In the early nineties cannabinoid receptors, the main target for Δ9-tetrahydrocannabinol (THC), the psychoactive component of marijuana were identified. Shortly after their endogenous ligands, N-arachidonoylethanolamine (anandamide, AEA) and 2-diacylglycerol (2-AG) were characterized. The enzymes primarily responsible for catalysing the degradation of AEA and 2-AG are fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MGL) respectively. AEA is a member of the N-acylethanolamine (NAE) class of lipids, which depending on the acyl chain length and number of double bonds can act as ligands for a variety of biological targets. Exogenous cannabinoids have long been reported to have analgesic effects, however the clinical usefulness of such substances is limited by their psychoactive effects. Inhibition of endocannabinoid degradation would mean enhancing the therapeutic effects without producing these unwanted side effects. In order to succeed in developing such compounds the pharmacology of the enzymes responsible for the degradation of endocannabinoids has to be thoroughly understood. When the preclinical part of this thesis was planned, FAAH had been well characterized whereas little was known as to the pharmacology of MGL. A series of compounds were tested in this first study aiming to find MGL-selective compounds. Although no compounds showed selectivity for MGL over FAAH, several interesting agents affecting both enzymes were identified. In order to increase the knowledge concerning which patient group would benefit from such treatment strategies it is important to investigate in which pain states the endocannabinoids/NAEs are altered. Thus the general aim of the clinical part of this thesis was to investigate the levels of endocannabinoids/NAEs in the interstitium of the trapezius muscle in women suffering from chronic neck/shoulder pain (CNSP) and chronic wide spread pain (CWP) and in healthy pain-free controls. Furthermore for the CNSP the effect of training, which is a commonly recommended treatment for these patients, on the levels of endocannabinoids/NAEs was also investigated. Microdialysis technique in the trapezius muscle was used for sampling and masspectrometry was used for analysing. Two NAEs, N-palmitoylethanolamine (PEA) and N-stearoylethanolamine (SEA), could be repeatedly measured. The levels of these two lipids were significantly higher in CNSP compared to CON. The result showed also that PEA and SEA mobilize differently in CWP compared to both CNSP and CON. Taken together the results presented in thesis represent an early characterization of the pharmacology of MGL and provides novel information on NAEs in chronic muscle pain.

The endogenous cannabinoids (endocannabinoids) anandamide (AEA) and 2-arachidonylglycerol (2-AG) exert their effects predominantly through cannabinoid CB1 and CB2 receptors, but these actions are short-lived because of rapid hydrolysis by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), respectively. Selective inhibition of either enzyme elevates CNS levels of the appropriate endocannabinoid and produces analgesic effects with fewer psychomimetic side effects than Δ9-tetrahydrocannabinol (THC), the primary active constituent of marijuana. While cannabinoid receptor agonists and FAAH inhibitors reliably produce anti-inflammatory and anti-hyperalgesic effects in the carrageenan test and other inflammatory pain models, much less is known about the consequences of inhibiting MAGL in these assays. Here, we tested whether the selective MAGL inhibitor JZL184 would reduce nociceptive behavior in the carrageenan test. JZL184 significantly attenuated carrageenan-induced paw edema and mechanical allodynia, whether administered before or after carrageenan. Complementary genetic and pharmacological approaches revealed that JZL184’s anti-allodynic effects required both CB1 and CB2 receptors, but only CB2 receptors mediated its anti-edematous actions. Importantly, the anti-edematous and anti-allodynic effects of JZL184 underwent tolerance following repeated injections of high dose JZL184 (16 or 40 mg/kg), but repeated administration of low dose JZL184 (4 mg/kg) retained efficacy. Interestingly, the anti-allodynic effects of the combination of low dose of JZL184 (4mg/kg) and high dose of the selective and long-acting FAAH inhibitor PF-3845 (10 mg/kg) was augmented compared with each drug alone. On the contrary, the combination treatment did not reduce edema more than either JZL184 or PR-3845 given alone. These results suggest that low doses of MAGL inhibitors alone or in combination with FAAH inhibitors, reduce inflammatory nociception through the activation of both CB1 and CB2 receptors with no evidence of tolerance following repeated administration.

