Stimulating intestinal GIP release reduces food intake and body weight in mice

Abstract

Objective: Glucose dependent insulinotropic polypeptide (GIP) is well established as an incretin hormone, boosting glucose-dependent insulin secretion. However, whilst anorectic actions of its sister-incretin glucagon-like peptide-1 (GLP-1) are well established, a physiological role for GIP in appetite regulation is controversial, despite the superior weight loss seen in preclinical models and humans with GLP-1/GIP dual receptor agonists compared with GLP-1R agonism alone.

Methods: We generated a mouse model in which GIP expressing K-cells can be activated through hM3Dq Designer Receptor Activated by Designer Drugs (DREADD, GIP-Dq) to explore physiological actions of intestinally-released GIP.

Results: In lean mice, Dq-stimulation of GIP expressing cells increased plasma GIP to levels similar to those found postprandially. The increase in GIP was associated with improved glucose tolerance, as expected, but also triggered an unexpected robust inhibition of food intake. Validating that this represented a response to intestinally-released GIP, the suppression of food intake was prevented by injecting mice peripherally or centrally with antagonistic GIPR-antibodies, and was reproduced in an intersectional model utilising Gip-Cre/Villin-Flp to limit Dq transgene expression to K-cells in the intestinal epithelium. The effects of GIP cell activation were maintained in diet induced obese mice, in which chronic K-cell activation reduced food intake and attenuated body weight gain.

Conclusions: These studies establish a physiological gut-brain GIP-axis regulating food intake in mice, adding to the multi-faceted metabolic effects of GIP which need to be taken into account when developing GIPR-targeted therapies for obesity and diabetes.

Keywords: Diabetes; Enteroendocrine cells; Feeding behaviour; GIP; GLP-1; Obesity.

Figure 1

GIP-Cre::hM3Dq activation significantly improves glucose tolerance. (A) Schematic for the GIP-Dq mouse model. (B) Representative section from the small intestine of GIP-Dq mice demonstrating Dq (green) and DAPI (white) expression. Scale bar – 100um. (C) Plasma GIP of WT mice receiving oral gavage of liquid Ensure, a mixed meal. (D–G) Plasma (D) GIP, (E) insulin, (F) mPYY and (G) total GLP-1 (TGLP-1) of GIP-Dq mice treated with CNO (1 mg/kg BW ip). (H) ipgtt (2 g/kg BW glucose, admin of VEH or CNO (at 1 mg/kg ip, delivered contralaterally to glucose at time 0) with AUC (inset) (n = 16–24 per group). (I–K) Plasma (I) GIP (one-way ANOVA: effect of treatment F_(3,24) = 133.3, p < 0.0001. Post hoc p < 0.0001), (J) insulin (effect of treatment F_(3,20) = 10.95, p < 0.0001. Post hoc p = 0.0017) and (K) TGLP-1 (effect of treatment F_(3,24) = 1.275, p = 0.3053) at basal and +15 mins post glucose (as previous). Animals were subsequently pre-treated with a GIPR monoclonal antibody antagonist or isotype control antibody 48 h prior to (L) ipgtt (as previous) and (M) AUC (n = 7 per group, effect of treatment F_(3,24) = 38.73, p < 0.0001. Post hoc p = 0.0001). Values are presented as group mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001 and ∗∗∗∗p < 0.0001 by paired students T test (C-H[inset]) and one-way ANOVA (I-K, M[inset]).

Figure 2

GIP-Cre::hM3Dq activation significantly reduces food intake. (A) Food intake of GIP-Dq mice 1hr post ipgtt (n = 15 per treatment). (B) Overnight fast (16 h)-refeeding(1hr) food intake of GIP-Dq mice treated with VEH/CNO (n = 12–15 per treatment). (C) Food intake of ad lid fed GIP-Dq mice treated with VEH/CNO (at 1 mg/kg BW ip) at the onset of the dark phase (n = 5–6 per treatment, treatment F_(1,8) = 20.66, p = 0.0042, time F_{(1.628, 13.03)} = 180.6, p < 0.0001, interaction F_(2,16) = 28.31, p < 0.0001). (D) HPM intake of ad lib fed mice treated with CNO at the onset of the dark phase (n = 8 per treatment). (E) Food intake (interaction F_(2,50) = 5.578, p = 0.0065. Post hoc p = 0.001), (F) Total meal duration (treatment F_(1,26) = 6.007), p = 0.0213, time F_{(1.933,50.27)} = 4.181, p = 0.022. Post hoc p = 0.0225 (G) Inter-meal interval (interaction F_(2,76) = 5.269, p = 0.0072, time F_{(1.659,63.02)} = 16.88, p < 0.0001). Post hoc p = 0.0001), (H) Cumulative food intake (treatment F_(1,25)] = 4.775, p = 0.0385, time F_{(2.093,52.33)} = 178.6, p < 0.0001, (I) RER (interaction F_(24,610) = 2.033, p = 0.0027, time F(_6.816,173.2) = 42.37, p < 0.0001), (J,K) Energy expenditure (time F_{(9.234,234.7)} = 25.67, p < 0.0001), (L) Ambulatory activity (time F_{(7.607, 193.3)} = 13.30, p < 0.0001) and (M) body weight change of ad lib fed GIP-Dq mice treated with VEH/CNO at the onset of the dark phase (n = 14 per treatment, interaction F_(1,26) = 4.242, p = 0.0496. Post hoc p = 0.0189). Values are presented as group mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 by paired (A, D) and students T test (B) and two-way ANOVA (C, E-J, L,M) and ANCOVA (body weight as covariate, K).

