Expression of Putative Umami/L‐Amino Acid Receptors in Mouse Intestine and Colon, and a Murine Enteroendocrine Cell Line

Abstract

The ability of the gastrointestinal tract to adequately regulate motility and secretion is dependent on the interplay of both sensory and motor system. Chemosensation of nutrients in luminal contents is critical to this process and is largely dependent on activation of receptors present on enteroendocrine cells. G‐protein coupled nutrient receptors for free fatty acids, carbohydrates, protein and proteolytic digestion products as well as receptors for other components of luminal contents such as bile acids have been identified in the gut and postulated to play a role in chemosensation. Several receptors have been postulated to play a role in the sensing of L‐amino acids (L‐AA) in the gut as well as the initiation of umami taste sensation in taste buds. These include the metabotropic glutamate receptors 1 and 4 (mGluR1 and mGluR4), the calcium‐sensing receptor (CaR), the GPRC family C group 6 subtype A receptor (GPRC6A), and the taste receptor heterodimer T1R1/T1R3.AimTo test the hypothesis that the putative L‐AA receptors CaR, mGluR1, mGluR4, and T1R1 are differentially expressed in mouse intestine, colon, and a murine enteroendocrine cell line, the STC‐1 cells.MethodsIntestine and colon were removed from mice, and separated into the mucosal layer containing enteroendocrine cells (EEC) and the smooth muscle layer. Expression of mRNA for each receptor was measured in mRNA isolated from each layer by RT‐PCR using specific primers. STC‐1 cells were grown in DMEM‐10; mRNA for each receptor type was measured by RT‐PCR and protein expression was measured by immunohistochemistry. Co‐localization of receptors with each other and with endocrine peptides Peptide YY (PYY) and Glucagon‐Like Peptide 1 (GLP‐1) was also examined.ResultsAll receptors were expressed in intestine and colon. The pattern of mRNA expression was similar for CaR, mGluR1, and mGluR4 such that (1) there was greater expression in colon than in intestine and (2) there was greater expression in mucosa than in muscle. In contrast, T1R1 mRNA expression was greater in intestine than colon although, similar to other L‐AA receptors, T1R1 expression in mucosa was greater than in muscle. In the EEC cell line (STC‐1), mRNA for all receptors was expressed; however, the rank order of expression was CaR>mGluR1=mGlu4>T1R1. Immunohistochemical studies showed that protein for all receptors was expressed on EEC membranes and demonstrated a high degree of co‐expression of CaR with mGluR1 and mGluR4. In contrast, T1R1 receptors were rarely co‐expressed with CaR or mGluR1 and mGluR4. STC‐1 cells expressing each receptor type also expressed GLP‐1 and PYY confirming the EEC phenotype of these STC‐1 cells.ConclusionMultiple L‐amino acid sensing receptors are expressed in murine intestine and colon, and a murine enteroendocrine cell line. CaR and mGluRs are more likely to be co‐localized in enteroendocrine cells and mediate L‐AA sensing in colon whereas T1R1 may be more important in intestine. The co‐localization of these receptors with important gut hormones in EEC suggests a role of nutrient L‐AA in regulating the release of GLP‐1 and PYY.Support or Funding InformationSupported by DK15564, DK28300, and DK34153