The molecular mechanisms regulating secretion of the orexigenic-glucoregulatory hormone ghrelin remain unclear. Based on qPCR analysis of FACS-purified gastric ghrelin cells, highly expressed and enriched 7TM receptors were comprehensively identified and functionally characterized using in vitro, ex vivo and in vivo methods. Five Gαs-coupled receptors efficiently stimulated ghrelin secretion: as expected the β1-adrenergic, the GIP and the secretin receptors but surprisingly also the composite receptor for the sensory neuropeptide CGRP and the melanocortin 4 receptor. A number of Gαi/o-coupled receptors inhibited ghrelin secretion including somatostatin receptors SSTR1, SSTR2 and SSTR3 and unexpectedly the highly enriched lactate receptor, GPR81. Three other metabolite receptors known to be both Gαi/o- and Gαq/11-coupled all inhibited ghrelin secretion through a pertussis toxin-sensitive Gαi/o pathway: FFAR2 (short chain fatty acid receptor; GPR43), FFAR4 (long chain fatty acid receptor; GPR120) and CasR (calcium sensing receptor). In addition to the common Gα subunits three non-common Gαi/o subunits were highly enriched in ghrelin cells: GαoA, GαoB and Gαz. Inhibition of Gαi/o signaling via ghrelin cell-selective pertussis toxin expression markedly enhanced circulating ghrelin. These 7TM receptors and associated Gα subunits constitute a major part of the molecular machinery directly mediating neuronal and endocrine stimulation versus metabolite and somatostatin inhibition of ghrelin secretion including a series of novel receptor targets not previously identified on the ghrelin cell.

Triglycerides (TGs) are among the most efficacious stimulators of incretin secretion; however, the relative importance of FFA1 (G Protein-coupled Receptor [GPR] 40), FFA4 (GPR120), and GPR119, which all recognize TG metabolites, ie, long-chain fatty acid and 2-monoacylglycerol, respectively, is still unclear. Here, we find all 3 receptors to be highly expressed and highly enriched in fluorescence-activated cell sorting-purified GLP-1 and GIP cells isolated from transgenic reporter mice. In vivo, the TG-induced increase in plasma GIP was significantly reduced in FFA1-deficient mice (to 34%, mean of 4 experiments each with 8-10 animals), in GPR119-deficient mice (to 24%) and in FFA1/FFA4 double deficient mice (to 15%) but not in FFA4-deficient mice. The TG-induced increase in plasma GLP-1 was only significantly reduced in the GPR119-deficient and the FFA1/FFA4 double deficient mice, but not in the FFA1, and FFA4-deficient mice. In mouse colonic crypt cultures the synthetic FFA1 agonists, TAK-875 stimulated GLP-1 secretion to a similar extent as the prototype GLP-1 secretagogue neuromedin C; this, however, only corresponded to approximately half the maximal efficiency of the GPR119 agonist AR231453, whereas the GPR120 agonist Metabolex-209 had no effect. Importantly, when the FFA1 agonist was administered on top of appropriately low doses of the GPR119 agonist, a clear synergistic, ie, more than additive, effect was observed. It is concluded that the 2-monoacylglycerol receptor GPR119 is at least as important as the long-chain fatty acid receptor FFA1 in mediating the TG-induced secretion of incretins and that the 2 receptors act in synergy, whereas FFA4 plays a minor if any role.

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