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Neurobiology of Disease 5, 502–533 (1998)
Article No. NB980219
Cannabinoid Transmission
and Reward-Related Events
Eliot L. Gardner1 and S. Robert Vorel
Department of Psychiatry and Department of Neuroscience,
Albert Einstein College of Medicine, New York, New York 10461-1602
The reward/reinforcement circuitry of the mammalian brain consists of synaptically interconnected
neurons associated with the medial forebrain bundle, linking the ventral tegmental area, nucleus
accumbens, and ventral pallidum. Electrical stimulation of this circuit supports intense self-
stimulation in animals and, in humans, produces intense pleasure or euphoria. This circuit is strongly
implicated in the neural substrates of drug addiction and in such addiction-related phenomena as
withdrawal dysphoria and craving. This circuit is also implicated in the pleasures produced by natural
rewards (e.g., food, sex). Cannabinoids are euphorigenic in humans and have addictive liability in
vulnerable persons, but were long considered ‘‘anomalous’’ drugs of abuse, lacking pharmacological
interaction with these brain reward substrates. It is now clear, however, that cannabinoids activate
these brain substrates and influence reward-related behaviors. From these actions, presumably,
derive both the abuse potential of cannabinoids and the possible clinical efficacy in dysphoric
states.
1998 Academic Press
Key Words: cannabis; cannabinoid; marijuana; reward; reinforcement; brain stimulation; ICSS;
microdialysis; place preference; dopamine; nucleus accumbens; medial forebrain bundle; ventral
tegmental area; addiction; dependence; drug abuse.
The past decade has witnessed a thoroughly remark-
able advance in our understanding of the action of
marijuana, other cannabis constituents, and cannabi-
noid-like drugs on brain and behavior (1–3). This
cascade of increased understanding stems in large
measure from the discovery that cannabinoids interact
with specific G-protein-coupled brain receptors to
modulate neuronal function and behavior (4–10). It is
now clear that cannabinoids act through both a central
(11) cannabinoid receptor (termed the CB1 receptor)
and a peripheral (12) receptor (CB2), both of which
have been characterized at the molecular level (11–13).
The CB1 receptor is widely distributed throughout the
brain, with especially high densities in the basal
ganglia, hippocampus, and cerebellum (14,15). Worthy
of note is the fact that CB1 receptors occur in signifi-
cant densities in brain areas associated with reward
and reinforcement functions, including the ventral
striatum and ventral mesencephalon (14,15) (see below
for implication of these areas in brain reward func-
502
tions). Functional variants and isoforms of these endog-
enous cannabinoid receptors may well exist in the
body (16,17). An exceedingly wide variety of selective
ligands with partial or full agonist action at cannabi-
noid receptors has been developed (18–27) (many by
Pfizer Central Research and by Sanofi Winthrop, Inc.),
allowing a formidable array of pharmacological agents
to probe cannabinoid receptor-mediated neuronal
events and behaviors, as well as conducting structure–
activity studies. Highly selective and potent antago-
nists have also been developed for both the CB1 (28,29)
and the CB2 (30) receptors. At least two endogenous
ligands, arachidonyl ethanolamide (anandamide)
(3,31,32) and 2-arachidonylglycerol (3,32–34), have been
identified as binding to cannabinoid receptors and
fulfilling requirements for neurotransmitter or neuro-
modulator functions.Anandamide—discovered in 1992
and thus studied longer—has in particular been shown
to bind to cannabinoid receptors, to be synthesized
and released by neurons, to be subject to a high-affinity
reuptake process, and to be enzymatically degraded in
1To whom correspondence and reprint requests should be ad-
brain (35–39). When administered to laboratory ani-
dressed. Fax: (718) 430-8772. E-mail: gardner@aecom.yu.edu.
mals, anandamide shows a classic constellation of
0969-9961/98 $25.00
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
Cannabinoids and Brain Reward 503
cannabinoid effects—including locomotor inhibition,
COMMON PROPERTIES
hypothermia, analgesia, and memory impairment
(40–43). Yet more endogenous ligands of the anan-
OF HABIT-FORMING DRUGS
damide family may well exist in brain and function as
Evidence that recreational and abuse-prone drugs
neurotransmitters or neuromodulators (44,45). In addi-
derive their rewarding properties by activating brain
tion, other endogenous cannabinoid ligands—not
reward circuits either directly or indirectly was pre-
chemically related to the anandamides—may well also
sented as early as 1957 by Killam and his colleagues
exist in brain and function as neurotransmitters or
(57). In the years since, many different lines of evi-
neuromodulators (44,46).
