Proceedings of The Physiological Society
University of Oxford (2008) Proc Physiol Soc 12, C6 and PC16
Mechanisms of GIP secretion from primary intestinal K-cells
H. E. Parker1, A. M. Habib1, G. J. Rogers1, F. M. Gribble1, F. Reimann1
1. Cambridge Institute of Medical Research, Cambridge, United Kingdom.
Glucose-dependent insulinotropic polypeptide (GIP) is an incretin hormone that promotes insulin release and coordinates the fate of dietary fat . It is released from K-cells located in the duodenal and jejunal epithelia, which are thought to directly sense the presence of nutrients in the gut lumen. However, at the cellular and molecular level, little is known about how K-cells respond to stimuli. The aim of this study was to generate transgenic mice with fluorescently labelled K-cells and to use these to investigate pathways by which K-cells detect nutrients. Transgenic mice were generated in which the GIP promoter drives expression of a fluorescent protein, Venus. Fluorescent cells from intestinal tissue were purified by flow cytometry and analysed by quantitative RT-PCR. GIP secretion in response to various stimuli was assayed in cultures of adult mouse small intestine. Our data establish for the first time that K-cells express glucose-sensing proteins, Kir6.2, SUR1, glucokinase and SGLT1, at levels 190-fold (p=0.005), 470-fold (p=0.0002) and 420-fold (p=0.007) greater than control cells respectively, but barely detectable levels of sweet taste receptor subunits. Expression of the lipid receptors, GPR40, GPR119 and GPR120, postulated to play a role in the regulation of enteroendocrine hormone release, was also enhanced in the K-cell population, 99-fold (p=0.007), 330-fold (p=0.0009) and 76-fold (p=0.002), respectively. In primary cultures, GIP secretion was enhanced 1.34-fold by 10mM glucose (n=27, p=0.003), 1.25-fold by 500μM tolbutamide (n=19, p=0.01) and 1.21-fold by 10mM fructose (n=9, p=0.03) confirming that KATP channels in K-cells are functional. Secretion was also enhanced 2.0-fold by PMA (n=3, p=0.005) and 3.8-fold by a combination of forskolin/IBMX (n=44, p=10-13) highlighting an important role for pathways coupled to the activation of adenylate cyclase and/or PKC. In the presence of forskolin/IBMX, we observed a 1.7-fold stimulation of GIP release by 10mM α-methylglucose (αMG; n=18, p=0.0001), demonstrating that glucose metabolism is not required for monosaccharide triggered secretion. The measured ED50 for glucose of 0.6mM is close to the Km for glucose and αMG of SGLT1 (~0.3mM ), suggesting that SGLT1 plays a key role in glucose-triggered GIP release. Forskolin/IBMX potentiated the response to 10mM glucose by 3.1-fold (n=17, p=2x10-6) indicating a synergistic interaction between these two pathways. GIP release was unaffected by the artificial sweetener, 1mM sucralose. In conclusion, glucose-dependent GIP secretion was found to be SGLT1-dependent, and modulated by KATP channels but not determined by sweet taste receptors. Synergistic stimulation by elevated cAMP and glucose suggests that targeting appropriate G-protein coupled receptors may provide opportunities to modulate GIP release in vivo.
Where applicable, experiments conform with Society ethical requirements