Fig 1.
GLP-1Rimmunostaining in colon.
GLP-1R positive nerve fibres (Abcam antibody, left panels) and PGP9.5 (right panels) in mucosal sigmoid colon biopsy specimens, from top—control, middle and bottom–IBD, magnification x40. GLP-1R expressing nerve fibres were increased in IBD compared to controls.
Fig 2.
Quantitation of GLP-1R expression in colon.
Bar charts showing an overall increase in expression of nerve fibres expressing GLP-1R in the IBD group by image analysis of GLP-1R (A, B) and PGP9.5 (C) in colonic biopsies (mean ± SEM, Mann Whitney U test). There was no difference in the ratio of GLP-1R to PGP9.5, indicating an overall increase in the neuronal fibres expressing GLP-1R.
Fig 3.
CGRP and GLP-1R immunostaining in colon and DRG.
CGRP neuronal staining (arrowed) in mucosal sigmoid colon biopsy specimens from control (A), IBD (B and C), magnification x40. Bar chart showing image analysis of CGRP in colonic biopsies (mean ± SEM, Mann Whitney U test) (D). GLP-1R (Abcam antibody, E), and PGP9.5 (F), staining in avulsion injury DRG, magnification x20.
Fig 4.
Specificity of GLP-1R immunostaining in DRG tissue.
Serial sections of GLP-1R-immunoreactive DRG immunostained with no peptide (A) or with 1 μg peptide (B), 100 ng peptide (C), 10 ng peptide (D), 1 ng peptide (E) or 100 pg peptide (F); Abcam antibody at a dilution of 1:1500; magnification x40.
Fig 5.
Specificity of GLP-1R immunostaining in colon.
Serial sections of GLP-1R immunoreactivity in sub-mucous plexus of a full thickness normal colon immunostained with no peptide (A) or with 1 μg peptide (B), 100 ng peptide (C), 10 ng peptide (D), 1 ng peptide (E) or 100 pg peptide (F) Abcam antibody at a dilution of 1:1500; magnification x40.
Fig 6.
Co-localization of GLP-1R-like immunoreactivity and β tubulin.
Double staining of GLP-1R (Abcam GLP-1R antibody at a dilution of 1:1000, black) with the neuronal marker β Tubulin (at a dilution of 1:1000, red) in, A, sub-mucous plexus of colon from a patient with IBD and, B, human DRG; magnification x20.
Fig 7.
GLP-1R immunostaining in colon.
GLP-1R (Acris antibody, A,B) and Neurofilaments (C, D) positive nerve fibres in mucosal sigmoid colon biopsy specimens from top—control and middle–IBD, magnification x20. Bar charts showing an overall increase in expression of neuronal fibres expressing GLP-1R in the IBD group (n = 4) by image analysis of GLP-1R (E) and NF (F) in colonic biopsies (mean ± SEM, Mann Whitney U test).
Fig 8.
GLP-1R immunofluorescence in DRG neurons.
Cultured adult rat DRG neurons (A, B, C) showing β tubulin immunofluorescence (green in A), GLP-1R expression (red in B), and the merged image (C). Bar = 200 μm. High power image of cultured human DRG neurons positive for β tubulin (green in D), intense GLP-1R expression (red) in a small neuron (E), and merged image (F). Bar = 20 μm.
Fig 9.
Effect on neurite outgrowth and capsaicin responses.
Representative images showing β tubulin immunofluorescence in rat DRG neurons cultured with vehicle (A, control), with Oxm (B), exendin-4 (C), or GLP-1 (D). Bar = 100 microns. Graph showing significantly increased neurite lengths after 48 h treatment with oxyntomodulin (*p = 0.04), exendin-4 (*p = 0.01) and GLP-1 (*p = 0.04) using Student’s unpaired t-test (E). Graph showing lack of significant effect of acute treatment with incretin hormones on capsaicin sensitivity in adult rat DRG neurons (F).
Fig 10.
Effect of exendin-4 on ATP responses in human DRG neurons.
Sample trace showing responses to ATP in the absence and presence of exendin-4 (A). Graph showing responses to ATP are significantly enhanced in the presence of exendin-4 (*p = 0.02, Student’s unpaired t-test) (B).