dence have converged to confirm this hypothesis (see
Cannabinoid-containing botanical preparations
Ref. 58 for extensive review). First, virtually all ad-
(marijuana, hashish) are some of the oldest psychoac-
equately studied recreational and abuse-prone drugs
tive substances known to mankind (47). Among the
(including those in such disparate chemical and phar-
many pharmacological effects produced by cannabi-
macological classes as opiates, stimulants, sedative-
noids is a dose-dependent euphoric high, which in
natural cannabinoid preparations such as marijuana
and hashish derives primarily from the psychoactive
constituent 9-tetrahydrocannabinol ( 9-THC). From
this euphorigenic property derives marijuana’s use as
a recreational drug and its habit-forming and abuse
potential. Estimates of current marijuana users among
the U.S. population range from 12 to 20 million (48,49).
Among teenagers and young adults, approximately
45% of all current U.S. 12th-grade students have used
cannabis (50), and marijuana currently serves (along
with ethanol) in both North America and Europe as a
‘‘gateway drug,’’introducing individuals (often teenag-
ers) to the use of euphoriant drugs, from which some
then progress to more addicting and destructive drugs
such as cocaine and heroin (48,51). In fact, Crowley
and colleagues have recently shown that, among teen-
agers (especially those with conduct disorder, atten-
tion deficit/hyperactivity disorder, or major depres-
sion), progression from first cannabis use to regular
cannabis use/dependence is as rapid as tobacco pro-
gression and more rapid than that for alcohol (52).
Outright cannabis addiction, with obsessional drug-
seeking and compulsive drug-taking behavior, is rela-
tively rare with low-dose cannabis preparations (e.g.,
hypnotics, ethanol, anxiolytics, and anesthetics) en-
hance brain-stimulation reward or lower brain reward
thresholds (58–61). Second, virtually all adequately
studied recreational and abuse-prone drugs enhance
basal neuronal firing and/or basal neurotransmitter
release in reward-relevant brain circuits (58,62–66).
Third, laboratory animals will work for microinjec-
tions of abused drugs into brain-reward loci, but not
into other brain loci (58,67–71). Fourth, lesions or
pharmacological blockade of brain reward circuits
markedly inhibit the rewarding properties of systemi-
cally administered drugs of abuse (58,72,73). Thus,
acute enhancement of brain reward mechanisms ap-
pears to be the single essential commonality of abuse-
prone drugs, and the hypothesis that recreational and
abused drugs act on these brain mechanisms to pro-
duce the subjective reward that constitutes the ‘‘high’’
or ‘‘rush’’ or ‘‘hit’’ sought by drug users is, at present,
the most compelling hypothesis available on the neuro-
biology ofrecreationaldrug useand abuse (58–61,64,74–
76). In addition, compelling evidence now exists that
aberrations within these brain reward circuits may
confer vulnerability to drug addiction and dependence
(77–81). At the same time, the involvement of brain
reward systems in mediating drug craving, relapse,
and vulnerability to drug abuse may well be complex
marijuana) but common with high-dose preparations
and multifaceted (58,82–85).
(e.g., hashish). Furthermore, physical dependence on
cannabis and physical withdrawal upon cannabis ces-
sation are clearly documented (52). Thus, cannabi-
NEUROANATOMY, NEUROPHYSIOLOGY,
noids can be categorized as abusable and habit-
AND NEUROCHEMISTRY
forming drugs and are currently the most commonly
used illicit drugs in North America and Europe (48),
OF BRAIN REWARD
with as many as 9% of current users meeting criteria
The present senior author has recently reviewed the
for substance dependence (53) and as many as 25% of
current understanding of the neuroanatomical, neuro-
heavy users meeting criteria for substance dependence
physiological, and neurochemical substrates of brain
(54). Not irrelevantly, a significant percentage of heavy
reward mechanisms (58) and the interested reader is
users appears to suffer significant deleterious residual
referred to that review, as a detailed explication of this
health consequences (54–56).
topic is beyond the scope of the present paper.