Les dérivés de l’opium (opioïdes) et du cannabis (cannabinoïdes) présentent de nombreuses propriétés intéressantes. Suite à l’identification de leurs récepteurs respectifs, diverses stratégies pharmacologiques ont tenté d’exploiter leurs propriétés analgésiques. Le clonage des récepteurs cannabinoïdes CB1 et CB2 a favorisé la découverte de composés endogènes pour ces récepteurs, les endocannabinoïdes, dont les deux plus étudiés sont l’anandamide et le 2-arachidonyl glycérol (2-AG). Cette découverte a également mené à l’identification d’enzymes qui catalysent l’inactivation de ces cannabinoïdes endogènes : une amidohydrolase des acides gras ou FAAH ainsi qu’une monoacylglycérol lipase ou MAGL. Le système cannabinoïde endogène est régulé à la hausse dans une variété de processus pathologiques, tels que les douleurs inflammatoire et neuropathique. Cette augmentation est habituellement interprétée comme une réaction physiologique visant à rétablir l’homéostasie et elle a notamment été observée en périphérie. Les endocannabinoïdes semblent donc agir de façon spécifique à des moments clés dans certains tissus ciblés afin de minimiser les conséquences reliées au déclenchement de ces douleurs. Cette observation est très intéressante d’un point de vue thérapeutique puisqu’elle suggère la possibilité de cibler les enzymes de dégradation des endocannabinoïdes dans le but d’augmenter leurs concentrations locales et d’ainsi prolonger leur action neuromodulatrice. En périphérie, l’activation des récepteurs cannabinoïdes induit des effets antinociceptifs bénéfiques tout en minimisant les effets indésirables souvent associés à leur activation centrale. Nous avons orienté nos travaux vers la modulation périphérique de ce système endogène à l’aide d’inhibiteurs des enzymes de dégradation des endocannabinoïdes afin d’évaluer leur potentiel thérapeutique et d’élucider les mécanismes d’action qui sous-tendent leurs effets dans des modèles animaux de douleurs inflammatoire et neuropathique. Nous avons démontré que cette approche permet de soulager les symptômes associés à ces deux types de douleurs, et ce via les récepteurs CB1 et CB2. Les systèmes cannabinoïde et opioïde présentent des similitudes, dont des localisations similaires le long des voies de la douleur, des mécanismes d’action relayés par des récepteurs couplés aux protéines G et des propriétés pharmacologiques communes telles que l’analgésie. Le système opioïde est impliqué dans les effets antinociceptifs induits par les cannabinoïdes. À l’inverse, le rôle joué par le système cannabinoïde dans ceux induits par la morphine demeure incertain. Nous avons démontré que les effets antinociceptifs périphériques et spinaux produits par la morphine sont diminués chez les souris génétiquement modifiées chez lesquelles l’expression des récepteurs CB1 ou CB2 a été éliminée, laissant supposer un rôle pour ces récepteurs dans les effets de la morphine. Nous avons de plus démontré que la diminution de l'analgésie produite par la morphine dans ces souris n'est pas causée par un dysfonctionnement des récepteurs opioïdes mu (MOP) ni par une régulation à la baisse de ces récepteurs. Nos résultats confirment l'existence d'interactions fonctionnelles entre les systèmes cannabinoïde et opioïde au niveau périphérique et spinal. Ces observations sont prometteuses d’un point de vue thérapeutique puisqu’une modulation périphérique ciblée des niveaux d’endocannabinoïdes et d’opioïdes endogènes permettrait de produire des effets analgésiques bénéfiques potentiellement synergiques tout en minimisant les effets indésirables associés à l’activation centrale de ces systèmes. / Opium (opioids) and cannabis (cannabinoids) derivatives present many interesting properties. Following the identification of their respective receptors, various pharmacological strategies have tried to exploit their analgesic properties. The cloning of cannabinoid CB1 and CB2 receptors has promoted the discovery of endogenous agonists of these receptors named endocannabinoids. The two mostly studied endocannabinoids are anandamide and 2-arachidonoyl glycerol (2-AG). This has also led to the identification of enzymes that catalyze the inactivation of these endogenous cannabinoids: a fatty acid amide hydrolase or FAAH and a monoacylglycerol lipase or MAGL. It is known that the endogenous cannabinoid system is upregulated in a variety of pathological processes, such as inflammatory and neuropathic pain. This increase is usually interpreted as a physiological response to restore homeostasis and it was particularly observed in the periphery. Endocannabinoids seem to act specifically at key moments in targeted tissues to minimize the consequences related to the onset of pain. This observation is very interesting from a therapeutic perspective because it suggests the possibility of targeting the endocannabinoid degrading enzymes in order to increase their local concentrations and thus prolong their neuromodulatory action. At the peripheral level, the activation of cannabinoid receptors induces beneficial antinociceptive effects while minimizing side effects often associated with their central activation. We focused our work on the peripheral modulation of this endogenous system using inhibitors of endocannabinoid degrading enzymes to assess their therapeutic potential and to elucidate the mechanisms of action underlying their effects in animal models of inflammatory and neuropathic pain. We have demonstrated that this approach can relieve the symptoms associated with these two types of pain, through the activation of CB1 and CB2 receptors. The opioid and cannabinoid systems have similarities, including comparable locations along the pain pathways, mechanisms of action relayed by G protein-coupled receptors and common pharmacological properties such as analgesia. The opioid system is involved in the antinociceptive effects induced by cannabinoids. In contrast, the participation of the cannabinoid system in those induced by morphine remains uncertain. We have demonstrated that peripheral and spinal antinociceptive effects induced by morphine are reduced in genetically modified mice in which the expression of CB1 and CB2 receptors was eliminated, suggesting a role for these receptors in the effects of morphine. We have further demonstrated that the decrease in morphine-induced analgesia in these mice is not caused by a malfunction of the mu opioid receptors (MOP) or by a down-regulation of these receptors. Our results confirm the existence of functional interactions between cannabinoid and opioid systems at the peripheral and spinal levels. These findings are promising from a therapeutic perspective since a targeted modulation of the levels of endocannabinoids and endogenous opioids would induce potentially synergistic beneficial analgesic effects while minimizing side effects associated with the central activation of these systems.