Amino acid taste receptor regulates insulin secretion in pancreatic β-cell line MIN6 cells

Abstract

Amino acids such as L-glutamate and L-arginine are potent stimuli for insulin secretion from pancreatic b-cells. However, the precise molecular mechanisms of amino acid-induced insulin secretion have only partly understood. In this study, we show here that mouse pancreatic b-cell line MIN6 cells expressed amino acid taste receptor (heterodimer of type 1 taste G protein-coupled receptor member 1 and member 3; Tas1R1 and Tas1R3, respectively). We found that administration of L-glutamate or L-arginine to MIN6 cells caused the increase in free intracellular concentrations of both inositol-1,4,5-triphosphate (IP 3 ) and Ca 2+ , and umami substance inocinate enhanced the effects of L-glutamate. Effects of amino acids on intracellular IP 3 and Ca 2+ concentration were diminished by application of lactisole, a Tas1R3 receptor antagonist. Furthermore, we investigated the effect of amino acids on the insulin release from MIN6 cells by both ELISA and total internal reflection fluorescence microscopy. Application of L-glutamate or L-arginine significantly increased the release of insulin, whereas inhibited by the application of lactisole. Based on these findings, we propose that heterodimer of Tas1R1 and Tas1R3 is the fundamental receptor for the sensing amino acids and regulates the amino acid-induced insulin secretion in pancreatic b-cells.

The G Protein-coupled Receptor Family C Group 6 Subtype A (GPRC6A) Receptor Is Involved in Amino Acid-induced Glucagon-like Peptide-1 Secretion from GLUTag Cells

“…Total Internal Reflection Fluorescence (TIRF) Imaging of tPA-GFP Release-To monitor the release of tPA-GFP at the single vesicle level, we used a TIRF microscope similar to that described previously (14,19,20). Imaging was performed in modified RB (130 mM NaCl, 3.6 mM KCl, 0.5 mM NaH 2 PO 4 , 0.5 mM MgSO 4 , 1.5 mM CaCl 2 , 10 mM HEPES, 2 mM NaHCO 3 , 5 mM glucose, pH 7.4).…”

“…To analyze the TIRF imaging data, single exocytotic events were selected manually, and the average fluorescence intensity of an individual vesicle in a 0.7 m ϫ 0.7 m square placed over the vesicle center was calculated (14,19,20). To distinguish between fusion events and retreated vesicles, we focused on fluorescence changes just before the disappearance of fluorescent signals.…”

“…We therefore examined the effects of PLC and IP 3 ] i changes (Fig. 2, C and D (14,19,20,30). Preferentially, the visualization of GLP-1-containing secretory vesicles with this technique would employ fluorescent protein-tagged GLP-1.…”

Lactisole inhibits the glucose-sensing receptor T1R3 expressed in mouse pancreatic β-cells

“…These reports negate the function of lactisole as an inhibitor of the glucose-sensing receptor in rodents. In this regard, Oya et al (2011) reported recently that lactisole inhibits the umami receptor function in MIN6 cells. Their results suggest that mouse T1R3 (mT1R3) is not totally insensitive to lactisole.…”

Low glucose-induced ghrelin secretion is mediated by an ATP-sensitive potassium channel

“…To observe the release of BDNF-Venus at the single-vesicle level, we used a total internal reflection fluorescence (TIRF) microscope similar to those described previously (Mori et al 2011, Oya et al 2011, 2013a,b, Sato et al 2012, Kitaguchi et al 2013. Briefly, a high numerical aperture objective lens (CFI Apochromat TIRF, 100!, NAZ1.49, Nikon, Tokyo, Japan) was mounted on an inverted microscope (Ti-E-Perfect focus system, Nikon), and incident light for total internal reflection illumination was introduced from the high numerical aperture objective lens through a single-mode optical fiber and two illumination lenses (TI-TIRF, Nikon).…”