endocannabinoid roles in neurochemical and reinforcing effects of abused drugs

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2/2/13 NIH Annual Report DA000569

nidb.nih.gov/search/searchview.taf?ipid=75507

NIH Annual Intramural Research Report

DA000569-03

Report Title

Endocannabinoid roles in neurochemical and reinforcing effects of abused drugs

2012 Fiscal Year

October 01, 2011 - September 30, 2012

Principal Investigator

Jonathan L. Katz PhD

Research Organization

Psychobiology Section, NIDA

Lab Staff

Gianluigi Tanda PhDMaddalena Mereu

Takato Hiranita

Jessica Patricia Lopez

Keywords

Cannabinoid receptors and addiction, cannabinoids, dopamine, Nucleus Accumbens

Goals and Objectives

The main goal of this proposal is to provide a better understanding of the involvement of the

brain endocannabinoid system in the effects of drugs abused by humans tested in preclinical

procedures. The focus is on behavioral and neurochemical effects of drugs acting at

cannabinoid receptors, or that modulate brain endocannabinoid levels. We are also testing theinvolvement of the endocannabinoid system in the behavioral and neurochemical effects

produced by selected drugs of abuse, cocaine, amphetamine, methamphetamine, heroin,

delta-9-tetrahydrocannabinol (THC), ethanol, and nicotine. These studies will provide valuable

information about cannabinoid brain functions related to drug abuse and addiction, and

cannabinoid-based pharmacological interventions/strategies against drug dependence.

Cannabinoids, initially thought to act through non-specific interactions related to cell

membrane fluidity, were shown to activate specific cannabinoid receptors. Two types of these

receptors exist, CB1, most expressed in the brain, and CB2 mostly expressed in the periphery.

Cannabinoids have been shown to activate other receptors, for example, TRPV1, PPAR, and

GPR55. Different endogenous compounds bind to cannabinoid receptors, among them,

anandamide (AEA), and 2-arachidonyl-glycerol (2AG). These compounds are not stored in

vesicles, like other transmitters, but are released on demand by cleavage of membrane

precursor components. The recent availability of natural and synthetic ligands, and drugs that

block the metabolism or the uptake of endocannabinoids, has facilitated the study of the role

of cannabinoids in physiological functions, for example in synaptic plasticity. Dysfunctions of

the endocannabinoid system could also result in neurological disorders, including drug

addiction.

Summary

We have shown that the endocannabinoids AEA (and its metabolically stable analogue

methAEA) and 2AG share with drugs abused by humans the ability to acutely stimulate DA

levels in the rat NAC shell, suggesting endocannabinoids may possess reinforcing effects.

Indeed, AEA promotes and maintains intravenous self-administration behavior in monkeys.

These effects are blunted by pretreatment with a selective CB1 antagonist, indicating a direct


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CB1 receptor involvement in elevation of DA and reinforcing effects. Compounds known to

increase brain endocannabinoid levels may also elicit DA release. URB597 inhibits the enzyme

Fatty Acid Amide Hydrolase (FAAH) that metabolizes AEA. This drug alone (at doses that fully

block FAAH) did not produce DA stimulation, suggesting that, in animals at rest, levels of AEA

are not high enough to activate CB1 receptors, even after FAAH blockade. Alternatively, FAAH

might not be sufficiently expressed in areas related to DA transmission. However, URB597

enhanced the effects of exogenous AEA on DA levels, supporting the hypothesis of low levels

of AEA in animals at rest, and suggesting that URB597 can block AEA metabolism resulting in

increased levels of AEA in areas related to DA transmission. URB597 alone does not elevate DA

levels, and its degree of abuse liability appears low, in agreement with negative findings of self-

administration in monkeys, and place preference in rats. AM404, another endocannabinoidenhancer, produces cannabinoid-like behaviors in rodents, but in our experiments it did not

alter DA levels, and surprisingly, did not enhance the effect of AEA on NAC shell DA. In

agreement, in rats trained to discriminate THC from saline (a behavior selectively mediated by

CB1 receptors), AEA alone did not produce full THC-like effects (likely due to its rapid

metabolism), while after pretreatments with URB597, but not with AM404, AEA produced full

THC-like effects.

DA D2 receptor activation stimulates AEA levels, thus, drugs that increase DA levels in the

brain, like abused drugs, can 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 in specific brain areas 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 brain actions of anandamide and blockers of anandamide

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 anandamidemetabolism 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,

using self-administration and reinstatement behavioral models of nicotine dependence.

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 thisproject 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


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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.

We are also studying the role of CB1 receptors on the reinforcing effects of food. This studyuses genetically modified CB1-receptor KO mice and their wild-type littermates. Mice are

trained to emit operant responses to get a food reward. As the number of required responses

is increased, food consumption decreases. This demand function is derived from behavioral

economic theory, and has been validated as a behavioral index of reinforcing efficacy. Studies

are underway to evaluate the role of CB1 receptors in the value of specific reinforcers using

these genetically engineered mice.

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 are going

to test drugs like JHW007, a DAT blocker that also show allosteric CB1 activity, in preclinical

procedures that would evaluate the cocaine antagonist effects of drugs when administered

together with cocaine. For example, we could test some behavioral/ reinforcing effects of

cocaine under conditions in which CB1 receptors have been deleted, thus ruling out any

possible interference of allosteric CB1 modulation induced by the test compounds.

Publications Generated during the 2012 Reporting Period

Ordered by author name.

1. Mascia P, Pistis M, Justinova Z, Panlilio LV, Luchicchi A, Lecca S, Scherma M, Fratta W,

Fadda P, Barnes C, Redhi GH, Yasar S, Le Foll B, Tanda G, Piomelli D, Goldberg SR (2011)

Blockade of nicotine reward and reinstatement by activation of alpha-type peroxisomeproliferator-activated receptors. Biol Psychiatry 69:633-41.

[PMID 20801430] [PMC 2994947]

endocannabinoid roles in neurochemical and reinforcing effects of abused drugs

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