Glucagon‑like Peptide‑1 (GLP- 1) and Receptors

After consuming a meal, the L cells of the intestine produce and release the 30 amino acid peptide GLP- 1, which in turn promotes insulin secretion [6]. GLP- 1-based therapy exerts its pharmacological effects by activating the GLP- 1 receptor, a class of G protein-coupled receptor consisting of 463 amino acids. The receptor is a glycoprotein that, like all G protein-coupled receptors, has a seven-transmembrane helix domain and an N-terminal extracellular signal peptide [7]. When GLP- 1 receptors are activated, adenosine triphosphate (ATP) is converted to cyclic adenosine monophosphate (cAMP). Increased signaling through exchange proteins directly activated by cAMP (EPACs), specifically EPAC2, and activation of protein kinase A (PKA) follow the rise in cAMP. An increase in PKA activity also causes ATP-sensitive potassium channels to close, thereby depolarizing the cell membrane and opening voltage-gated calcium channels. This absorbs calcium and stimulates exocytosis of secretory granules, which in turn causes the release of insulin. Furthermore, the endoplasmic reticulum releases calcium in response to EPAC2 activation, increasing intracellular calcium levels and promoting exocytosis. Activation of the GLP- 1 receptor also prevents beta cell apoptosis. Additional consequences of GLP- 1 receptor activation include appetite suppression, cardioprotective effects, and inhibition of glucagon secretion associated with inhibition of gastric emptying [6]. Whether GLP- 1 increases energy expenditure has also been studied. Although there are data in animal models supporting that this is true, particularly in diet-induced obese mice, in which brown adipose tissue activity is regulated [8], there is no convincing evidence that this occurs in humans [6]. As a result, GLP- 1's primary effects include decreasing body weight, protecting against cardiovascular disease, and lowering fasting and postprandial blood glucose levels—all of which are therapeutic targets in T2DM. The short half-life of native GLP- 1-based therapy in the circulation (approximately two minutes after intravenous administration and one to five hours after subcutaneous administration) posed a problem during its early development [9]. For this reason, the first experiments of the glucose-lowering effect of GLP- 1 used continuous intravenous administration or intramuscular injections [10]. The short duration of action needed to be addressed to exploit the promising effects of GLP- 1 for the development of a clinically beneficial drug.

Glucose-dependent Insulinotropic Polypeptide (GIP) Receptor

During fasting, K cells (found in the duodenum and jejunum) constitutively secrete the 42 amino acid protein GIP, which exhibits a marked increase in secretion after meals [11,12,13]. GIP was initially discovered because it inhibited the secretion of gastric acid. Since then, numerous investigations have revealed the effects of GIP on multiple organs [14]. The GIP receptor, a class of B G-protein-coupled receptors (GPCRs) that is widely expressed in a variety of tissues, is the binding site that mediates the systemic effects. The stomach, adipose tissue, bone, and pancreas all express GIP receptors, and these have also been found in the cerebral cortex, hippocampus, and olfactory bulb, among other parts of the brain [15, 16]. GIP is an incretin hormone that indirectly regulates glucagon secretion and increases glucose-stimulated insulin secretion [17]. GIP and GLP- 1 have opposing effects on glucagon release, despite their similarities. Studies conducted in isolated rat islets have shown that GIP does, in fact, increase intracellular cAMP levels, which in turn stimulate glucagon secretion [18]. The clinical application of GIP agonists for the treatment of diabetes is hampered by this increase in glucagon secretion, which has also been verified in both healthy and subjects with T2DM [19, 20]. In addition to its effects on insulin, GIP plays a crucial role in fat accumulation by promoting the activity of adipocyte-expressed lipoprotein lipase and promoting bone formation by preventing osteoclast apoptosis, as evidenced by the thinner bone trabeculae seen in mice lacking GIP receptors [20,21,22]. Better memory was also demonstrated by GIP-transgenic mice, which is most likely connected to increased neurogenesis [23]. In support to this theory, it has been demonstrated that GIP infusion increases the proliferation of neuronal progenitors in the dentate gyrus, while mice lacking GIP displayed deterioration in memorial tasks as a result of reduced neurogenesis [24].

