We characterized an activation mechanism of the human LTRPC2 protein, a member of the transient receptor potential family of ion channels, and demonstrated that LTRPC2 mediates Ca2+ influx into immunocytes. Intracellular pyrimidine nucleotides, adenosine 5'-diphosphoribose (ADPR), and nicotinamide adenine dinucleotide (NAD), directly activated LTRPC2, which functioned as a Ca2+-permeable nonselective cation channel and enabled Ca2+ influx into cells. This activation was suppressed by intracellular adenosine triphosphate. These results reveal that ADPR and NAD act as intracellular messengers and may have an important role in Ca2+ influx by activating LTRPC2 in immunocytes.

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 ...

To find a novel human ion channel gene we have executed an extensive search by using a human genome draft sequencing data base. Here we report a novel twopore domain K ؉ channel, TRESK (TWIK-related spinal cord K ؉ channel). TRESK is coded by 385 amino acids and shows low homology (19%) with previously characterized two-pore domain K ؉ channels. However, the most similar channel is TREK-2 (two-pore domain K ؉ channel), and TRESK also has two pore-forming domains and four transmembrane domains that are evolutionarily conserved in the two-pore domain K ؉ channel family. Moreover, we confirmed that TRESK is expressed in the spinal cord. Electrophysiological analysis demonstrated that TRESK induced outward rectification and functioned as a background K ؉ channel. Pharmacological analysis showed TRESK to be inhibited by previously reported K ؉ channel inhibitors Ba 2؉ , propafenone, glyburide, lidocaine, quinine, quinidine, and triethanolamine. Functional analysis demonstrated TRESK to be inhibited by unsaturated free fatty acids such as arachidonic acid and docosahexaenoic acid. TRESK is also sensitive to extreme changes in extracellular and intracellular pH. These results indicate that TRESK is a novel two-pore domain K ؉ channel that may set the resting membrane potential of cells in the spinal cord.

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The response to intracellular ADP-ribose in the rat CRI-G1 insulinoma cell line was studied using a patch-clamp method. Dialysis of ADP-ribose into cells induced a response in a dose-dependent manner. The reversal potentials in various solutions showed that the ADP-ribose-gated channel was a Ca2+-permeable nonselective cation channel. In inside-out recordings, ADP-ribose and b-NAD induced responses in the same patch. The single-channel current-voltage relationships for ADP-ribose- and b-NAD-induced responses were almost identical, indicating that ADP-ribose and b-NAD activated the same channel. The physiological properties of the ADP-ribose-gated channel are similar to those we reported previously for the cloned transient receptor potential channel TRPM2. Moreover, RT-PCR analysis showed that TRPM2 was abundantly expressed in CRI-G1 cells, suggesting that the ADP-ribose-gated channel represents the native TRPM2 channel in CRI-G1 cells. These results suggest that ADP-ribose can be an endogenous modulator of Ca2+ influx through the TRPM2 channel into CRI-G1 cells.