut incretin hormone that has an antioxidative protective effect on various tissues. Here, we determined whether GLP-1 has a role in the pathogenesis of diabetic nephropathy using nephropathy-resistant C57BL/6-Akita and nephropathy-prone KK/Ta-Akita mice. By in situ hybridization, we found the GLP-1 receptor (GLP-1R) expressed in glomerular capillary and vascular walls, but not in tubuli, in the mouse kidney. Next, we generated C57BL/6-Akita Glp1r knockout mice. These mice exhibited higher urinary albumin levels and more advanced mesangial expansion than wild-type C57BL/6-Akita mice, despite comparable levels of hyperglycemia. Increased glomerular superoxide, upregulated renal NAD(P)H oxidase, and reduced renal cAMP and protein kinase A (PKA) activity were noted in the Glp1r knockout C57BL/6-Akita mice. Treatment with the GLP-1R agonist liraglutide suppressed the progression of nephropathy in KK/Ta-Akita mice, as demonstrated by reduced albuminuria and mesangial expansion, decreased levels of glomerular superoxide and renal NAD(P)H oxidase, and elevated renal cAMP and PKA activity. These effects were abolished by an adenylate cyclase inhibitor SQ22536 and a selective PKA inhibitor H-89. Thus, GLP-1 has a crucial role in protection against increased renal oxidative stress under chronic hyperglycemia, by inhibition of NAD(P)H oxidase, a major source of superoxide, and by cAMP-PKA pathway activation.

Central GLP-2 enhances hepatic insulin sensitivity via activating PI3K signaling in POMC neurons.

Glucagon-like peptides (GLP-1/GLP-2) are coproduced and highlighted as key modulators to improve glucose homeostasis and insulin sensitivity after bariatric surgery. However, it is unknown if CNS GLP-2 plays any physiological role in the control of glucose homeostasis and insulin sensitivity. We show that mice lacking GLP-2 receptor (GLP-2R) in POMC neurons display glucose intolerance and hepatic insulin resistance. GLP-2R activation in POMC neurons is required for GLP-2 to enhance insulin-mediated suppression of hepatic glucose production (HGP) and gluconeogenesis. GLP-2 directly modulates excitability of POMC neurons in GLP-2R- and PI3K-dependent manners. GLP-2 initiates GLP-2R-p85α interaction and facilitates PI3K-Akt-dependent FoxO1 nuclear exclusion in POMC neurons. Central GLP-2 suppresses basal HGP and enhances insulin sensitivity, which are abolished in POMC-p110α KO mice. Thus, CNS GLP-2 plays a key physiological role in the control of HGP through activating PI3K-dependent modulation of membrane excitability and nuclear transcription of POMC neurons in the brain.

Hypothalamic glucagon signaling inhibits hepatic glucose production.

Glucagon activates hepatic protein kinase A (PKA) to increase glucose production, but the gluco-stimulatory effect is transient even in the presence of continuous intravenous glucagon infusion. Continuous intravenous infusion of insulin, however, inhibits glucose production through its sustained actions in both the liver and the mediobasal hypothalamus (MBH). In a pancreatic clamp setting, MBH infusion with glucagon activated MBH PKA and inhibited hepatic glucose production (HGP) in rats, as did central glucagon infusion in mice. Inhibition of glucagon receptor-PKA signaling in the MBH and hepatic vagotomy each negated the effect of MBH glucagon in rats, whereas the central effect of glucagon was diminished in glucagon receptor knockout mice. A sustained rise in plasma glucagon concentrations transiently increased HGP, and this transiency was abolished in rats with negated MBH glucagon action. In a nonclamp setting, MBH glucagon infusion improved glucose tolerance, and inhibition of glucagon receptor-PKA signaling in the MBH enhanced the ability of intravenous glucagon injection to increase plasma glucose concentrations. We also detected a similar enhancement of glucose concentrations that was associated with a disruption in MBH glucagon signaling in rats fed a high-fat diet. We show that hypothalamic glucagon signaling inhibits HGP and suggest that hypothalamic glucagon resistance contributes to hyperglycemia in diabetes and obesity.

Glucagon-like peptide-1 receptor-mediated endosomal cAMP generation promotes glucose-stimulated insulin secretion in pancreatic ß-cells.

Glucagon-like peptide-1 receptor (GLP-1R) plays a major role in promoting glucose-stimulated insulin secretion in pancreatic ß-cells. In the present study, we synthesized a novel functional analog of GLP-1 conjugated to tetramethyl rhodamine to monitor the internalization of the receptor. Our data show that after being internalized the receptor is sorted to lysosomes. In endosomes, receptor-ligand complex is found to be colocalized with adenylate cyclase. Pharmacological inhibition of endocytosis attenuates GLP-1R-mediated cAMP generation and consequent downstream protein kinase A substrate phosphorylation and glucose-stimulated insulin secretion. Our study underlines a paradigm shift in GLP-1R signaling and trafficking. The receptor ligand complex triggers cAMP generation both in plasma membrane and in endosomes, which has implications for receptor-mediated regulation of insulin secretion.

High-fat diet changes the temporal profile of GLP-1 receptor-mediated hypophagia in rats.

