Niforatos Wende, Zhang Xu-Feng, Lake Marc R, Walter Karl A, Neelands Torben, Holzman Thomas F, Scott Victoria E, Faltynek Connie R, Moreland Robert B, Chen Jun
Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064-6125, USA.
Mol Pharmacol. 2007 May;71(5):1209-16. doi: 10.1124/mol.106.033621. Epub 2007 Feb 21.
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 (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 and therefore has been pursued as an antinociceptive and antiepileptic drug target. Using Ca(2+) influx assays and patch-clamp techniques, we demonstrated that URB597 could activate heterologously expressed human and rat TRPA1 channels, whereas 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 probably activates the channel through a direct gating mechanism.
作为瞬时受体电位（TRP）离子通道超家族的一员，配体门控离子通道TRPA1与伤害感受功能和疼痛状态有关。激活TRPA1的内源性配体尚不清楚。然而，已鉴定出多种激动剂，包括环境刺激物（如丙烯醛）和辛辣天然产物的成分[如异硫氰酸烯丙酯（ITC）、肉桂醛、大蒜素和姜辣素]。一般来说，这些试剂要么具有高反应性、非选择性，要么效力或效果不佳，这严重限制了它们在TRPA1通道特性和生物学功能研究中的应用。在寻找新型TRPA1激动剂的过程中，我们鉴定出了3'-氨基甲酰基联苯-3-基环己基氨基甲酸酯（URB597），一种强效且具有全身活性的脂肪酸酰胺水解酶（FAAH）抑制剂。这种酶负责花生四烯乙醇胺的降解，因此一直被作为抗伤害感受和抗癫痫药物靶点进行研究。使用钙离子内流测定法和膜片钳技术，我们证明URB597可以激活异源表达的人类和大鼠TRPA1通道，而另外两种FAAH抑制剂（即URB532和化合物7）则没有效果。当应用于表达大鼠TRPA1的内向外膜片时，URB597引发了单位电导为40 pS的单通道活动。此外，URB597激活了在一群大鼠背根神经节神经元中内源性表达的TRPA1通道，这些神经元也对ITC有反应。与它对TRPA1的作用相反，URB597抑制TRPM8，对TRPV1或TRPV4没有影响。因此，我们得出结论，URB597是一种新型的TRPA1激动剂，可能通过直接门控机制激活该通道。
