NON-SELECTIVE EFFECTS OF THE MONOACYLGLYCEROL LIPASE INHIBITOR URB602 ON ENDOCANNABINOID LEVELS IN AN INFLAMMATORY PAIN MODEL

Ian Robinson1, David A Barrett2, David A Kendall1, Victoria Chapman1
1University of Nottingham, School of Biomedical Sciences, Nottingham, United Kingdom, 2University of Nottingham, School of Pharmacy, Nottingham, United Kingdom

Cannabinoids and endocannabinoids (ECs) produce well described anti-nociceptive effects via the activation of cannabinoid (CB) receptors, which are present on primary afferent sensory nerve fibres as well as spinal and supraspinal neurones. ECs are metabolised by a number of enzymes including fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). 2-Arachidonoylglycerol (2AG) is primarily metabolised by MAGL. Systemic administration of the MAG lipase inhibitor (URB602) attenuated hyperalgesia in animal models of inflammatory pain (Hohmann et al. 2005; Guindon et al 2006). The aim of the present study was to investigate the local effects of the MAGL inhibitor URB602 on hyperalgesia and levels of ECs in the carrageenan model of inflammatory pain.

Male Sprague-Dawley rats (240-260g) received an intraplantar injection of URB602 (70μg in 50μl), or vehicle (50μl 3% Tween 80 in saline), 30 minutes prior to intraplantar injection of 2% carrageenan or saline (100μl). Weight bearing, measured with an Incapacitance Tester (Linton Instrumentation, U.K.), on the left (injected) and right hindpaw was assessed 10 minutes prior to injection of carrageenan / saline and at 90, 150 and 210 minutes post-injection. 3 hours after carrageenan injection, rats were killed and ECs extracted and purified from hindpaw tissue using an ethyl acetate/hexane solvent extraction, followed by solid phase extraction (SPE). Extracts were then analysed by liquid chromatography-coupled tandem mass spectrometry (Richardson et.al., 2007). Weight bearing was calculated by subtracting the weight on the left (ipsilateral) hindpaw from that on the right (contralateral) hindpaw. Data were then analysed using a two-way ANOVA with Newman Keul’s post-hoc test and are expressed as mean ± SEM. EC data were analysed using Mann-Whitney t-tests and are expressed as mean ± SEM.

Intraplantar injection of carrageenan in vehicle pre-treated rats produced a significant change in weight bearing (45.25 ± 3.23g) compared to vehicle-saline controls (-13.67 ± 4.95g). Intraplantar injection of URB602 in carrageenan-treated rats significantly (p<0.05) reduced changes in weight bearing (6.31 ± 4.9g) compared to vehicle-carrageenan treated rats (45.25 ± 3.23g). Thus, URB602 was anti-hyperalgesic in the carrageenan model of inflammatory pain. As previously reported, levels of anandamide (AEA) and palmitoylethanolamide (PEA) in the ipsilateral hindpaw of carrageenan treated rats were significantly lower (P < 0.01) than levels in the ipsilateral hindpaw of saline treated rats (data not shown). In carrageenan-treated rats, intraplantar injection of URB602 (70μg in 50 μl) produced a significant (P < 0.001) increase in levels of AEA (108 ± 33.2 nmol/g), 2-AG (15.73 ± 3.2 pmol/g) OEA (15.62 ± 3.25 pmol/g) and PEA (7.39 ± 1.55pmol/g) in the ipsilateral hindpaw, compared to vehicle–treated carrageenan rats (12.8 ± 2.85 nmol/g and, 1.87 ± 0.35, 3.33 ± 0.41, and 2.09 ± 0.27 pmol/g respectively). These data indicate that, although URB602 attenuates inflammatory hyperalgesia, these effects are not due to a specific increase in levels of 2-AG levels in hindpaw tissue, but might also be mediated by increases in AEA and PEA.

Guindon et al. (2006). Pain 121(1-2): 85-93.
Hohmann, et al. (2005) Nature 435(7045): 1108-12.
Richardson et al. (2007) Anal Biochem, 360, 216-26.