salicylates and allyl-isothiocyanate

TRPA1 is a nonselective cation channel belonging to the transient receptor potential (TRP) family that is expressed in peripheral sensory neurons and may play important roles in pain perception and inflammation. We found that agonist stimulation of TRPA1, along with other members of the TRP family (TRPV1-4 and TRPM8), can induce the appearance of a large pore permeable to large organic cations such as Yo-Pro (YP) and N-methyl-d-glucamine, in an agonist and divalent cation-dependent manner. YP uptake was not inhibited by a panel of putative gap junction/pannexin blockers, suggesting that gap junction proteins are not required in this process. Our data suggest that changes in the TRP channel selectivity filter itself result in a progressive but reversible pore dilation process, a process that is under strong regulation by external calcium ions. Our data suggest that calcium plays a novel role in setting the amount of time TRPA1 channels spend in a dilated state providing a mechanism that may limit sensory neuron activation by painful or irritating substances.

Topics: Animals; Benzamides; Benzoxazoles; Calcium; Calcium Channels; Carbamates; Cell Membrane; Cell Membrane Permeability; CHO Cells; Cricetinae; Cricetulus; Dogs; Dose-Response Relationship, Drug; Farnesol; Humans; Ion Channel Gating; Isothiocyanates; Kinetics; Meglumine; Membrane Transport Modulators; Nerve Tissue Proteins; Purinergic P2 Receptor Agonists; Quinolinium Compounds; Rats; Receptors, Purinergic P2; Receptors, Purinergic P2X7; Salicylates; Transfection; Transient Receptor Potential Channels; TRPA1 Cation Channel; TRPM Cation Channels; TRPV Cation Channels

The nonselective cation channel TRPA1 (ANKTM1, p120) is a potential mediator of pain, and selective pharmacological modulation of this channel may be analgesic. Although several TRPA1 activators exist, these tend to be either reactive or of low potency and/or selectivity. The aim of the present study, therefore, was to identify novel TRPA1 agonists. Using a combination of calcium fluorescent assays and whole-cell electrophysiology, we discovered several compounds that possess potent, selective TRPA1-activating activity, including several lipid compounds (farnesyl thiosalicylic acid, farnesyl thioacetic acid, 15-deoxy-Delta(12,14)-prostaglandin J(2), and 5,8,11,14-eicosatetraynoic acid), and two marketed drugs: disulfiram (Antabuse; a compound used in the treatment of alcohol abuse) and the antifungal agent chlordantoin. Farnesyl thiosalicylic acid activates the channel in excised patches and in the absence of calcium. Furthermore, using a quadruple TRPA1 mutant, we show that the mechanism of action of farnesyl thiosalicylic acid differs from that of the reactive electrophilic reagent allylisothiocyanate. As a TRPA1 agonist with a potentially novel mechanism of action, farnesyl thiosalicylic acid may be useful in the study of TRPA1 channels.

Topics: Animals; Calcium; Calcium Channels; Cells, Cultured; CHO Cells; Cricetinae; Cricetulus; Dogs; Electrophysiology; Farnesol; Fluorescence; Ganglia, Spinal; Humans; Ion Channel Gating; Isothiocyanates; Male; Nerve Tissue Proteins; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Ruthenium Red; Salicylates; Transfection; Transient Receptor Potential Channels; TRPA1 Cation Channel