Glucagon Receptor

The first description of the hyperglycemic effects of glucagon was made more than a century ago [25, 26]. By attaching itself to its receptors, this hormone works in tandem with insulin to control blood sugar levels [14]. Glucagon's capacity to regulate hepatic glucose metabolism is primarily responsible for its hyperglycemic effects [27]. In order to maintain a steady supply of glucose, glucagon strongly stimulates glycogenolysis and suppresses glycogenesis in the liver [28, 29]. Many studies have revealed numerous additional functions of glucagon in the brain, liver, heart, and adipose tissue in addition to its well-known function in regulating blood sugar levels [30,31,32]. Nearly 40 years ago, it was discovered that the rat brain contained glucagon receptors, indicating that the hormone may play a part in controlling brain activity [33]. Accordingly, glucagon has been shown to reduce appetite, food intake, and promote weight loss in both humans and rodents [34, 35]. Despite the fact that the relationship between glucagon and body weight is well established, some research found no changes in food intake following glucagon administration, which suggests that appetite-independent mechanisms may play a role in the glucagon-mediated regulation of body weight [14]. Rats that receive a single subcutaneous dose of glucagon exhibit a sharp rise in metabolic rate, which is consistent with this theory [36]. By promoting oxygen consumption in brown adipose tissue (BAT), glucagon increases metabolic rate. This has been demonstrated by elevated BAT temperature in rats [37, 38]. By preventing lipogenesis and promoting lipolysis, glucagon also has lipolytic effects on white adipose tissue [14]. Hormone-sensitive lipase (HSL) in adipocytes is activated to produce the lipolytic effect, which is then enhanced by indirect processes such as secretion of growth hormone, cortisol, and adrenaline [39]. Adipose tissue is not the only area where glucagon affects lipid metabolism; the liver also experiences an increase in ketogenesis [40]. In fact, by continuously depriving the liver of esterified fatty acids and inhibiting the hepatic glycolytic pathway, glucagon promotes the production of ketone bodies [41,42,43] Hepatic ketogenesis is boosted as a result, and mitochondrial fatty acid oxidation is improved [14, 43]. By attaching itself to its receptors and activating adenylate cyclase (AC), which raises cAMP levels in the myocardium, glucagon also increases cardiac output. Glucagon has very rapid chronotropic and inotropic effects, peaking five minutes after administration and lasting for twenty minutes [14, 32].

Pathways for Intracellular Signaling in the Langerhans Islets

GLP- 1 receptor, GIP receptor, and glucagon receptor are mainly expressed on beta, alpha, and delta cells in the islets of Langerhans, where they influence hormone secretion and regulate the survival and proliferation of endocrine cells. The search for GLP- 1 receptor agonists to treat T2DM was sparked by studies showing the insulinotropic and glucose-lowering effects of GLP- 1 in beta cells, as well as the hyperglycemic effects of GLP- 1 receptor antagonism [44]. Importantly, individuals with T2DM continue to benefit from GLP- 1's glucose-lowering effects. Insulin gene transcription and translation, as well as the enhancement of glucose-stimulated insulin secretion (insulinotropic effects), are caused by GLP- 1 receptor activation in the beta cell. GLP- 1 receptor activation has longer-term consequences, such as increased b cell proliferation and neogenesis along with cytoprotective (non-insulinotropic) effects [45,46,47,48,49,50,51].

GLP- 1 Receptor Signaling

The heterotrimeric G proteins are known to mediate signaling through the canonical GLP- 1 receptor. These G proteins have both a Gb/c dimer subunit and an independent Ga subunit. GPCRs that are agonist-activated promote the synthesis of guanine triphosphate (GTP), which when bound to the G-protein causes the Ga and Gbc subunits to separate, which in turn can activate signaling proteins downstream [52]. It was initially discovered that GLP- 1 activates adenylate cyclase in tissue from the central nervous system (CNS) [53]. It has been demonstrated that ligand-activated GLP- 1 receptor interacts with the Gas subunit and activates adenylate cyclase to generate cAMP. Both the EPAC2 and PKA are activated in response to elevated cAMP [54]. By increasing Ca2+ influx and consequently insulin granule exocytosis, both pathways work together to promote insulin secretion. PKA directly phosphorylates sulfonylurea receptor 1 (SUR1), a regulatory subunit of voltage-dependent K+ channels and KATP channels, increasing membrane depolarization and voltage-gated Ca2+ channel activation. Through several mechanisms, including direct interaction with SUR1 and membrane depolarization, intracellular calcium mobilization, and insulin granule priming modification, EPAC2 activation may promote the exocytosis of insulin granules [55, 56].

