Ligand determinants of fatty acid activation of the pronociceptive ion channel TRPA1

William John Redmond; Liuqiong Gu; Maxime Camo; Peter McIntyre; Mark Connor

doi:10.7717/peerj.248

Ligand determinants of fatty acid activation of the pronociceptive ion channel TRPA1

PeerJ. 2014 Jan 28:2:e248. doi: 10.7717/peerj.248. eCollection 2014.

Authors

William John Redmond¹, Liuqiong Gu², Maxime Camo¹, Peter McIntyre³, Mark Connor¹

Affiliations

¹ Australian School of Advanced Medicine, Macquarie University , NSW , Australia.
² Department of Pharmacology, University of Melbourne , Parkville , Victoria , Australia.
³ Department of Pharmacology, University of Melbourne , Parkville , Victoria , Australia ; Health Innovations Research Institute and School of Medical Sciences, RMIT University , Melbourne , Victoria , Australia.

Abstract

Background and purpose. Arachidonic acid (AA) and its derivatives are important modulators of cellular signalling. The transient receptor potential cation channel subfamily A, member 1 (TRPA1) is a cation channel with important functions in mediating cellular responses to noxious stimuli and inflammation. There is limited information about the interactions between AA itself and TRPA1, so we investigated the effects of AA and key ethanolamide and amino acid/neurotransmitter derivatives of AA on hTRPA1. Experimental approach. HEK 293 cells expressing hTRPA1 were studied by measuring changes in intracellular calcium ([Ca] i ) with a fluorescent dye and by standard whole cell patch clamp recordings. Key results. AA (30 μM) increased fluorescence in hTRPA1 expressing cells by 370% (notional EC 50 13 μM). The covalent TRPA1 agonist cinnamaldehyde (300 μM) increased fluorescence by 430% (EC 50, 11 μM). Anandamide (230%) and N-arachidonoyl tyrosine (170%) substantially activated hTRPA1 at 30 μM, however, N-arachidonoyl conjugates of glycine and taurine were less effective while N-acyl conjugates of 5-HT did not affect hTRPA1. Changing the acyl chain length or the number and position of double bonds reduced fatty acid efficacy at hTRPA1. Mutant hTRPA1 (Cys621, Cys641 and Cys665 changed to Ser) could be activated by AA (100 μM, 40% of wild type) but not by cinnamaldehyde (300 μM). Conclusions and implications. AA is a more potent activator of TRPA1 than its ethanolamide or amino acid/neurotransmitter derivatives and acts via a mechanism distinct from that of cinnamaldehyde, further underscoring the likelyhood of multiple pharmacologically exploitable sites on hTRPA1.

Keywords: Arachidonic acid; Arachidonoyl amino acid/neurotransmitter conjugate; Inflammation; Pain; TRP channel.

Grants and funding

This work was supported by NHMRC Project Grant 1002680 to MC Sr and PM. WR and MC Jr were supported by Macquarie University International Research Scholarships, and received top up funding from the Australian School of Advanced Medicine. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.