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
504 Gardner and Vorel
FIG. 1. Schematic diagram of the brain reward circuitry of the mammalian (laboratory rat) brain, with sites at which various
abusable
substances appear to act to enhance brain reward and thus to induce drug-taking behavior and possibly drug craving. ‘‘ICSS’’
indicates the
descending, myelinated, moderately fast-conducting component of the brain reward circuitry that is preferentially activated by
electrical
intracranial self-stimulation. ‘‘DA’’ indicates the subcomponent of the ascending mesolimbic dopaminergic system that appears
preferentially
activated by abusable substances. ‘‘Raphe´’’ indicates the brain-stem serotonergic raphe´ nuclei, ‘‘LC’’ indicates the locus
coeruleus, ‘‘VTA’’
indicates the ventral tegmental area, ‘‘Acc’’ indicates the nucleus accumbens, ‘‘VP’’ indicates the ventral pallidum, ‘‘ABN’’
indicates the anterior
bed nuclei of the medial forebrain bundle, ‘‘AMYG’’ indicates the amygdala, and ‘‘FCX’’ indicates the frontal cortex. ‘‘5HT’’
indicates the
serotonergic (5-hydroxytryptamine) fibers which originate in the anterior raphe´ nuclei and project to both the cell body region
(ventral
tegmental area) and the terminal projection field (nucleus accumbens) of the DAreward neurons. ‘‘NE’’ indicates the
noradrenergic fibers which
originate in the locus coeruleus and synapse into the general vicinity of the ventral mesencephalic DA cell fields of the ventral
tegmental area.
‘‘GABA’’ indicates the GABAergic inhibitory fiber systems synapsing upon the locus coeruleus noradrenergic fibers, the ventral
tegmental area,
and the nucleus accumbens, as well as the GABAergic outflow from the nucleus accumbens. ‘‘Opioid’’ indicates the endogenous
opioid peptide
neural systems synapsing into both the ventral tegmental DA cell fields and the nucleus accumbens DA terminal projection
loci. ‘‘ENK’’
indicates the enkephalinergic outflow from the nucleus accumbens. ‘‘DYN’’ indicates the dynorphinergic outflow from the
nucleus accumbens.
‘‘GLU’’ indicates the glutamatergic neural systems originating in frontal cortex and synapsing in both the ventral tegmental area
and the nucleus
accumbens. (Reprinted, with permission of the publisher, from Ref. 58.)
Suffice it to say that, at best present levels of
understanding, the core reward-related circuitry of the
mammalian brain appears to consist of a synaptically
interconnected series of neuronal tracts closely associ-
ated with the medial forebrain bundle (Fig. 1). The
‘‘first-stage’’ reward-related neurons appear to origi-
nate diffusely within the anterior ventral limbic fore-
brain—in the anterior bed nuclei of the medial fore-
brain bundle(anterior lateral hypothalamus, horizontal
limb of the diagonal band of Broca, interstitial nucleus
of the stria medullaris, lateral preoptic area, magnocel-
lular preoptic nucleus, olfactory tubercle, substantia
innominata, and ventral pallidum), presumably consti-
tuting an anatomic convergence of disparate neurally
encoded information critical to the set point of hedonic
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.

tone. These first-stage reward neurons run posteriorly


within the medial forebrain bundle in a myelinated
moderately fast-conducting pathway of unknown neu-
rotransmitter type and synapse on dopamine (DA)
cells in the ventral tegmental area of the ventral
mesencephalon. These first-stage reward neurons ap-
pear preferentially activated by electrical brain stimu-
lation reward or ‘‘intracranial self-stimulation’’ (ICSS).
The ‘‘second-stage’’ DA neurons project anteriorly
within the medial forebrain bundle to the nucleus
accumbens, where they make synaptic connections
with a variety of cell types. It is on this second-stage
DAconvergence—with itsDAcell bodies in the ventral
tegmental area and DA axon terminals in the nucleus
accumbens—that abusable substances appear to act to

Amphetamine Cocaine Opiates Cannabinoids Phencyclidine Ketamine


ICSS
Cannabinoids and Brain Reward 505
enhance brain reward functions and produce the
hedonic tone. The neurotransmitters of some of these
pleasurable and/or euphoric effects that constitute the
additional modulatory circuits are known—including
‘‘high’’or ‘‘rush’’ soughtby substance abusers. Further-
opioid peptides, serotonin, glutamate, and GABA.
