ARTICLESAnandamide, the naturally occurring amide of arachidonic acid with ethanolamine, meets all key criteria of an endogenous cannabinoid substance 1 : it is released on demand by stimulated neurons 2,3 ; it activates cannabinoid receptors with high affinity 1 ; and it is rapidly eliminated through a two-step process consisting of carrier-mediated transport followed by intracellular hydrolysis 2,4 . Anandamide hydrolysis is catalyzed by the enzyme fatty acid amide hydrolase (FAAH), a membrane-bound serine hydrolase 5,6 that also cleaves other bioactive fatty acid ethanolamides such as oleoylethanolamide 7 and palmitoylethanolamide 8 . Mutant mice lacking the gene encoding FAAH (Faah) cannot metabolize anandamide 9 and, although fertile and generally normal, show signs of enhanced anandamide activity at cannabinoid receptors such as reduced pain sensation 9 . This is suggestive that drugs targeting FAAH may heighten the tonic actions of anandamide, while possibly avoiding the multiple and often unwanted effects produced by ∆ 9 -tetrahydrocannabinol (∆ 9 -THC) and other direct-acting cannabinoid agonists 10,11 . To test this hypothesis, potent, selective and systemically active inhibitors of intracellular FAAH activity are needed. However, most current inhibitors of this enzyme lack the target selectivity and biological availability required for in vivo studies [12][13][14] , whereas newer compounds, though promising, have not yet been characterized 15,16 . Thus, the therapeutic potential of FAAH inhibition remains essentially unexplored. Lead identification and optimizationDespite its unusual catalytic mechanism 6 , FAAH is blocked by a variety of serine hydrolase inhibitors, including compounds with activated carbonyls 16 . Therefore we examined whether esters of carbamic acid such as the anti-cholinesterase agent carbaryl (compound 1; Table 1) might inhibit FAAH activity in rat brain membranes. Although compound 1 was ineffective, its positional isomer 2 produced a weak inhibition of FAAH (half-maximal inhibitory concentration (IC 50 ) = 18.6 ± 0.7 µM; mean ± s.e.m., n = 3), which was enhanced by replacing the N-methyl substituent with a cyclohexyl group (compound 3; IC 50 = 324 ± 31 nM). The aryl ester 4, the benzyloxyphenyl group of which can be regarded as an elongated bioisosteric variant of the naphthyl moiety of compound 2, inhibited the activity of FAAH with a potency (IC 50 = 396 ± 63 nM) equivalent to that of compound 3. A conformational analysis of compound 4 revealed families of accessible conformers differing mainly in the torsion angle around the O-CH 2 bond, with substituents in anti or gauche conformations (data not shown). As the latter conformations more closely resembled the shape of the naphthyl derivative 3, we hypothesized that they might be responsible for the interac-

The potent analgesic effects of cannabis-like drugs and the presence of CB1-type cannabinoid receptors in pain-processing areas of the brain and spinal cord indicate that endogenous cannabinoids such as anandamide may contribute to the control of pain transmission within the central nervous system (CNS). Here we show that anandamide attenuates the pain behaviour produced by chemical damage to cutaneous tissue by interacting with CB1-like cannabinoid receptors located outside the CNS. Palmitylethanolamide (PEA), which is released together with anandamide from a common phospholipid precursor, exerts a similar effect by activating peripheral CB2-like receptors. When administered together, the two compounds act synergistically, reducing pain responses 100-fold more potently than does each compound alone. Gas-chromatography/mass-spectrometry measurements indicate that the levels of anandamide and PEA in the skin are enough to cause a tonic activation of local cannabinoid receptors. In agreement with this possibility, the CB1 antagonist SR141716A and the CB2 antagonist SR144528 prolong and enhance the pain behaviour produced by tissue damage. These results indicate that peripheral CB1-like and CB2-like receptors participate in the intrinsic control of pain initiation and that locally generated anandamide and PEA may mediate this effect.

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Severe pain remains a major area of unmet medical need. Here we report that agonists of the nuclear receptor PPAR-␣ (peroxisome proliferator-activated receptor-␣) suppress pain behaviors induced in mice by chemical tissue injury, nerve damage, or inflammation. The PPAR-␣ agonists GW7647 [2-(4-(2-(1-cyclohexanebutyl)-3-cyclohexylureido)ethyl)phenylthio)-2-methylpropionic acid], Wy-14643 [4-chloro-6-(2,3-xylidino)-2-pyrimidinylthioacetic acid], and palmitoylethanolamide (PEA) reduced nocifensive behaviors elicited in mice by intraplantar (i.pl.) injection of formalin or i.p. injection of magnesium sulfate. These effects were absent in PPAR-␣-null mice yet occurred within minutes of agonist administration in wild-type mice, suggesting that they were mediated through a transcriptionindependent mechanism. Consistent with this hypothesis, blockade of calcium-operated IK ca (K Ca 3.1) and BK ca (K Ca 1.1) potassium channels prevented the effects of GW7647 and PEA in the formalin test. Three observations suggest that PPAR-␣ agonists may inhibit nocifensive responses by acting on peripheral PPAR-␣. (i) PEA reduced formalin-induced pain at i.pl. doses that produced no increase in systemic PEA levels; (ii) PPAR-␣ was expressed in dorsal root ganglia neurons of wildtype but not PPAR-␣-null mice; and (ii) GW7647 and PEA prevented formalin-induced firing of spinal cord nociceptive neurons in rats. In addition to modulating nociception, GW7647 and PEA reduced hyperalgesic responses in the chronic constriction injury model of neuropathic pain; these effects were also contingent on PPAR-␣ expression and were observed following either acute or subchronic PPAR-␣ agonist administration. Finally, acute administration of GW7647 and PEA reduced hyperalgesic responses in the complete Freund's adjuvant and carrageenan models of inflammatory pain. Our results suggest that PPAR-␣ agonists may represent a novel class of analgesics.Current therapies do not control safely and effectively severe pain states-a broad spectrum of debilitating conditions that comprises acute, persistent inflammatory, and neuropathic pain. Even widely used drugs, such as opiates (e.g., morphine) or anticonvulsants (e.g., gabapentin), are only active in a fraction of the patient population and produce multiple, often serious, side effects. Thus, despite continuing progress in analgesic drug discovery, the need for therapeutic agents capable of blocking abnormal pain sensation without impairing normal abilities remains largely unmet.PPAR-␣ is a nuclear receptor that serves important functions in lipid nutrient utilization and inflammation (Taylor et al., 2002;Kostadinova et al., 2005). Like other members of the nuclear receptor superfamily, PPAR-␣ is activated

Palmitoylethanolamide (PEA), the naturally occurring amide of ethanolamine and palmitic acid, is an endogenous lipid that modulates pain and inflammation. Although the anti-inflammatory effects of PEA were first characterized nearly 50 years ago, the identity of the receptor mediating these actions has long remained elusive. We recently identified the ligand-activated transcription factor, peroxisome proliferator-activated receptor-alpha (PPAR-a), as the receptor mediating the anti-inflammatory actions of this lipid amide. Here we outline the history of PEA, starting with its initial discovery in the 1950s, and discuss the pharmacological properties of this compound, particularly in regards to its ability to activate PPAR-a. Discovery of PEAThe discovery of naturally occurring fatty acid ethanolamides (FAEs) (Fig. 1) stems from an interesting clinical finding in the early 1940s, when investigators noted that supplementing the diets of 0024-3205/$ -see front matter D