There have also been reports of non-insulinotropic effects of GLP- 1 receptor activation. In fact, the GLP- 1 receptor-cAMP-PKA axis stimulates the growth of beta cells by activating the transducer of regulated cAMP response element-binding protein (CREB) (TORC2) and the expression of the CREB and insulin receptor substrate 2 genes [57] Additionally, by boosting bcl- 2 activity and blocking the proapoptotic Bax, activated CREB also supports beta cell survival [58]. GLP- 1's anti-apoptotic and proliferative effects are also mediated by the PI3 K/Akt axis through the transactivation of the epidermal growth factor receptor (EGFR), which is connected to GLP- 1 receptor through c-Src activation and the generation of endogenous EGF like ligands [59]. GLP- 1 receptor agonism has not been thoroughly studied, but it has been demonstrated in certain cell lines to activate additional G-protein subunits and alternative downstream signaling pathways [60,61,62]

GIP Receptor Signaling

The mature 42 amino acid GIP attaches itself to the b cell surface and activates its cognate receptor, the GIP receptor. Ligand binding to the GIP receptor activates Gas, which in turn activates adenylate cyclase, resulting in the production of cAMP, in accordance with GLP- 1 receptor activation. PKA and EPAC are activated by elevated cAMP. Additionally, cAMP and PKA phosphorylate ERK1/2, which controls genes involved in proliferative and anti-apoptotic processes, and activate several proteins, such as the mitogen activated protein kinase (MAPK) cascades [63]. By the same mechanisms (potassium channel closure-mediated membrane depolarization) as the GLP- 1 receptor, the GIP receptor activation of PKA results in the secretion of insulin. Additionally, non-insulinotropic effects like regulating the survival and proliferation of pancreatic beta cells can be facilitated by GIP receptor activation [64, 65]. Furthermore, PKA activation is known to inhibit AMPK in GIP receptor and GLP- 1 receptor signaling, which results in the transfer of TORC2 into the nucleus [66]. The anti-apoptotic gene bcl- 2 is a promoter of transcription in the nucleus by a complex between CREB and TORC2. Additionally, the nuclear transcription factor (Foxo1) is phosphorylated upon activation of phosphoinositide- 3-kinase–protein kinase B/Akt (PI3 K-PKB/Akt), which in turn deactivates molecules to restrict the activity of proapoptotic pathways [66].

Intracellular Signaling Pathways: Interaction and Cooperation in Beta Cells

The signaling pathways in beta cells that are triggered by ligand binding to the GIP receptor and GLP- 1 receptor share a lot of similarities. The PKA and EPAC2 pathways are triggered by both receptors'activation of adenylate cyclase, which raises intracellular cAMP and causes insulin synthesis and release, cellular proliferation, and antiapoptotic effects. Recruitment of beta arrestins desensitizes both receptors, and this is followed by internalization, recycling, and inactivation of Gas. There are also several distinctions. First, beta arrestins and Gaq can both internalize the GLP- 1 receptor, and multiple ligands can affect internalization. On the other hand, GIP receptor internalization depends solely on arrestins and is not easily impacted by new ligands [67]. By inhibiting the Gas proteins and facilitating receptor trafficking through internalization and recycling, these beta arrestins are essential for GIP receptor desensitization. Second, it has been demonstrated that GIP receptor activation in beta cells leads to MAPK-induced signaling pathways, whereas GLP- 1 receptor activation does not. Third, GIP receptor activation has not been shown to cause proliferation and anti-apoptosis; instead, GLP- 1 receptor activation stimulates EGF receptor signaling [20].

