Received for publication December 20, 2006.
Revised February 15, 2007.
Accepted for publication February 15, 2007.

Activation of TRPA1 Channels by the Fatty Acid Amide Hydrolase Inhibitor URB597

Abstract

As a member of the transient receptor potential (TRP) ion channel superfamily, the ligand-gated ion channel TRPA1 has been implicated in nociceptive function and pain states. The endogenous ligands that activate TRPA1 remain unknown. However various agonists have been identified, including environmental irritants (e.g., acrolein) and ingredients of pungent natural products (e.g., allyl isothiocyanate or ITC, cinnamaldehyde, allicin and gingerol). In general, these agents are either highly reactive, nonselective, or not potent or efficacious, significantly limiting their utilities in the study of TRPA1 channel properties and biological functions. In a search for novel TRPA1 agonists, we identified 3'-carbamoylbiphenyl-3-yl cyclohexylcarbamate (URB597), a potent and systemically active inhibitor of fatty acid amide hydrolase (FAAH). This enzyme is responsible for anandamide degradation therefore has been pursued as an anti-nociceptive and anti-epleptic drug target. Using Ca²⁺ influx assays and patch-clamp techniques, we demonstrated that URB597 could activate heterologously expressed human and rat TRPA1 channels, while two other FAAH inhibitors (i.e., URB532 and Compound 7) had no effect. When applied to inside-out membrane patches expressing rat TRPA1, URB597 elicited single-channel activities with a unitary conductance of 40 pS. Furthermore, URB597 activated TRPA1 channels endogenously expressed in a population of rat dorsal root ganglion neurons that also responded to ITC. In contrast to its effect on TRPA1, URB597 inhibited TRPM8 and had no effects on TRPV1 or TRPV4. Thus we conclude that URB597 is a novel agonist of TRPA1 and likely activates the channel through a direct gating mechanism.

Key words: Ion channel regulation, Func. analysis receptor/ion channel mutants, Single channel kinetics

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