The glucose-dependent insulinotropic polypeptide (GIP) regulates body weight and food intake via CNS-GIPR signaling

Abstract

Uncertainty exists as to whether the glucose-dependent insulinotropic polypeptide receptor (GIPR) should be activated or inhibited for the treatment of obesity. Gipr was recently demonstrated in hypothalamic feeding centers, but the physiological relevance of CNS Gipr remains unknown. Here we show that HFD-fed CNS-Gipr KO mice and humanized (h)GIPR knockin mice with CNS-hGIPR deletion show decreased body weight and improved glucose metabolism. In DIO mice, acute central and peripheral administration of acyl-GIP increases cFos neuronal activity in hypothalamic feeding centers, and this coincides with decreased body weight and food intake and improved glucose handling. Chronic central and peripheral administration of acyl-GIP lowers body weight and food intake in wild-type mice, but shows blunted/absent efficacy in CNS-Gipr KO mice. Also, the superior metabolic effect of GLP-1/GIP co-agonism relative to GLP-1 is extinguished in CNS-Gipr KO mice. Our data hence establish a key role of CNS Gipr for control of energy metabolism.

Keywords: CNS; GIP; GIPR CNS KO; body weight; diet-induced obesity; food intake; glucose metabolism; incretin; type 2 diabetes.

Graphical abstract

Figure 1

Mice with CNS deletion of murine Gipr are protected from diet-induced obesity and glucose intolerance (A–E) Body weight (A), body composition at the age of 28 weeks (B and C), food intake (D), and assimilated energy (E) in 42-week-old male C57BL/6J WT and CNS-Gipr KO mice (N = 7–8 mice each group) fed with a high-fat diet (HFD). Food intake and assimilated energy were assessed per cage in double-housed mice. (F–I) Locomotor activity (N = 7–8 mice each group) (F), hypothalamic expression of genes related to food intake (6–7 mice each group) (G), and total energy expenditure (H) and resting metabolic rate (I) in 29-week-old male mice (N = 7–8 mice each group). (J) Expression of genes related to BAT thermogenesis in HFD-fed male mice (N = 8 each genotype). (K) Respiratory exchange ratio (RER) in 29-week-old male mice (N = 7–8 mice each group). (L–O) Plasma levels of triglycerides (L) and cholesterol (M) (N = 6–7 each group) and intraperitoneal glucose tolerance (N and O) (N = 6–8 mice each group) in 42-week-old male mice. (P) HbA1c (N = 18 mice each group; p = 0.0033). (Q and R) Fasting levels of blood glucose (Q) and insulin (R) as well as HOMA-IR (S) in 42-week-old male mice (N = 7–8 each group). (T and U) Relative expression of Gipr (corrected to housekeeping gene peptidylprolyl isomerase B; Ppib) in the hypothalamus (N = 8 mice each genotype) (T) and in isolated islets from WT and CNS-Gipr KO mice (N = 3 each group) (U). (V and W) Glucose-stimulated insulin secretion (GSIS) in isolated islets under conditions of low (2.8 mM) and high glucose (16.8 mM) (V) and GSIS of isolated islets treated with or without 10 nM of either acyl-GLP-1 or acyl-GIP (W) (N = 4 mice each group). y axis in (W) represents the ratio of secreted insulin stimulated with high glucose (16.8 mM) to low glucose (2.8 mM). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A and N) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs (B–G, J–M, and O–W) were analyzed using two-tailed, two-sided t test. Data in (H) and (I) were analyzed using ANCOVA with body weight as co-variate.

Figure 2

Humanized (h)GIPR knockin mice with conditional CNS-specific hGIPR deletion are protected from diet-induced obesity and glucose intolerance (A–D) Body weight (A), body composition (B and C), and food intake (D) in male C57BL/6N WT and CNS-hGIPR KO mice (N = 6–8 mice each group). (E) Hypothalamic expression of proopiomelanocortin (Pomc), brain-derived neurotrophic factor (Bdnf), cocaine-and-amphetamine-regulated transcript (Cart), agouti-related peptide (Agrp), and neuropeptide y (Npy) in 20-week-old male mice (N = 6–7 mice each group). (F and G) Energy expenditure (F) and locomotor activity (G) in 20-week-old male mice (N = 6 mice each group). (H–P) Intraperitoneal glucose tolerance (H) and fasting levels of blood glucose (I), insulin (J), GLP-1 (K), leptin (L), triglycerides (M), free fatty acids (N), GIP (O), and cholesterol (P) in WT and CNS-hGIPR KO mice (N = 6–8 mice each group). (Q) H&E staining of hepatic lipid accumulation (scale bar represents 100 μm). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A and H) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs (B–E and G–P) were analyzed using two-tailed, two-sided t test. Data in (F) were analyzed using ANCOVA with body weight as co-variate.

