During the present reporting period, our research in this area was limited due to severe reductions in available research resources. Nevertheless, we carried out research in three distinct domains - first, continued research into the existence of cannabinoid CB2 receptors in the brain, second, preliminary experiments with a reward delay discounting behavioral model, and third, setting up an alcohol binge-drinking behavioral model in our laboratory. In the cannabinoid and endocannabinoid realm of research, the existence of cannabinoid CB2 receptors in the brain has been heretofore controversial. Most evidence has heretofore suggested that only CB1 cannabinoid receptors are found in brain and central nervous system while cannabinoid CB2 receptors are restricted to the body's periphery - primarily in the immune system. However, this view has been challenged by recent claims that CB2 receptors are present in the central nervous system and by recent claims that CB2 receptors modulate synaptic activity. Therefore, we used highly selective CB2 agonists and antagonists, combined with the use of CB1 and CB2 receptor gene-deleted mice, to study CB2 involvement in cocaine's behavioral and neurochemical effects. We found that the CB2 receptor-selective agonist JWH133 attenuates intravenous cocaine self-administration in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. This effect was abolished by the CB2 receptor-selective antagonist AM630. To confirm our findings, we also used the CB2-selective agonist GW405833 and found a similar inhibition of intravenous cocaine self-administration in wild-type mice. Under progressive-ratio reinforcement conditions, we found that JWH133 inhibits incentive motivation to self-administer cocaine, as evidenced by strong reductions in the progressive-ratio break-point. Similar effects were found when JWH133 was administered intra-nasally (for direct passage into the brain via the cribiform plate) or administered by direct intracerebral microinjections of JWH133 into the nucleus accumbens. Again, the effect was seen in wild-type but not in CB2 receptor gene-deleted mice. JWH133 by itself was found to have no reinforcing or aversive effects, as assessed by intravenous self-administration and by conditioned place preference/aversion experiments. Further, JWH133 inhibited cocaine-enhanced locomotion in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. JWH133 by itself had an inhibitory effect on locmotion, both with systemic administration and with intracerebral microinjection into the nucleus accumbens in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. The CB2 selective antagonist AM630 had a stimulatory effect on locomotion, both with systemic administration and with intracerebral microinjection into the nucleus accumbens in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. JWH133 by itself inhibited extracellular nucleus accumbens dopamine as measured by real-time in vivo brain microdialysis. JWH133 also inbited basal and cocaine-enhanced extracellular nucleus accumbens dopamine as measured by real-time in vivo brain microdialysis. This effect was blocked by the CB2-selective antagonist AM630. By itself, AM630 - microinjected intracerebrally into the nucleus accumbens - aumented basal extracellular nucleus accumbens dopamine. We conclude that CB2 cannabinoid receptors exist in the brain, that CB2 receptors functionally modulate the meso-accumbens dopamine system, that CB2 receptors functionally modulate dopamine-mediated behaviors, and that the brain CB1 and CB2 receptor-linked neural systems may functionally antagonize each other in a reciprocal mutually antatagonistic manner. Such mechanistic knowledge can aid in the search for new and effective pharmacotherapeutic compounds for the treatment of drug addiction and dependence. In addition, during this reporting period, we carried out preliminary experiments with a reward-delay-discounting animal model of addiction. At the human level, inability to delay gratification is a pathognomonic symptom of drug addiction. Therefore, we added the reward-delay-discounting task to our battery of preclinical animal models. In this task, laboratory rats are presented with two wall-mounted levers in their test chambers. Depression of one lever delivers a food reward immediately. Depression of the other lever delivers a larger reward after a delay period ranging up to 60 seconds. The animal must choose whether it desires a small immediate reward or a larger delayed reward. In this manner, impulsive choice can be measured and quantified. This model gives us an entirely new clinically-relevant perspective to evaluate potentially therapeutic anti-addiction anti-craving anti-relapse medications at the preclinical animal model level. Using this model during the reporting period, we found that laboratory rats self-assort themselves bimodally into animals with high impulsive choice and others with low impulsive choice. We further found that in the high impulsive choice animals, cocaine produces a shift toward less impulsivity. In contrast, in the low impulsive choice animals, cocaine produces a shift toward higher impulsivity. We intend to further explore the effects of addictive drugs on impulsive choice behavior using this new animal model. In addition, during this reporting period, we introduced an alcohol binge-drinking animal model into our battery of preclinical animal models of addiction. At the human level, binge-type drug taking behavior is often characteristic of drug addiction. Therefore, we have added an alcohol binge-drinking animal model. In this task, laboratory rats are allowed to drink water, near beer (containing no ethanol), or beer (containing 2% ethanol) for 2 hours per day. Binge drinking is operationally defined as bringing blood ethanol levels up to at least 80mg/dl within 2 hours of the initial drink. It is our intention to use this new animal model to determine whether any of our putative anti-addiction pharmacotherapies can alter binge drinking in a putatively therapeutic direction. We believe that the addition of this new model gives us yet another new clinically-relevant perspective to evaluate potentially therapeutic anti-addiction anti-craving anti-relapse medications at the preclinical animal model level. If successful, we intend to investigate binge consumption of other addictive drugs.
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