2-Arachidonoylglycerol (2-AG) and anandamide are endocannabinoids that activate cannabinoid receptors CB1 and CB2. Endocannabinoid signaling is terminated by enzymatic hydrolysis, a process that, for anandamide, is mediated by fatty acid amide hydrolase (FAAH) and, for 2-AG, is thought to involve monoacylglycerol lipase (MAGL). FAAH inhibitors produce a select subset of the behavioral effects observed with CB1 agonists, intimating a functional segregation of endocannabinoid signaling pathways in vivo. Testing this hypothesis, however, requires specific tools to independently block anandamide and 2-AG metabolism. Here, we report a potent and selective inhibitor of MAGL, JZL184, that, upon administration to mice, raises brain 2-AG by 8-fold without altering anandamide. JZL184-treated mice exhibited a broad array of CB1-dependent behavioral effects, including analgesia, hypothermia, and hypomotility. These data indicate that 2-AG endogenously modulates several behavioral processes classically associated with the pharmacology of cannabinoids and point to overlapping and unique functions for 2-AG and anandamide in vivo.

Prolonged exposure to drugs of abuse, such as cannabinoids and opioids, leads to pharmacological tolerance and receptor desensitization in the nervous system. Here we show that a similar form of functional antagonism is produced by sustained inactivation of monoacylglycerol lipase (MAGL), the principal degradative enzyme for the endocannabinoid 2-arachidonoylglycerol (2-AG). After repeated administration, the MAGL inhibitor JZL184 lost its analgesic activity and produced cross-tolerance to cannabinoid receptor (CB1) agonists in mice, effects that were phenocopied by genetic disruption of MAGL. Chronic MAGL blockade also caused physical dependence, impaired endocannabinoid-dependent synaptic plasticity, and desensitization of brain CB1 receptors. These data contrasted with blockade of fatty acid amide hydrolase (FAAH), an enzyme that degrades the other major endocannabinoid anandamide, which produced sustained analgesia without impairing CB1 receptors. Thus, individual endocannabinoids generate distinct analgesic profiles that are either sustained or transitory and associated with agonism and functional antagonism of the brain cannabinoid system, respectively.

⌬ 9 -Tetrahydrocannabinol (THC), the psychoactive component of marijuana, and other direct cannabinoid receptor (CB1) agonists produce a number of neurobehavioral effects in mammals that range from the beneficial (analgesia) to the untoward (abuse potential). Why, however, this full spectrum of activities is not observed upon pharmacological inhibition or genetic deletion of either fatty acid amide hydrolase (FAAH) or monoacylglycerol lipase (MAGL), enzymes that regulate the two major endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG), respectively, has remained unclear. Here, we describe a selective and efficacious dual FAAH/MAGL inhibitor, JZL195, and show that this agent exhibits broad activity in the tetrad test for CB1 agonism, causing analgesia, hypomotilty, and catalepsy. Comparison of JZL195 to specific FAAH and MAGL inhibitors identified behavioral processes that were regulated by a single endocannabinoid pathway (e.g., hypomotility by the 2-AG/MAGL pathway) and, interestingly, those where disruption of both FAAH and MAGL produced additive effects that were reversed by a CB1 antagonist. Falling into this latter category was drug discrimination behavior, where dual FAAH/MAGL blockade, but not disruption of either FAAH or MAGL alone, produced THC-like responses that were reversed by a CB1 antagonist. These data indicate that AEA and 2-AG signaling pathways interact to regulate specific behavioral processes in vivo, including those relevant to drug abuse, thus providing a potential mechanistic basis for the distinct pharmacological profiles of direct CB1 agonists and inhibitors of individual endocannabinoid degradative enzymes.hydrolase ͉ inhibitor ͉ metabolism N -arachidonoyl ethanolamine (anandamide or AEA) (1) and 2-arachidonoylglycerol (2-AG) (2, 3) are lipid transmitters that serve as endogenous ligands for the cannabinoid G-proteincoupled receptors CB1 and CB2. These lipids and receptors, along with the enzymes that biosynthesize and degrade AEA and 2-AG, form the endogenous cannabinoid (endocannabinoid) system, which regulates a diverse number of physiological processes in mammals, including pain, cognition, emotionality, neurodegeneration, feeding, and inflammation (4).CB1 and CB2 receptors are also activated by ⌬ 9 -tetrahydrocannabinol (THC), the psychoactive component of marijuana (4). Most of the neurobehavioral effects of THC and other direct cannabinoid receptor agonists are mediated by the CB1 receptor (5, 6), likely reflecting its widespread and abundant expression in the nervous system (7,8). CB1 agonism produces medicinally useful activities, such as analgesia, but also a number of undesirable side effects, including locomotor and cognitive impairments, as well as abuse liability. To date, it has proved difficult to uncouple these beneficial and untoward properties, thus limiting the therapeutic utility of direct CB1 agonists.Inhibitors of AEA and 2-AG degradation offer a potentially attractive alternative strategy to stimulate the endocannabinoid system (9-12). Indeed, d...

