Insulin is the only hormone able to lower blood glucose levels. In order to maintain glucose homeostasis, insulin secretion from β-cells of pancreatic islets is subject to a well-coordinated regulation. Secretion of insulin is primarily controlled by the entry of glucose into β-cells. However, glucose-stimulated insulin secretion (GSIS) is modulated by multiple factors including mediators of the autonomous nervous system as well as various hormones and nutrients. Most of these molecules act through G-protein-coupled receptors expressed by β-cells. Among them is FFAR1, a G q/G 11-coupled receptor which shows high expression in β-cells and can be activated by various long- and medium-chain free fatty acids (FFAs). Shortly after the discovery of FFAR1, synthetic agonists of the receptor have been developed which promote GSIS, and FFAR1 is now an established target for drugs used to increase insulin secretion in type-2 diabetes.
Despite the role of FFAR1 as a therapeutic drug target and the rapid development of synthetic FFAR1 agonists, the physiological function of FFAR1 in regulating insulin secretion from β-cells remains unclear. In transgenic mice with β-cell specific overexpression of FFAR1, acute stimulation of insulin secretion by a bolus injection of glucose was increased whereas it was impaired in Ffar1-deficient mice kept on a high-fat diet. Insulin secretion in response to a sustained glucose stimulation in hyperglycemic clamp experiments was even more strongly reduced in Ffar1-deficient mice both under fasted and fed conditions. These data indicate that FFAR1 plays an important role in the regulation of insulin secretion by glucose under physiological and pathological conditions, but it is unclear which physiological ligands are responsible for FFAR1-dependent regulation of insulin secretion.
Based on earlier data, which showed that the artificial lowering of FFA plasma levels resulted in reduced GSIS, whereas elevation of plasma FFA levels increased GSIS, it was assumed that FFAR1 functions as a receptor for dietary FFAs. This notion was supported by the observation that in vivo infusion of dietary FFAs as well as in vitro exposure of pancreatic islets to palmitate potentiated GSIS, and these effects were reduced by about 50% in the absence of FFAR1, while the remaining half of this GSIS-promoting effect of FFAs appears to require FFA metabolism to long-chain co-enzyme A esters. Since dietary FFAs were always added exogenously, it remains unclear whether dietary FFAs regulate insulin secretion through FFAR1 under physiological and pathophysiological conditions. In addition, dietary FFAs are strongly bound to plasma proteins, and their free circulating levels appear to be rather low to activate FFAR1. Finally, physiological variations of plasma FFA levels, which are primarily the result of adipocyte lipolytic activity, are rather inversely correlated with insulin secretion as lipolysis and FFA plasma levels decrease postprandially when insulin levels go up while their levels increase during starvation when insulin levels are low.
We therefore searched for other endogenous fatty acid ligands of FFAR1 which promote insulin secretion through FFAR1 under physiological conditions. Here we show that glucose promotes the formation of 20-hydroxyeicosatetraenoic acid (20-HETE) in pancreatic islets and that 20-HETE activates FFAR1 with higher efficacy than dietary FFAs. In addition, we provide evidence that 20-HETE acting through FFAR1 mediates a positive feedback regulation during GSIS, a mechanism which is impaired under type-2-diabetic conditions in mice and humans.
In a screen for lipid mediators which are able to activate FFAR1, we found that 20-HETE was significantly more efficacious than dietary FFAs including linoleic and pinolenic acid as well as palmitic and α-linolenic acid in activating FFAR1 when tested for their ability to induce FFAR1-mediated induction of Ca 2+ transients in intact cells. Very similar data were obtained when FFAR1-mediated increases in GTPγS binding were determined in plasma membranes. 20-HETE was also slightly more potent than dietary FFAs with regard to the induction of FFAR1-mediated effects. In contrast to 20-HETE, arachidonic acid was less active, and 18-HETE as well as 19-HETE showed no activity on FFAR1. The FFAR1 antagonist GW1100 concentration-dependently inhibited α-linolenic acid as well as 20-HETE-induced elevation in [Ca 2+]i in cells transfected with FFAR1, confirming that the effect of 20-HETE is mediated by the receptor. In addition, 20-HETE was able to displace 3 H-palmitic acid from the FFAR1 receptor expressed in COS-1 cells whereas 18- and 19-HETE, which were inactive in cellular FFAR1 activation assays, were unable to compete with 3 H-palmitic acid. Recent data indicate that FFAR1 can not only signal through G q/G 11 but also through G s and β-arrestin, depending on the ligand. Similar to dietary FFAs, 20-HETE acting through FFAR1 did not lead to activation of G s but was able to recruit β-arrestin, although with relatively low potency. Interestingly, 20-HETE at 30 μM had no activity on FFAR4 (GPR120), a FFA receptor activated by dietary long-chain fatty acids. We conclude that 20-HETE is a specific orthosteric agonist of FFAR1, which shows significantly increased efficacy compared to other long-chain fatty acids described as FFAR1 agonists so far.