more, it seems likely that only a small subset of these
Complicating this picture is recent evidence (again,
DA neurons are specialized for carrying reward-
reviewed in Ref. 58) that these reward-related neurons
relevant information. From the nucleus accumbens,
may be functionally heterogeneous—with some neu-
‘‘third-stage’’ reward-relevant neurons appear to carry
rons encoding reward magnitude per se while others
the reward/reinforcement neural signal further. This
encode expectancy of reward, errors in reward predic-
third-stage pathway uses the endogenous opioid pep-
tion, prioritized reward, and other more complex as-
tide enkephalin as its primary neurotransmitter and
pects of reward-driven learning and reward-related
projects anatomically to the ventral pallidum. Electro-
incentive motivation. While these complexities of func-
physiologic studies show that a large majority of
tion within the reward substrates of the forebrain do
nucleus accumbens output neurons are opiate-sensi-
appear to exist, it is equally clear that one of the
tive and naloxone-blockade, tending to confirm that
primary functions of those reward substrates is to
this is the major efferent pathway of the accumbens
compute hedonic tone and neural ‘‘payoffs,’’ that this
and tending to confirm its opioid nature. This third-
computation takes place in large measure within the
stage output pathway appears critical for the expres-
medial forebrain bundle (MFB)-associated circuits de-
sion of reward-related and incentive-related behaviors
lineated above, that the second-stage DAcomponent is
(as is the next-following transsynaptic pathway from
the common site of action for addicting drugs and
the ventral pallidum to the pedunculopontine tegmen-
crucial to their addictive features, that drug reward per
tal nucleus). Manipulationsof this nucleus accumbens–
se and drug potentiation of electrical brain stimulation
ventral pallidum pathway have clear effects on brain-
reward have common mechanisms, and that electrical
stimulation reward and reward-driven behaviors,
including drug-seeking behavior in animals. Mutually
reciprocal anatomic interconnections exist between the
ventral pallidum, the nucleus accumbens, and the
ventral tegmental area, which appear functionally
relevant to the set-point of reward functions and
reward-driven behaviors. Another nucleus accumbens
output pathway—the medium spiny output neurons
which use -aminobutyric acid (GABA)asaneurotrans-
mitter—may constitute yet another brain reward final
common output path, in which the critical event is
inhibition of the GABAergic medium spiny output
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
brain stimulation reward and the pharmacological
rewards of addicting drugs are habit-forming because
they act in the brain circuits that subserve more
natural, biologically significant rewards (86–89).
Of crucial importance to the present discussion is the
fact that the second-stage DAcomponent appears to be
the crucial convergence upon which drugs with eu-
phorigenic properties and/or abuse potential (regard-
less of chemical structure or pharmacological cat-
egory) act to enhance neural reward functions,
subjective experience of reward, and reward-related
behaviors.
neurons in nucleus accumbens. However, since GABA
appears to be colocalized with the opioid neuropep-
tide enkephalin in at least a portion of the medium
NEUROBIOLOGICAL TECHNIQUES
spiny projection pathway from nucleus accumbens to
FOR STUDYING BRAIN REWARD
pallidum and from nucleus accumbens to ventral
tegmental area (as well as apparently being colocalized
Among the many techniques that have been devel-
with the opioid neuropeptide dynorphin and the
oped for studying the neurobiological substrates of
nonopioid neuropeptide Substance P in at least a
brain reward, the following have proven useful and
portion of the medium spiny projection pathway from
will be dealt with below under the rubric of cannabi-
nucleus accumbens to ventral tegmental area), this
noid interactions with brain reward substrates: (a)
suggestion may well be fully congruent with the
electrical brain stimulation reward or ICSS, (b) single-
above-noted hypothesis that the third-stage reward
neuron electrophysiological recordings, (c) in vivo brain
neurons are coextensive with at least a portion of the
microdialysis, (d) in vivo brain voltammetric electro-
enkephalinergic pathway from nucleus accumbens to
chemistry, and (e) in vitro neurochemical assays and
ventral pallidum. A number of additional circuits
receptor binding measurements. As a detailed explica-
synapse onto the first-stage, second-stage, or third-
tion of these laboratory techniques is beyond the scope
stage elements of this brain reward system, apparently
of the present paper, the interested reader is referred to
to regulate and modulate the overall set-point of
recent reviews by the senior author (58,90).
506 Gardner and Vorel
TECHNIQUES FOR STUDYING
REWARD-RELATED BEHAVIORS
Many techniques have been developed for studying
reward-related behaviors in controlled laboratory set-
tings. Among those that have been used for studying
cannabinoids, and which will be dealt with below
under the rubric of cannabinoid effects on reward-
related behaviors, are: (a) conditioned taste preference/
aversion, (b) conditioned cue or place preference/
aversion, and (c) drug self-administration. The
interested reader is referred to reviews by the senior
author and others for detailed explications of these
laboratory techniques (58,91).
