Full inhibition of spinal FAAH leads to TRPV1-mediated analgesic effects in neuropathic rats and possible lipoxygenase-mediated remodeling of anandamide metabolism.

Katarzyna Starowicz,Michal Korostynski,Magdalena Zychowska,Natalia Malek,Vincenzo Di Marzo,Luigia Cristino,Wioletta Makuch,Stefania Petrosino,Barbara Przewlocka,Luciano De Petrocellis,Michal Slezak,Bernard Le Foll

doi:10.1371/journal.pone.0060040

Abstract

Neuropathic pain elevates spinal anandamide (AEA) levels in a way further increased when URB597, an inhibitor of AEA hydrolysis by fatty acid amide hydrolase (FAAH), is injected intrathecally. Spinal AEA reduces neuropathic pain by acting at both cannabinoid CB1 receptors and transient receptor potential vanilloid-1 (TRPV1) channels. Yet, intrathecal URB597 is only partially effective at counteracting neuropathic pain. We investigated the effect of high doses of intrathecal URB597 on allodynia and hyperalgesia in rats with chronic constriction injury (CCI) of the sciatic nerve. Among those tested, the 200 µg/rat dose of URB597 was the only one that elevated the levels of the FAAH non-endocannabinoid and anti-inflammatory substrates, oleoylethanolamide (OEA) and palmitoylethanolamide (PEA), and of the endocannabinoid FAAH substrate, 2-arachidonoylglycerol, and fully inhibited thermal and tactile nociception, although in a manner blocked almost uniquely by TRPV1 antagonism. Surprisingly, this dose of URB597 decreased spinal AEA levels. RT-qPCR and western blot analyses demonstrated altered spinal expression of lipoxygenases (LOX), and baicalein, an inhibitor of 12/15-LOX, significantly reduced URB597 analgesic effects, suggesting the occurrence of alternative pathways of AEA metabolism. Using immunofluorescence techniques, FAAH, 15-LOX and TRPV1 were found to co-localize in dorsal spinal horn neurons of CCI rats. Finally, 15-hydroxy-AEA, a 15-LOX derivative of AEA, potently and efficaciously activated the rat recombinant TRPV1 channel. We suggest that intrathecally injected URB597 at full analgesic efficacy unmasks a secondary route of AEA metabolism via 15-LOX with possible formation of 15-hydroxy-AEA, which, together with OEA and PEA, may contribute at producing TRPV1-mediated analgesia in CCI rats.

Highlights

The endocannabinoid system includes the cannabinoid CB1 and CB2 receptors, and the endogenous agonists at these receptors, i.e. the endocannabinoids anandamide (AEA) and 2arachidonoyl glycerol (2-AG)]
Fatty acid amide hydrolase (FAAH) catalyzes the hydrolysis of AEA and its two congeners, the acylethanolamides oleoylethanolamide (OEA) and palmitoylethanolamide (PEA), which occur in higher tissue concentrations than AEA and produce their effects in a mostly cannabinoid receptor-independent manner
We assessed the selective FAAH inhibitor, URB597, on signs of neuropathic pain in rats subjected to constriction injury (CCI)

Summary

Introduction

The endocannabinoid system includes the cannabinoid CB1 and CB2 receptors (two G-protein-coupled receptors activated by the main psychotropic component of Cannabis sativa, D9tetrahydrocannabinol [1]), and the endogenous agonists at these receptors, i.e. the endocannabinoids anandamide (AEA) and 2arachidonoyl glycerol (2-AG)]. AEA activates the transient receptor potential vanilloid-1 (TRPV1) channel [2], which transduces the pronociceptive and heat-like effects of another plant natural product, the pungent component of hot peppers, capsaicin [3,4,5,6]. TRPV1 usually, but not always, plays in pain transmission a role opposite to cannabinoid receptors [7,8] and, the possible effects on pain of the activation of this channel by AEA have been thoroughly studied [9,10,11]. Lipoxygenase (LOX)- and cyclooxygenase (COX)-2- catalyzed oxygenation of endocannabinoids generate several bioactive compounds [15,16]. Whilst COX-2 derivatives are inactive at cannabinoid receptors and TRPV1 [16], LOX products of AEA are still able to activate these targets [17,18]

Methods

Results

Conclusion