BackgroundClinical studies of osteoarthritis (OA) suggest central sensitization may contribute to the chronic pain experienced. This preclinical study used the monosodium iodoacetate (MIA) model of OA joint pain to investigate the potential contribution of spinal sensitization, in particular spinal glial cell activation, to pain behaviour in this model. Experimental OA was induced in the rat by the intra-articular injection of MIA and pain behaviour (change in weight bearing and distal allodynia) was assessed. Spinal cord microglia (Iba1 staining) and astrocyte (GFAP immunofluorescence) activation were measured at 7, 14 and 28 days post MIA-treatment. The effects of two known inhibitors of glial activation, nimesulide and minocycline, on pain behaviour and activation of microglia and astrocytes were assessed.ResultsSeven days following intra-articular injection of MIA, microglia in the ipsilateral spinal cord were activated (p < 0.05, compared to contralateral levels and compared to saline controls). Levels of activated microglia were significantly elevated at day 14 and 21 post MIA-injection. At day 28, microglia activation was significantly correlated with distal allodynia (p < 0.05). Ipsilateral spinal GFAP immunofluorescence was significantly (p < 0.01) increased at day 28, but not at earlier timepoints, in the MIA model, compared to saline controls. Repeated oral dosing (days 14-20) with nimesulide attenuated pain behaviour and the activation of microglia in the ipsilateral spinal cord at day 21. This dosing regimen also significantly attenuated distal allodynia (p < 0.001) and numbers of activated microglia (p < 0.05) and GFAP immunofluorescence (p < 0.001) one week later in MIA-treated rats, compared to vehicle-treated rats. Repeated administration of minocycline also significantly attenuated pain behaviour and reduced the number of activated microglia and decreased GFAP immunofluorescence in ipsilateral spinal cord of MIA treated rats.ConclusionsHere we provide evidence for a contribution of spinal glial cells to pain behaviour, in particular distal allodynia, in this model of osteoarthritic pain. Our data suggest there is a potential role of glial cells in the central sensitization associated with OA, which may provide a novel analgesic target for the treatment of OA pain.

1 Marijuana's appetite-increasing effects have long been known. Recent research suggests that the CB 1 cannabinoid receptor antagonist SR141716A may suppress appetite. This study represents a further, systematic investigation of the role of CB 1 cannabinoid receptors in the pharmacological effects of cannabinoids on food intake. 2 Mice were food-restricted for 24 h and then allowed access to their regular rodent chow for 1 h. Whereas the CB 1 antagonist SR141716A dose-dependently decreased food consumption at doses that did not affect motor activity, D 9 -tetrahydrocannabinol (D 9 -THC) increased food consumption at doses that had no effect on motor activity. O-3259 and O-3257, structural analogs of SR141716A, produced effects similar to those of the parent compound. 3 Amphetamine (a known anorectic) and diazepam (a benzodiazepine and CNS depressant) decreased food consumption, but only at doses that also increased or decreased motor activity, respectively. The CB 2 cannabinoid receptor antagonist SR144528 and the nonpsychoactive cannabinoid cannabidiol did not affect food intake nor activity. 4 SR141716A decreased feeding in wild-type mice, but lacked pharmacological activity in CB 1 knockout mice; however, basal food intake was lower in CB 1 knockout mice. Amphetamine decreased feeding in both mouse genotypes. 5 These results suggest that SR141716A may affect the actions of endogenous cannabinoids in regulating appetite or that it may have effects of its own aside from antagonism of cannabinoid effects (e.g., decreased feeding behavior and locomotor stimulation). In either case, these results strongly suggest that CB 1 receptors may play a role in regulation of feeding behavior.