DA D2 receptor activation stimulates AEA levels, thus, drugs that increase DA levels in the brain, like abused drugs, activate DA D2 receptors, and in turn, increase AEA levels. We have tested the effects of abused drugs in producing CB1-receptor mediated generalization in THC-discrimination tests. Cocaine, and amphetamine injected alone did not produce effects significantly different from vehicle, but potentiated the THC-like effects of THC. Nicotine and the D2/3 DA receptor agonist quinpirole alone did not generalize to the THC cue, but both drugs did so in animals pretreated with the inhibitor of FAAH that metabolizes AEA. Nicotine and quinpirole also potentiated the effects of THC. We have suggested that AEA is released by these drugs by a D2 receptor mediated mechanism. So, administered alone these drugs do not stimulate AEA levels sufficiently to provide CB1-mediated THC-like effects, but potentiate ineffective small doses of THC. However, when the same drugs are administered in combination with URB-597, AEA levels are magnified by blockade of its metabolism, and its concentration could thus activate CB1 receptors producing THC discriminative effects. Recently it has also been shown that effects of anandamide and blockers of its metabolism (e.g. URB-597) might be mediated not only by the endocannabinoid system, but also by PPAR-alpha receptors. It has also been demonstrated that blockade of anandamide metabolism through fatty acid amide hydrolase enzymes by URB-597 might lead to increased levels of oleoylamide (OEA) or palmytoilamide (PEA), as well as anandamide. While anadamide has both CB1 and PPAR-alpha receptor activities, OEA and PEA are selective ligands for PPAR receptors, with negligible activity at CB1 receptors. We showed that drugs acting specifically at brain PPAR-alpha receptors can block the addictive actions of nicotine in rats and monkeys. Psychostimulant sensitization might play a role in the path to abuse and addiction, and even a single exposure to psychostimulants could produce long-term sensitization by increasing strength of excitatory synapses in midbrain dopaminergic regions. Sensitization can be viewed as a type of synaptic plasticity, which is also related to alterations in the cannabinoid system. We hypothesized that development of psychostimulant sensitization might involve stimulation of brain endocannabinoid levels that can bind to and activate CB1 receptors. We started this project studying cocaine sensitization in mice, measured as increased stimulation of behavioral activities before and after sensitizing doses of cocaine. We are testing the hypothesis that the development of cocaine sensitization requires release of endocannabinoids, and can be reversed by CB1 receptor blockade. Based on our original hypothesis, low doses of cocaine that do not induce behavioral sensitization might become effective when animals are pretreated with enhancers of endocannabinoid levels. DA transmission, believed to mediate behavioral and reinforcing effects of cocaine, will also be measured before and after cocaine sensitization. Our results have confirmed that a single exposure to cocaine induces behavioral sensitization in mice. Rimonabant, a CB1 antagonist, injected before the sensitizing dose of cocaine, reduced the sensitization produced by cocaine. In addition, cocaine-induced sensitization was paralleled by a sensitized, larger stimulation of DA levels, compared to saline treated animals in the nucleus accumbens core, but not in the NAC shell. Our results suggest also that blockade of endocannabinoid metabolism (obtained by pretreatment with URB-597 in mice) enhances the extracellular levels of endocannabinoids released by cocaine, and this enhancement could be related to the induction of behavioral sensitization by doses of cocaine otherwise not effective in inducing behavioral or neurochemical sensitization. We have also found that the enhancement of cocaine-induced anandamide levels in the brain will also result in a specific related neurochemical sensitization of DA stimulation in the core but not in the shell of the nucleus accumbens. It has been recently suggested that selected blockers of the dopamine transporter might possess the ability to negatively interact with an allosteric site of the cannabinoid CB1 receptors. It has also been suggested that this negative allosteric modulation of CB1 receptors might be important in these DAT blockers for expressing antagonism to the behavioral/reinforcing effects of cocaine. To test the validity of these suggestions, we tested antagonism of cocaine by JHW007, a DAT blocker that also show allosteric CB1 activity. Antagonism of cocaine was obtained in both CB1 knockout and wild-type mice suggesting that allosteric modulation of CB1 receptor actions has a minimal role in the cocaine-antagonist effects of JHW 007. The number of people seeking treatment for marijuana use in the United States per year (1,243,000) is higher than the number seeking treatment for cocaine or heroin use (787,000 or 507,000, respectively)1. THC, the main psychoactive ingredient in marijuana, activates brain pathways mediating its reinforcing effects by enhancing the firing of DA neurons in the ventral tegmental area (VTA), resulting in increased release of DA from nerve terminals in the shell of the nucleus accumbens (NAc). Developing medications that modulate these effects of THC as a reinforcer might provide a therapeutic approach for the treatment of marijuana dependence. For example, previously we found that reward-related behavioral and neurochemical effects of THC could be blocked by methyllycaconitine (MLA), a selective antagonist of α7nAChRs that are present in both the VTA and the NAc shell on glutamatergic nerve terminals. Their activation elicits glutamate release, which in turn acts at ionotropic glutamate receptors on dopaminergic terminals to stimulate dopamine release. Unfortunately, systemic use of direct antagonists of α7nAChRs is associated with side effects that limit their therapeutic utility. To avoid these unwanted effects, we tested a compound 3,4-dimethoxy-N-4-(3-nitrophenyl)thiazol-2-ylbenzenesulfonamide (Ro 61-8048), a potent, selective, peripherally acting kynurenine 3-monooxygenase (KMO) inhibitor, to indirectly increase brain KYNA, an endogenous negative allosteric modulators of α7nAChRs that might be better tolerated than directly acting cholinergic antagonists. Indeed, allosteric modulators change receptor conformations in the presence of orthosteric ligands and often have no effect on their own, acting only when physiological receptors are activated. Newly formed KYNA is promptly released into the extracellular compartment. Notably, no reuptake processes exist for KYNA, and extracellular KYNA is not degraded enzymatically but is slowly eliminated from the brain by a non-specific acid transporter. We found that administration of the kynurenine 3-monooxygenase (KMO) inhibitor Ro 61-8048 increases brain KYNA levels and attenuates THC-induced stimulation of extracellular levels of dopamine in reward-related brain areas. Administration of Ro 61-8048 also reduced the reinforcing effects of THC measured under self-administration behavioral procedures, also preventing relapse to drug-seeking induced by re-exposure to cannabinoids or cannabinoid-associated cues. Pharmacologic proof of involvement of α7nAChRs was obtained by administration of positive allosteric modulators of α7nAChRs. These results suggest a therapeutic strategy for treatment of marijuana dependence.
