Unravelling the mechanisms by which GLP-1 modulates glucagon secretion

The secretion of glucagon by pancreatic alpha-cells plays a critical role in the regulation of glycaemia1. This endocrine hormone is essential to prevent hypoglycaemia. Generally, it opposes the actions of insulin by promoting hepatic glucose production, thereby increasing blood glucose concentrations. Despite the importance of glucagon, very little is known about the precise regulatory machinery that controls its secretion from alpha-cells.

It has been established that, in addition to glucose, a number of both endocrine and paracrine hormones play a role in glucagon secretion. For example, insulin, somatostatin and γ-aminobutyric (GABA) all decrease glucagon exocytosis while the incretin hormone, gastric inhibitory polypeptide (GIP), increases its release2,3. However, the actions of the mature form of GLP-1 [GLP-1 (7-36) amide] are far from clear, since its ability to inhibit glucagon release in vivo contrasts with its effects in single alpha-cells. Likewise, our preliminary data indicate that GLP-1 (9-36) amide, the inactivated form of GLP-1 generated following degradation by dipeptidyl peptidase IV (DPP-IV), is also able to reduce secretion of glucagon, with high potency, in a cAMP-independent manner. In contrast, exenatide, a synthetic and stable mimetic of GLP-1 (7-36) amide has been separately documented to both increase and decrease glucagon secretion from mouse alpha-cells4.

Moreover, there are reports that glucagon has both positive (feed-forward) and negative (feed-back) effects upon its own secretion5,6. Finally, and somewhat intriguingly, expression levels of both the glucagon receptor (GCGR) and the GLP-1R found within alpha-cells are between 30-100 fold lower than those detected in pancreatic beta-cells7. Recent work from our laboratory has shown that GLP-1 (7-36) amide as well as GLP-1 (9-36) can strongly supress glucagon secretion from pancreatic islets isolated from GLP-1 receptor -/- ’knockout’ mice; This implies that GLP-1 utilizes a novel mechanism for modulating glucagon secretion. Given these seemingly conflicting reports, and a lack of a clear definitive mechanism, the aim of this project is to determine how GLP-1 regulates the secretion of glucagon in a GLP-1R-independent manner.

The complex nature of islets means that many of the components for modulating glucagon secretion are also found in beta-cells and delta-cells. Therefore simpler model systems to enable pharmacological characterization of candidate receptors for GLP-1 binding are required. Our initial receptors of choice are the GCGR and the GIPR since recent reports suggest that both exhibit a preference for binding GLP-1 (7-36) amide over exenatide. Further preliminary data suggests that administration of a GCGR antagonist blocks GLP-1 (9-36) amide-mediated glucagon secretion from human islets. GPCR pharmacology will be studied as part of an ongoing collaboration with Dr Graham Ladds at the University of Cambridge and islet physiological evaluation will be undertaken at the Oxford Centre for Diabetes, Endocrinology and Metabolism.

The specific objectives of the proposed work are as follows:
1.Pharmacological characterization of the GCGR and GIPR as targets for GLP-1 activity;
2.Evaluation of other GPCRs in modulating glucagon secretion;
3.Analysis of the GPCR-based signaling components found in alpha-cells – including GPCRs, G-proteins and RAMPs.
4.Evaluation of the role of somatostatin in modulating glucagon secretion. This work is essential since somatostatin receptors are predominately Gαi-coupled in alpha-cells.
5.Assessment of receptor physiology in islets by knockdown of GCGR, GIPR and GLP-1R to pharmacologically and physiologically characterise their responses to both GLP-1 (7-36) amide and GLP-1 (9-36) amide.

These experiments will establish the mechanisms by which both GLP-1 (7-36) amide and GLP-1 (9-36) amide modulate glucagon secretion. In the long-term, the insights gained here will also facilitate the design of more efficacious drugs aimed at regulating glycaemia.