4-hydroxy-2-nonenal and allyl-isothiocyanate

TRPA1 (transient receptor potential ankyrin 1), the lone member of the mammalian ankyrin TRP subfamily, is a Ca

Topics: Aldehydes; Animals; Calcitonin; Calcium; Calcium Channels; Cardiovascular System; Crotalus; Endothelial Cells; Gene Expression Regulation; Humans; Hypertension; Inflammation; Isothiocyanates; Molecular Conformation; Mustard Plant; Nerve Tissue Proteins; Plant Oils; Protein Conformation; Protein Domains; Stroke; Transient Receptor Potential Channels; TRPA1 Cation Channel; Vasodilation

Several transient receptor potential (TRP) channels are expressed in pancreatic beta cells and have been proposed to be involved in insulin secretion. However, the endogenous ligands for these channels are far from clear. Here, we demonstrate the expression of the transient receptor potential ankyrin 1 (TRPA1) ion channel in the pancreatic beta cells and its role in insulin release. TRPA1 is an attractive candidate for inducing insulin release because it is calcium permeable and is activated by molecules that are produced during oxidative glycolysis.. Immunohistochemistry, RT-PCR, and Western blot techniques were used to determine the expression of TRPA1 channel. Ca²⁺ fluorescence imaging and electrophysiology (voltage- and current-clamp) techniques were used to study the channel properties. TRPA1-mediated insulin release was determined using ELISA.. TRPA1 is abundantly expressed in a rat pancreatic beta cell line and freshly isolated rat pancreatic beta cells, but not in pancreatic alpha cells. Activation of TRPA1 by allyl isothiocyanate (AITC), hydrogen peroxide (H₂O₂), 4-hydroxynonenal (4-HNE), and cyclopentenone prostaglandins (PGJ₂) and a novel agonist methylglyoxal (MG) induces membrane current, depolarization, and Ca²⁺ influx leading to generation of action potentials in a pancreatic beta cell line and primary cultured pancreatic beta cells. Activation of TRPA1 by agonists stimulates insulin release in pancreatic beta cells that can be inhibited by TRPA1 antagonists such as HC030031 or AP-18 and by RNA interference. TRPA1-mediated insulin release is also observed in conditions of voltage-gated Na⁺ and Ca²⁺ channel blockade as well as ATP sensitive potassium (K(ATP)) channel activation.. We propose that endogenous and exogenous ligands of TRPA1 cause Ca²⁺ influx and induce basal insulin release and that TRPA1-mediated depolarization acts synergistically with K(ATP) channel blockade to facilitate insulin release.

Topics: Aldehydes; Animals; Blotting, Western; Calcium; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Hydrogen Peroxide; Immunohistochemistry; Insulin; Insulin-Secreting Cells; Isothiocyanates; Prostaglandin D2; Pyruvaldehyde; Rats; Reverse Transcriptase Polymerase Chain Reaction; TRPA1 Cation Channel; TRPC Cation Channels

Transient receptor potential ankyrin (TRPA) 1, an excitatory ion channel expressed by sensory neurons, mediates somatic and visceral pain in response to direct activation or noxious mechanical stimulation. Although the intestine is routinely exposed to irritant alimentary compounds and inflammatory mediators that activate TRPA1, there is no direct evidence for functional TRPA1 receptors on enteric neurons, and the effects of TRPA1 activation on intestinal function have not been determined. We characterized expression of TRPA1 by enteric neurons and determined its involvement in the control of intestinal contractility and transit.. TRPA1 expression was characterized by reverse-transcription polymerase chain reaction and immunofluorescence analyses. TRPA1 function was examined by Ca(2+) imaging and by assays of contractile activity and transit.. We detected TRPA1 messenger RNA in the mouse intestine and TRPA1 immunoreactivity in enteric neurons. The cecum and colon had immunoreactivity for neuronal TRPA1, but the duodenum did not. TRPA1 immunoreactivity was also detected in inhibitory motoneurons and descending interneurons, cholinergic neurons, and intrinsic primary afferent neurons. TRPA1 activators, including cinnamaldehyde, allyl isothiocyanate (AITC), and 4-hydroxynonenal, increased [Ca(2+)](i) in myenteric neurons. These were reduced by a TRPA1 antagonist (HC-030031) or deletion of Trpa1. TRPA1 activation inhibited contractility of the segments of colon but not stomach or small intestine of Trpa1(+/+) but not Trpa1(-/-) mice; this effect was reduced by tetrodotoxin or N(G)-nitro-l-arginine methyl ester. Administration of AITC by gavage did not alter gastric emptying or small intestinal transit, but luminal AITC inhibited colonic transit via TRPA1.. Functional TRPA1 is expressed by enteric neurons, and activation of neuronal TRPA1 inhibits spontaneous neurogenic contractions and transit of the colon.

Topics: Acrolein; Aldehydes; Animals; Carbachol; Cecum; Colon; Duodenum; Epithelial Cells; Female; Fluorescent Antibody Technique; Ganglia; Gastric Emptying; Gastric Mucosa; Gastrointestinal Motility; Ileum; Interneurons; Intestinal Mucosa; Isothiocyanates; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Neurons; Muscle Contraction; Muscle, Smooth; Neurons, Afferent; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stomach; Substance P; Transient Receptor Potential Channels; TRPA1 Cation Channel

The transient receptor potential subfamily A member 1 (TRPA1) is a non-selective cation channel implicated in the pathogenesis of several airway diseases like asthma and chronic obstructive pulmonary disease (COPD). Most of the research on TRPA1 focuses on its expression and function in neuronal context; studies investigating non-neuronal expression of TRPA1 are lacking. In the present study, we show functional expression of TRPA1 in human lung fibroblast cells (CCD19-Lu) and human pulmonary alveolar epithelial cell line (A549). We demonstrate TRPA1 expression at both mRNA and protein levels in these cell types. TRPA1 selective agonists like allyl isothiocyanate (AITC), 4-hydroxynonenal (4-HNE), crotonaldehyde and zinc, induced a concentration-dependent increase in Ca+2 influx in CCD19-Lu and A549 cells. AITC-induced Ca+2 influx was inhibited by Ruthenium red (RR), a TRP channel pore blocker, and by GRC 17536, a TRPA1 specific antagonist. Furthermore, we also provide evidence that activation of the TRPA1 receptor by TRPA1 selective agonists promotes release of the chemokine IL-8 in CCD19-Lu and A549 cells. The IL-8 release in response to TRPA1 agonists was attenuated by TRPA1 selective antagonists. In conclusion, we demonstrate here for the first time that TRPA1 is functionally expressed in cultured human lung fibroblast cells (CCD19-Lu) and human alveolar epithelial cell line (A549) and may have a potential role in modulating release of this important chemokine in inflamed airways.

Topics: Aldehydes; Calcium; Calcium Channels; Cations, Divalent; Cells, Cultured; Chlorides; Dose-Response Relationship, Drug; Epithelial Cells; Fibroblasts; Humans; Interleukin-8; Isothiocyanates; Nerve Tissue Proteins; Ruthenium Red; Transient Receptor Potential Channels; TRPA1 Cation Channel; Zinc Compounds