Figure 3

Central GIPR antagonism abolishes the effect on food intake but not glucose tolerance. Representative cfos staining of the ARC, AP and NTS of (A) GIP-Dq mice treated with VEH/CNO (at 1 mg/kg BW ip, n = 6 per group) and (B) WT/GIPR-Cre::GCaMP3 mice treated with VEH/[D-Ala2]-GIP; scale bars represent 100 μm and numbers at the top refer to Bregma. (C,D), quantification of conditions shown in a and b, respectively showing average c-Fos positive cells per section. Each point represents data from an individual mouse. (E) Food intake (at the onset of the dark phase) of ad lib fed GIP-Dq mice receiving ICV pre-treatment with IsoAb/GIPR Ab antagonist (n = 4–5 per group, interaction F(_6,26) = 7.174, p = 0.0001, time F_{(1.116,14.51)} = 137.5, p < 0.0001, treatment F_(1,13) = 9.690, p = 0.0013. Post hoc p = 0.030). (F) Food intake in fast/refeed paradigm (n = 8–11 per group, treatment F_(3,30) = 8.765, p = 0.0003. Post hoc p = 0.0019). (G) ipgtt (as previous) and (H) AUC (n = 3–6 per group, treatment F_(3,14) = 15.77, p < 0.0001. Post hoc p = 0.0079 and p = 0.0054 respectively). Values are presented as group mean ± SEM. ∗∗p < 0.01 by Students T test (C and D), two-way ANOVA (E) and one-way ANOVA (F, H).

Figure 4

The effects of GIP-Cre::hM3Dq activation are maintained in DIO mice. (A) ipgtt (2 g/kg BW glucose, admin of VEH or CNO (at 1 mg/kg ip, delivered contralaterally to glucose at time 0) with AUC (inset) (n = 8 per group). (B) Fast-refeed (as previous) food intake of GIP-Dq mice following treatment with VEH/CNO (at 1 mg/kg BW ip, n = 7–8 per group). (C) Blood glucose in ad lib fed state following treatment with VEH/CNO (n = 7–8 per group). (D) Food intake (interaction F_(2,56) = 6.432, p = 0.0031. Post hoc p = 0.0221), (E) Total meal duration (F) Inter-meal interval (treatment F_(1,28) = 4.478, p = 0.0434. Post hoc p = 0.0436), (G) Cumulative food intake (interaction F_(12,336) = 2.026, p = 0.0215), (H) RER (time F_(6.486,147) = 4.453, p = 0.0002), (I) Ambulatory activity (time F_{(5.666,132.7)} = 7.531, p < 0.0001) and (J,K) Energy expenditure (time F_{(4,692,109.5)} = 21.30, p < 0.0001) of GIP-Dq mice treated with VEH/CNO at the onset of the dark phase in ad lib fed animals (n = 15 per treatment). (L) Plasma GIP, (M) Body weight (interaction F_(7,91) = 6.117 p < 0.0001), (N) Body weight change (interaction F_(7,78) = 2.435, p = 0.0261, treatment F_(1,13) = 23.30, p = 0.0003, time F_{(2.596, 28.93)} = 10.74, p = 0.0001) and (O) Cumulative food intake (treatment F_(1,13) = 25.65, p = 0.0002), time F_{(1.607,20.89)} = 443.8, p < 0.0001) of GIP-Dq mice treated chronically with DCZ via drinking water (n = 7–8 per group). Values are presented as group mean ± SEM. ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001 by Students T test (A,B(inset)C, D) and two-way ANOVA (D-J, L-N) and ANCOVA (body weight as covariate, K).