RGS4 is a negative regulator of insulin release from pancreatic β-cells in vitro and in vivo

Abstract

Therapeutic strategies that augment insulin release from pancreatic β-cells are considered beneficial in the treatment of type 2 diabetes. We previously demonstrated that activation of β-cell M 3 muscarinic receptors (M3Rs) greatly promotes glucose-stimulated insulin secretion (GSIS), suggesting that strategies aimed at enhancing signaling through β-cell M3Rs may become therapeutically useful. M3R activation leads to the stimulation of G proteins of the G q family, which are under the inhibitory control of proteins known as regulators of G protein signaling (RGS proteins). At present, it remains unknown whether RGS proteins play a role in regulating insulin release. To address this issue, we initially demonstrated that MIN6 insulinoma cells express functional M3Rs and that RGS4 was by far the most abundant RGS protein expressed by these cells. Strikingly, siRNA-mediated knockdown of RGS4 expression in MIN6 cells greatly enhanced M3R-mediated augmentation of GSIS and calcium release. We obtained similar findings using pancreatic islets prepared from RGS4-deficient mice. Interestingly, RGS4 deficiency had little effect on insulin release caused by activation of other β-cell GPCRs. Finally, treatment of mutant mice selectively lacking RGS4 in pancreatic β-cells with a muscarinic agonist (bethanechol) led to significantly increased plasma insulin and reduced blood glucose levels, as compared to control littermates. Studies with β-cell-specific M3R knockout mice showed that these responses were mediated by β-cell M3Rs. These findings indicate that RGS4 is a potent negative regulator of M3R function in pancreatic β-cells, suggesting that RGS4 may represent a potential target to promote insulin release for therapeutic purposes. knockout mice | muscarinic receptor | RGS proteins | G protein-coupled receptor T ype 2 diabetes (T2D) has emerged as a major threat to human health worldwide. Besides peripheral insulin resistance, T2D is usually associated with β-cell dysfunction (1). Thus, the development of new drugs aimed at improving β-cell function, including stimulation of insulin release, is the focus of many laboratories (2).β-cell function is regulated by many hormones and neurotransmitters most of which act on specific G protein-coupled receptors (GPCRs) that are expressed on the surface of pancreatic β-cells (3, 4). Following ligand-induced activation, a specific GPCR interacts with and activates one or more classes of heterotrimeric G proteins (consisting of α, β, and γ subunits), which in turn modulate various intracellular signal transduction pathways (5).Stimulation of the muscarinic cholinergic (parasympathetic) nerves innervating the endocrine pancreas leads to a pronounced increase in glucose-stimulated insulin secretion (GSIS) (4, 6). Studies with pancreatic islets prepared from M 3 muscarinic acetylcholine receptor (M3R) KO mice demonstrated that muscarinic enhancement of GSIS is mediated by the M3R subtype (7,8). The M3R is a member of the muscarinic receptor family (M 1 -M 5 ) that is selectively coupled ...

Muscarinic Control of MIN6 Pancreatic β Cells Is Enhanced by Impaired Amino Acid Signaling

“…RGS4 terminates signaling from M3Rs, thereby inhibiting their function in MIN6 cells and primary mouse islets (7). It seemed possible that RGS4 might have been suppressed in T1R3 knockdown cells to increase M3R function, but experimental findings did not support this idea.…”

“…The M3 mAChR (M3R) is the predominant receptor subtype expressed in ␤ cells and insulin-secreting cell lines (6,7). Parasympathetic nerve endings that innervate the pancreas release acetylcholine during the preabsorptive and absorptive phases of feeding (8) to activate this receptor.…”

Chronic activation of a designer Gq-coupled receptor improves β cell function

“…Drug-induced increases in intracellular calcium levels were determined in 96-well plates using FLIPR technology (Molecular Devices) essentially as described (41).…”

G Protein-Coupled Receptor Signaling and Sphingosine-1-Phosphate Play a Phylogenetically Conserved Role in Endocrine Pancreas Morphogenesis

“…Rgs4 null mutants are viable and fertile (2,18,48), but Rgs4 exerts key functions in adult mouse physiology by regulating fatty acid and glucose homeostasis, parasympathetic signaling in heart rate control (2,18), and insulin release from ␤ cells (48). Since Rgs4 expression was not detected in endocrine cells at any developmental time point until and including P1, the expression and role of Rgs4 in the adult pancreas must be attributed to reactivation in mature ␤ cells during postnatal pancreas maturation.…”

“…RGS proteins are exclusive components of G protein-coupled receptor (GPCR) signaling and exert their effects by enhancing the intrinsic GTPase activity of activated GTP-bound G␣ subunits, thereby decreasing the duration of GPCR signaling in diverse processes (47). In the mature pancreas, GPCR signaling plays an important role in the regulation of normal ␤-cell function (43,48), and there is some evidence implicating it in cell fate specification during pancreas development (40,41).…”