228-LB: ß-arrestin-2 Deletion Influences GLP-1 Receptor Signaling in Pancreatic ß Cells In Vivo

Abstract

β-arrestin-2 (Barr2) is typically associated with G protein-coupled receptor desensitization and endocytosis, and reduced Barr2 recruitment is thought to enable prolongation of pharmacodynamic effects of biased glucagon-like peptide-1 receptor (GLP-1R) agonists. However, the precise role of Barr2 in GLP-1R function in β-cells is poorly understood. To study this role, a tamoxifen-inducible β-cell specific Barr2 knockout mouse model was generated (Pdx1CreERT/Barr2 fl/fl, referred to as β-Barr2 KO). β-Barr2 KO animals on high-fat high-sucrose diet were less glucose tolerant and tended to respond worse to acute exendin-4 (Ex4) treatment vs. controls (Barr2 fl/fl). However, Ex4 effects on blood glucose were more pronounced in KO mice 6h post-agonist injection [IPGTT AUC (6h) for Ex4/ vehicle: 0.84 ± 0.04 vs. 1.08 ± 0.05, p<0.001; ΔAUC (6h-0h): -49.8 ± 65.8 vs. 158.2 ± 43.6, p<0.05; KO vs. control]. This effect was not apparent with Ex-phe1, an Ex4 derivative biased away from Barr2 recruitment. Ex4-induced potentiation of insulin secretion in β-Barr2 KO islets was reduced acutely, but tended to increase overnight, resulting in significantly reduced desensitization vs. control islets. On chow diet, acute Ex4-stimulated calcium increases were unaffected, but acute cAMP responses were attenuated in KO islets [Emax (vs. baseline): 1.51 ± 0.17 vs. 1.72 ± 0.07, p<0.05; KO vs. control]. The latter was corroborated in INS1 832/3 Barr2 KO CRISPR cells, as NanoBiT™ functional assays showed reduced Ex4-induced Gs protein activation vs. control. GLP-1R trafficking profiles observed in vivo by imaging of Ex4-TMR uptake into islets implanted in the anterior chamber of the eye, as well as in vitro with GLP-1R-NLuc - KRAS/Rab5 NanoBRET assays were unaltered by Barr2 deletion. Thus, we provide evidence for opposing effects of Barr2 in acute vs. prolonged GLP-1R signaling, supporting a direct role for Barr2 in mediating the anti-hyperglycemic effects of biased GLP-1R agonists, but dispensability for GLP-1R trafficking. Disclosure S. Bitsi: None. K. Suba: None. N. Mohamed: None. I. Leclerc: Consultant; Spouse/Partner; Sun Pharmaceutical Industries Ltd. G. A. Rutter: Advisory Panel; Self; Sun Pharmaceutical Industries Ltd. V. Salem: None. B. Jones: Research Support; Self; Sun Pharmaceutical Industries Ltd. A. Tomas: Other Relationship; Self; Sun Pharmaceutical Industries Ltd. Funding Medical Research Council (MR/R010676/1)