Amino Acid Sensing Mechanisms in Enteroendocrine STC-1 Cells

STEVEN H YOUNG; OSVALDO REY; ENRIQUE ROZENGURT

Congreso; Digestive Diseases Week; 2009

Background: Recent studies provide support for the hypothesis that the molecular pathways that mediate oral taste signaling operate in specific cells of the gastrointestinal (GI) tract. It is now established that taste-specific molecular transducers, including sweet and bitter taste receptors (referred as T1Rs and T2Rs, respectively) 〈gustducin (G〈gust), 〈transducin (G〈t- 2), PLC2 and TRPM5, are expressed by a variety of cell types of the mammalian GI tract, including endocrine cells. These transducers are also expressed by enteroendocrine cell lines that serve as model systems to elucidate signal transduction pathways that mediate chemosensing in GI cells. Amino acid sensing plays an important role in the regulation of digestive functions but its effects on enteroendocrine cells have not been explored in detail.. Results: In order to determine whether STC-1 cells sense amino acids in the medium, we challenged these cells with 20 amino acids added at 5 mM. Cell response was monitored by determining the intracellular concentration of Ca2+ ([Ca2+]i). Peak transient [Ca2+]i and plateau values are the largest for L-proline, L-serine, and L-alanine, much reduced for L-leucine and L-phenylalanine, and range down to minimal response from L-glutamic acid, and L-isoluecine, and no response from L aspartic acid or L-tryptophan. The amplitude of the response is dependent on external Ca2+ concentration; as external Ca2+ concentration increases, both peak transient amplitude and plateau amplitude are increased. Sequential addition of L-proline desensitized the subsequent response to L-alanine, and reciprocally, L-alanine desensitized Ca2+ signaling in response to L-proline. The mouse amino acid taste receptor is a heterodimer of T1R1 and T1R3 that responds to a wide variety of L- amino acids with increases in [Ca2+]i . We found that siRNA targeting T1R3 caused a significant (~40%) decrease in the response to L-proline without interfering with Ca2+ signaling in response to bombesin. These results imply the operation of several mechanisms of amino acid sensing. Amino acid-induced Ca2+ signaling was completely prevented by nitrendipine, a specific blocker of L type voltage-sensitive channels. Conclusion: Our results show that STC-1 cells respond to several L amino acids with a fast, transient rise in [Ca2+]i followed by a slower decline to an extended plateau phase. The response can be explained, in part, by taste receptor broadly tuned to sense amino acids, but also suggest the operation of other sensing systems. The results show novel and unsuspected complexities in the sensing pathways initiated by amino acids in GI endocrine cells.