Figure 3

Acute central administration of acyl-GIP improves body weight, food intake, and glycemia in DIO mice (A–E) Body weight change (A), food intake (B and C), and plasma levels of blood glucose (D and E) in male DIO mice treated centrally (i.c.v.) with a single dose of 1, 3, or 6 nmol acyl-GIP (N = 7–8 mice each genotype). (F–I) Ad libitum plasma levels of insulin (F) and c-peptide (G) and plasma levels of triglycerides (H) and free fatty acids (I) in 32-week-old DIO mice (N = 6–8 each group). (J and K) cFOS immunofluorescence (J) and cFOS quantification (N = 6–7 mice each genotype) (K) in the hypothalamic arcuate nucleus (ARC) of DIO mice treated with acyl-GIP. Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Scale bar, 100 μm. Longitudinal data (A, B, and D) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs in (C), (E)–(I), and (K) were analyzed using one-way ANOVA.

Figure 4

Chronic central administration of acyl-GIP improves body weight, food intake, and glycemia in HFD-fed WT mice, but not in CNS-Gipr KO mice (A–C) Body weight (A), eWAT weight (B), and food intake (C) of HFD-fed mice treated with acyl-GIP (0.02 or 0.04 nmol/day) or liraglutide (0.04 nmol/day) or that were pair-fed to the acyl-GIP (0.04 nmol/day)-treated mice (N = 9–10 each group). (D–G) Fasting plasma levels of blood glucose (D), insulin (E), leptin (F), and HOMA-IR (G) after 14 days of treatment (N = 7–10 mice each group). (H) Expression of Gipr in iWAT, eWAT, and hypothalamus after 14 days of treatment (N = 7–10 mice each group). (I–K) Body weight change (I), food intake (J), and fasting blood glucose (K) in HFD-fed WT and CNS-Gipr KO mice following treatment with 0.02 nmol/day of acyl-GIP (N = 9–10 mice each genotype). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A, C, I, and J) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs in (B), (D)–(H), and (K) were analyzed using one-way ANOVA.

Figure 5

Peripheral administration of acyl-GIP decreases food intake and activates cFOS in the hypothalamic ARC and VMH in DIO mice (A–C) Body weight (A) and acute (B) and chronic (C) effects of peripherally (s.c.) administered acyl-GIP (30 nmol/kg/day) on food intake in 21-week-old male DIO mice (N = 8 mice each group). (D–I) Meal size (D) and frequency (E), acute acyl-GIP effects on fatty acid oxidation (F), respiratory exchange ratio (RER) (G), and acute and chronic effects of acyl-GIP on energy expenditure (H and I) in 21-week-old male DIO mice (N = 8 mice each group). (J and K) Assimilated energy (J) and assimilation efficiency (K) in mice chronically treated daily s.c. for 7 days with acyl-GIP (N = 8 mice each group). (L–O) Staining and quantification of cFOS in the ARC (L), DMH (M), and VMH (N) and cFOS/NPY co-staining (O) in the ARC of 19-week-old male HFD-fed NPY-GFP mice treated with a single peripheral (s.c.) injection of either vehicle or acyl-GIP (30 nmol/kg) (N = 5 mice each group; scale bar, 100 μm). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A–C, F, and H) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs in (E), (G), and (J)–(O) were analyzed using two-tailed, two-sided t test. Data in (I) was analyzed using ANCOVA with body weight as co-variate.

Figure 6

Chronic peripheral administration of acyl-GIP improves body weight, food intake, and glycemia via CNS-GIPR signaling (A–D) Body weight change (A), placebo-corrected total body weight loss (B), and food intake (C and D) of HFD-fed WT and CNS-Gipr KO mice treated with 100 nmol/kg/day of acyl-GIP (N = 8 mice each group). (E–H) Body weight change (E), placebo-corrected total body weight loss (F), and food intake (G and H) of HFD-fed WT and global Gipr KO mice treated with 100 nmol/kg/day of acyl-GIP (N = 12–13 mice each group). (I–L) Body weight change (I), placebo-corrected total body weight loss (J), and food intake (K and L) of HFD-fed WT and global GLP-1R KO mice treated with 100 nmol/kg/day of acyl-GIP (N = 6–8 mice each group). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A, C–E, G–I, K, and L) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs in (B), (F), and (J) were analyzed using one-way ANOVA.

Figure 7

GLP-1/GIP loses superior potency over GLP-1 upon chronic peripheral treatment in CNS-Gipr KO mice (A–D) Change in body weight (A), fat mass (B), lean mass (C), and food intake (D) of HFD-fed WT and CNS-Gipr KO mice treated with acyl-GLP-1 or GLP-1/GIP (MAR709) at a dose of 10 nmol/kg/day (N = 7–8 mice each group). (E and F) Intraperitoneal glucose tolerance after 12 days of treatment (N = 7–8 mice each group). Data represent means ± SEM. ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. Longitudinal data (A and E) were analyzed using two-way ANOVA with time and genotype as co-variables and Bonferroni post hoc analysis for individual time points. Bar graphs in (B)–(D) and (F) were analyzed using one-way ANOVA.