GIP and GLP- 1 Signaling in Alpha Cells

It has been reported that GLP- 1 and GIP influence alpha cells. GLP- 1 stimulation also lowers blood sugar levels by inhibiting α cell production of glucagon [68]. A subset of cultured cells expressed GLP- 1 receptor, and mice with a-cell specific GLP- 1 receptor knockout have mild glucose intolerance and higher glucagon secretion in response to glucose challenges than animals of the wild type, which may indicate that GLP- 1 directly affects this cell [69]. It's also possible that GLP- 1 affects cells indirectly. In fact, it has been demonstrated that several molecules secreted by beta cells, such as insulin, zinc, and c-aminobutyric acid, inhibit the secretion of glucagon. These molecules may also theoretically help to inhibit the secretory activity of alpha cells that are dependent on GLP- 1 [70]. Furthermore, it is possible that somatostatin directly inhibits glucagon secretion by binding to its receptor in the alpha cell, since GLP- 1 stimulates islet somatostatin secretion directly through the canonical GLP- 1 receptor, expressed on delta cells [71]. Research on isolated perfused rat pancreas provides evidence in this regard. Treatment with anti-somatostatin antibodies or co-infusion with a somatostatin receptor 2 antagonist eliminated the GLP- 1-mediated suppression of glucagon secretion [71]. The direct action of GIP on its receptor, which is expressed by these cells, is what causes GIP stimulation to increase glucagon secretion by alpha cells in contrast to GLP- 1 [72,73,74]. Functionally, GIP receptor activation improves intracellular Ca2+ concentration, glucagon secretion, and cell depolarization by increasing cAMP/PKA signaling pathways [75, 76]. The intact rat pancreas's GIP perfusion only stimulates glucagon secretion at low glucose levels (4.4 mM) and not at postprandial glucose concentrations (8.9 mM), suggesting that GIP receptor activity in the cell appears to be glucose dependent [18]. In healthy humans, GIP-mediated elevation of circulating glucagon concentrations has also been observed; like in preclinical research, this effect is glucose dependent, occurring only in hypoglycemia [72, 77]. Figure 1 presents the main effects of GLP‐1, GIP and glucagon in different body organs.

Fig. 1

Role of GLP‐1, GIP and glucagon in different body organs. This shows the physiological effects of GLP‐1, GIP and glucagon on different organs and peripheral tissues. Created in BioRender. Anastasiou IA. (2025) https://BioRender.com/ a49a870. Assessed on 28 January 2025

Preclinical data

In a randomized controlled trial, scientists reported the discovery and translational therapeutic effectiveness of a peptide with strong, balanced coagonism at both GLP- 1 and GIP receptors [78]. This unimolecular dual incretin, which was derived from a mixed sequence of GLP- 1 and GIP, showed improved antihyperglycemic and insulinotropic activity compared to selective GLP- 1 agonists. It is noteworthy that this superior effectiveness extended to primates (humans and cynomolgus monkeys) and rodent models of obesity and diabetes, such as db/db (diabetic) mice and Zucker diabetic fatty (ZDF) rat. In addition, a selective GIP agonist showed very low effectiveness in reducing weight, while this co-agonist showed synergism in reducing fat mass in obese rodents. The unimolecular dual incretins were more successful than selective monoagonists and tackled the two basic causes of diabetes: pancreatic insulin insufficiency and obesity-induced insulin resistance. To support less frequent administration, site-specific lipidation or PEGylation was performed to prolong the duration of action of the unimolecular dual incretins. These peptides offer pharmacology comparable to native peptides and improved effectiveness compared to similarly modified selective GLP- 1 agonists. The improvement in pharmacokinetics helped prevent the negative gastrointestinal side effects typical of GLP- 1-based selective agonists by reducing maximum drug exposure and reducing reliance on GLP- 1-mediated pharmacology. The identification and confirmation of a highly effective, balanced dual incretin agonist enables a more physiological approach to treating conditions associated with impaired glucose tolerance [78].

The combination of GLP- 1 and GIP is proposed as a promis