Serotonin (5-hydroxytryptamine; 5-HT) is abundantly present throughout the gastrointestinal tract and stored mostly in enterochromaffin (EC) cells, which are located on the mucosal surface. 5-HT released from EC cells stimulate both intrinsic and extrinsic nerves, which results in various physiological and pathophysiological responses, such as gastrointestinal contractions. EC cells are believed to have the ability to respond to the chemical composition of the luminal contents of the gut; however, the underlying molecular and cellular mechanisms have not been identified. Here, we demonstrate that the transient receptor potential (TRP) cation channel TRPA1, which is activated by pungent compounds or cold temperature, is highly expressed in EC cells. We also found that TRPA1 agonists, including allyl isothiocyanate and cinnamaldehyde, stimulate EC cell functions, such as increasing intracellular Ca 2؉ levels and 5-HT release, by using highly concentrated EC cell fractions and a model of EC cell function, the RIN14B cell line. Furthermore, we showed that allyl isothiocyanate promotes the contraction of isolated guinea pig ileum via the 5-HT 3 receptor. Taken together, our results indicate that TRPA1 acts as a sensor molecule for EC cells and may regulate gastrointestinal function.gastrointestinal tract ͉ RIN14B T he gastrointestinal tract has many functions, such as secretion, motility, and absorption. These functions are affected by various signals from the luminal contents, including nutrient and non-nutrient chemicals, mechanical factors, and microorganisms (1). The endocrine cells of the gut (hereafter enteroendocrine cells) are thought to be highly specialized mucosal cell subpopulations that receive luminal signals. There are more than 10 different types of enteroendocrine cells, and each type produces distinct transmitters/hormones (2). Serotonin (5-HT)-containing enterochromaffin (EC) cells, which are located throughout the gut, are considered to be the most prevalent enteroendocrine cells (3, 4). The 5-HT released from EC cells activate the submucosal sensory branch of the enteric nervous system and also control gastrointestinal motility and chloride secretion via interneurons and motor neurons (5, 6). Hence, EC cells are considered to be a major component of both the physiology and pathophysiology of gastrointestinal function (7,8). It has been suggested that EC cells respond to the contents of the lumen through the activation of receptor-operated or voltage-dependent Ca 2ϩ channels (9), however, the details of the cellular and molecular mechanisms have not yet been clarified.Many ion channels, like the transient receptor potential (TRP) channels expressed in sensory neurons, respond to natural compounds, especially spices and herbal medicines. For example, the vanilloid receptor (TRPV1) responds to the plant component capsaicin (the pungent ingredient in chili peppers), which produces the psychophysical sensation of heat or burning, whereas TRPM8 responds to menthol (found in peppermint), which produces ...

Recently, we discovered that transient receptor potential ankyrin1 channel (TRPA1) is highly expressed in human and rat enterochromaffin (EC) cells, and those TRPA1 agonists such as allyl isothiocyanates (AITC) and cinnamaldehyde (CA) enhance the release of serotonin (5-hydroxytryptamine; 5-HT) from EC cells in vitro. In this study, QGP-1 cells, a human pancreatic endocrine cell line, were found to highly express TRPA1 and EC cell marker genes, such as tryptophan hydroxylase 1 (TPH1), chromogranin A (CgA), synaptophysin, ATP-dependent vesicular monoamine transporter 1 (VMAT1), metabotropic glutamate receptor 4 (mGluR4), beta1-adrenergic receptor (ADB1), muscarinic 4 acetylcholine receptor (ACM4), substance P, serotonin transporter (SERT), and guanylin. Furthermore, the TRPA1 agonists AITC, CA, and acrolein concentration dependently evoked an increase in intracellular Ca(2+) influx and the release of 5-HT in QGP-1 cells. The effects of these TRPA1 agonists were inhibited by ruthenium red, a TRPA1 antagonist, and TRPA1-specific siRNA. These results indicate that the Ca(2+) influx increase and 5-HT release induced by AITC, CA and acrolein in QGP-1 cells were mediated by TRPA1, and that the QGP-1 cell line could be a new model for the investigation of TRPA1 function in the human EC cell.

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Our recent study found that TRPA1 is highly expressed in enterochromaffin cells and that stimulation of these cells with TRPA1 agonists enhances 5-hydroxytryptamine (5-HT) secretion in vitro. Here, to demonstrate the 5-HT-releasing effect of TRPA1 agonists in vivo, we examined the effect of TRPA1 agonists on gastric emptying in rats. The results showed that TRPA1 agonists dose-dependently delayed gastric emptying. Further, the effects of TRPA1 agonists on this delay were abolished in rats treated with a TRPA1 antagonist, an inhibitor of tryptophan hydroxylase, or a 5-HT(3) receptor antagonist. Taken together, these results indicate that TRPA1 agonists delay in vivo gastric emptying through serotonergic pathways.

Although it is known that both clonidine and loperamide cause delayed colonic transit in mice, these models of drug-induced experimental constipation have not yet been fully characterized. Therefore, the aims of this study were to validate the clonidine- and loperamide-induced delays of colonic transit in mice as models of atonic and spastic constipation, respectively, and to evaluate the effect of mustard oil, a TRPA1 agonist, in both models. Colonic transit was evaluated in mice by determining the time needed to evacuate a bead inserted into the distal colon. Both loperamide and clonidine dose-dependently prolonged the evacuation time. Clonidine (10 μg/kg) and loperamide (0.3 mg/kg) tripled the evacuation time compared to controls. These delays were antagonized by the administration of yohimbine and naloxone, respectively. Tegaserod, a gastrointestinal motor-stimulating drug, reversed the delay in both models, but the effects were diminished at high doses. Atropine, an antispastic drug, improved the loperamide-induced delay, but did not affect the clonidine-induced delay. Mustard oil accelerated the colonic transit dose-dependently in both models of drug-induced constipations. These results indicate that clonidine- and loperamide-induced delays in colonic transit are models of atonic and spastic constipation, respectively, and that mustard oil may be effective on both types of constipation.

The emergence of new molecularly targeted agents has contributed to reducing the approval lag, most likely due to improvements in R&D strategy.

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