Overconsumption of a high-fat diet promotes weight gain that can result in obesity and associated comorbidities, including Type 2 diabetes mellitus. Consumption of a high-fat diet also alters gut-brain communication. Glucagon-like peptide 1 (GLP-1) is an important gastrointestinal signal that modulates both short- and long-term energy balance and is integral in maintenance of glucose homeostasis. In the current study, we investigated whether high-fat diets (40% or 81% kcal from fat) modulated the ability of the GLP-1 receptor (GLP-1r) agonists exendin-4 (Ex4) and liraglutide to reduce food intake and body weight. We observed that rats maintained on high-fat diets had a delayed acute anorexic response to peripheral administration of Ex4 or liraglutide compared with low-fat diet-fed rats (17% kcal from fat). However, once suppression of food intake in response to Ex4 or liraglutide started, the effect persisted for a longer time in the high-fat diet-fed rats compared with low-fat diet-fed rats. In contrast, centrally administered Ex4 suppressed food intake similarly between high-fat diet-fed and low-fat diet-fed rats. Chronic consumption of a high-fat diet did not change the pharmacokinetics of Ex4 but increased intestinal Glp1r expression and decreased hindbrain Glp1r expression. Taken together, these findings demonstrate that dietary composition alters the temporal profile of the anorectic response to exogenous GLP-1r agonists.

Glucagon receptor is required for long-term survival: a natural history study of the Mahvash disease in a murine model.

BACKGROUND AND AIM: We have described a novel Mahvash disease of hyperglucagonemia and pancreatic neuroendocrine tumors (PNETs) associated with an inactivating glucagon receptor mutation, and identified the glucagon receptor-deficient (Gcgr(-/-)) mice as its murine model. We aim to elucidate the natural history of the rare Mahvash disease by long-term observation of the Gcgr(-/-) mice. MATERIALS AND METHOD: Wild type (WT) (n=52), heterozygous (n=127), and Gcgr(-/-) (n=56) mice living under standard vivarium conditions were observed without specific treatments over 22 months. Autopsy was performed on dead animals. RESULTS: The WT and heterozygous mice did not exhibit any measurable differences. The Gcgr(-/-) mice became progressively lethargic and cachexic after 12 months. Random glucose levels were stable in WT and heterozygous mice but decreased with age in the Gcgr(-/-) mice. At the end of observation, 28/56 Gcgr(-/-), 7/52 WT, and 24/127 heterozygous mice died. The survival curve of Gcgr(-/-) mice began to separate from those of WT and heterozygous mice at 12 months and the survival difference widened with age. At 18 months, survival probability was 17% for Gcgr(-/-) mice but 77% for WT and 81% for heterozygous mice. Autopsy revealed numerous PNETs up to 15 mm in diameter in most well-preserved Gcgr(-/-) pancreata (17/20) but none in WT or heterozygous ones. Four Gcgr(-/-) mice developed liver or subcutaneous metastasis. CONCLUSION: The untreated Mahvash disease may cause cachexia, severe hypoglycemia, and early death. Patients with Mahvash disease need to undergo life-long surveillance for PNETs. Functional glucagon receptor is thus required for long-term survival.

Physiology and emerging biochemistry of the glucagon-like peptide-1 receptor.

The glucagon-like peptide-1 (GLP-1) receptor is one of the best validated therapeutic targets for the treatment of type 2 diabetes mellitus (T2DM). Over several years, the accumulation of basic, translational, and clinical research helped define the physiologic roles of GLP-1 and its receptor in regulating glucose homeostasis and energy metabolism. These efforts provided much of the foundation for pharmaceutical development of the GLP-1 receptor peptide agonists, exenatide and liraglutide, as novel medicines for patients suffering from T2DM. Now, much attention is focused on better understanding the molecular mechanisms involved in ligand induced signaling of the GLP-1 receptor. For example, advancements in biophysical and structural biology techniques are being applied in attempts to more precisely determine ligand binding and receptor occupancy characteristics at the atomic level. These efforts should better inform three-dimensional modeling of the GLP-1 receptor that will help inspire more rational approaches to identify and optimize small molecule agonists or allosteric modulators targeting the GLP-1 receptor. This article reviews GLP-1 receptor physiology with an emphasis on GLP-1 induced signaling mechanisms in order to highlight new molecular strategies that help determine desired pharmacologic characteristics for guiding development of future nonpeptide GLP-1 receptor activators.

Targeting the glucagon receptor family for diabetes and obesity therapy.

Diabetes is a debilitating disease characterized by chronic hyperglycemia and is often associated with obesity. With diabetes and obesity incidence on the rise, it is imperative to develop novel therapeutics that will not only lower blood glucose levels, but also combat the associated obesity. The G protein-coupled receptors (GPCRs) for glucose-dependent insulinotropic polypeptide (GIP), glucagon-like peptide-1 (GLP-1) and glucagon are emerging as targets to treat both hyperglycemia and obesity. GIP is rapidly released from intestinal K-cells following food intake and stimulates glucose-dependent insulin secretion from ß-cells and the storage of fat in adipocytes. Both GIP receptor agonists and antagonists have been demonstrated to display therapeutic potential to treat diabetes and obesity. Similar to GIP, GLP-1 is released from intestinal L-cells following food intake and potentiates glucose-dependent insulin secretion from ß-cells. In addition, GLP-1 reduces glucagon levels, suppresses gastric emptying and reduces food intake. As such, GLP-1 receptor agonists effectively lower blood glucose levels and reduce weight. Finally, glucagon is released from α-cells and raises blood glucose levels during the fasting state by stimulating gluconeogenesis and glycogenolysis in the liver. Thus, molecules that antagonize the glucagon receptor may be used to treat hyperglycemia. Given the structural similarity of these peptides and their receptors, molecules capable of agonizing or antagonizing combinations of these receptors have recently been suggested as even better therapeutics. Here we review the biology of GIP, GLP-1 and glucagon and examine the various therapeutic strategies to activate and antagonize the receptors of these peptides.