Abstract
BackgroundCytochrome-P450 (CYP450) epoxygenases metabolise arachidonic acid (AA) into four different biologically active epoxyeicosatrienoic acid (EET) regioisomers. Three of the EETs (i.e., 8,9-, 11,12- and 14,15-EET) are rapidly hydrolysed by the enzyme soluble epoxide hydrolase (sEH). Here, we investigated the role of sEH in nociceptive processing during peripheral inflammation.ResultsIn dorsal root ganglia (DRG), we found that sEH is expressed in medium and large diameter neurofilament 200-positive neurons. Isolated DRG-neurons from sEH-/- mice showed higher EET and lower DHET levels. Upon AA stimulation, the largest changes in EET levels occurred in culture media, indicating both that cell associated EET concentrations quickly reach saturation and EET-hydrolyzing activity mostly effects extracellular EET signaling. In vivo, DRGs from sEH-deficient mice exhibited elevated 8,9-, 11,12- and 14,15-EET-levels. Interestingly, EET levels did not increase at the site of zymosan-induced inflammation. Cellular imaging experiments revealed direct calcium flux responses to 8,9-EET in a subpopulation of nociceptors. In addition, 8,9-EET sensitized AITC-induced calcium increases in DRG neurons and AITC-induced calcitonin gene related peptide (CGRP) release from sciatic nerve axons, indicating that 8,9-EET sensitizes TRPA1-expressing neurons, which are known to contribute to mechanical hyperalgesia. Supporting this, sEH-/- mice showed increased nociceptive responses to mechanical stimulation during zymosan-induced inflammation and 8,9-EET injection reduced mechanical thresholds in naive mice.ConclusionOur results show that the sEH can regulate mechanical hyperalgesia during inflammation by inactivating 8,9-EET, which sensitizes TRPA1-expressing nociceptors. Therefore we suggest that influencing the CYP450 pathway, which is actually highly considered to treat cardiovascular diseases, may cause pain side effects.
Highlights
Cytochrome-P450 (CYP450) epoxygenases metabolise arachidonic acid (AA) into four different biologically active epoxyeicosatrienoic acid (EET) regioisomers
Free AA is a major substrate for cyclooxygenases (COX), lipoxygenases (LOX) and cytochrome P450 (CYP450) epoxygenases which metabolize it to prostanoids, leukotrienes and epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetranoic acids (HETES), respectively [2,3,4]
We have investigated EETs in nociceptive processing by phenotyping mice with targeted deletion of soluble epoxide hydrolase (sEH), and find that the EETs metabolized by sEH sensitize TRPA1 expressing dorsal root ganglia (DRG) neurons and that sEH-/- mice show an increased mechanical hyperalgesia
Summary
Cytochrome-P450 (CYP450) epoxygenases metabolise arachidonic acid (AA) into four different biologically active epoxyeicosatrienoic acid (EET) regioisomers. EETs either bind intracellular targets, are released to act as auto- or paracrine mediators or are stored in cell membranes esterified to phospholipids [9]. They activate PPARg or the cAMP/PKA pathway and modulate and/or activate a variety of channels including several transient receptor potential (TRP) channel isoforms, large-conductance Ca2+-activated K+ channels (BK(Ca)) and L-type voltage gated calcium channels (Ca(v)) [9]. Three EETs (8,9-, 11,12- and 14,15-EET) are metabolized by soluble epoxide hydrolase (sEH) to their corresponding dehydro metabolites (DHET), which are thought to be less active [10]. A conditional knockout of cytochrome P450 reductase (CRP) in CNS neurons, which blocks CYP450 activity, largely abolishes morphineinduced anti-nociception, suggesting that EETs have an anti-nociceptive activity downstream of the μ-opioid receptor [7]
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