EFFECTS OF CANNABINOIDS
ON REWARD PROCESSES
As cannabinoids have been used for recreational
purposes by humans since ancient times (47), and as
cannabinoid receptors are found within brain loci and
circuits known to mediate reward functions (see be-
low), it may legitimately be inquired whether: (a)
cannabinoids act on brain reward mechanisms and on
behaviors mediated by such mechanisms and, if so, (b)
whether cannabinoids act on brain reward mecha-
nisms in a manner similar to that of other recreational
FIG. 2. Enhanced brain-stimulation reward following acute systemic administration of 9-THC (1.5 mg/kg, ip) and attenuation of
resulting
enhancement of brain-stimulation reward by acute naloxone. Enhanced brain reward is experimentally equivalent to a decrease in
electrical
brain-stimulation thresholds (see text) in the medial forebrain bundle. ‘‘PVP 20%’’ is the 20% polyvinylpyrrolidone vehicle used
for suspending
the 9-THC into an injectable form. The brain reward-enhancing effect is shown at both 15 and 30 min post- 9-THC. Probability
values shown
are for the specific comparisons indicated. (Reprinted, with permission of the publisher, from Refs. 93 and 94.)
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.

and abuse-prone drugs.Although there areinconsisten-


cies in the literature, the answers to such inquiries
appear to be affirmative.
INTERACTIONS OF CANNABINOIDS
WITH BRAIN REWARD SUBSTRATES
Cannabinoid Effects on Electrical Brain
Stimulation Reward
In the view of many, the electrical brain stimulation
reward paradigm offers the most direct in vivo neuro-
biological assay of drug effects on brain reward sub-
strates (58). In 1988, our research group demonstrated
that 9-THC enhances electrical brain-stimulation re-
ward (i.e., lowers brain reward thresholds) in the MFB
of laboratory rats (Fig. 2). For these experiments (92),
male rats were surgically implanted with chronic
brain-stimulation electrodes in the MFB and trained to
self-deliver rewarding electrical brain stimulation by a
titrating threshold stimulation procedure. This para-
digm allows the animal to indicate its threshold for
brain reward on a minute-to-minute basis throughout
the test session. The test chambers contain two re-
sponse levers. Each response by an animal on the
primary or ‘‘stimulation’’ lever delivered a 250-ms
stimulus train of 60-Hz bipolar rectangular pulse pairs

50 40 30 20
15 min POST 30 mín POST
Cannabinoids and Brain Reward 507
through the brain electrode. Initial current intensity for
separate pulse frequency, animals responded for two
each animal was set at the lowest intensity supporting
30-s time periods (‘‘bins’’), following which the pulse
consistent stable responding of 1000 lever presses per
frequency was decreased by 0.05 log units. Following
30-min test session. The current decremented by 1/16
each 30-s bin, the lever was retracted for 10 s. The two
of this initial intensity at every third press of the
30-s bins at each frequency were tested consecutively.
primary lever. At any point during the ensuing self-
Animals were trained on this rate-frequency proce-
administered decremental brain stimulation, the ani-
dure until reward thresholds were stable. Two com-
mal could reset the current back up to the initial
monly used measures of reward threshold were re-
maximum level by pressing the secondary or ‘‘reset’’
corded: M
50
, the pulse frequency at which the animals
lever (which did not itself deliver brain stimulation).
responded at half-maximum, and
0
, the frequency at
The current levels at which the animals reset were
which animals failed to respond for rewarding stimula-
recorded by microprocessors throughout each 30-min
tion. Both are reliable quantitative neurophysiological
test session, and the mean of the resulting frequency
indices of stimulation efficacy. Reward thresholds were
distribution of self-determined reset levels was opera-
considered stable when M
50
and
0
were within 0.01
tionally defined as the brain reward threshold. After
log units of their respective previous values for three
training to stable performance, each animal was tested
consecutive days. Animals were then injected with the
daily with intraperitoneal saline injections for a mini-
vehicle solution and tested in the brain reward proce-
mum of 6 weeks to ensure absolutely stable baselines
dure. The next day, animals were injected with 1.0
of brain reward threshold before drug trials began.
mg/kg 9-THC (ip) and tested in the brain reward
After this 6-week baseline period, each animal was
procedure. On each test day, testing began 20 min after
injected intraperitoneally on the next day with the 20%
vehicle or 9-THC injection. The findings are shown in
polyvinylpyrrolidone (PVP) vehicle and tested on the
brain reward paradigm. Two weeks of additional daily
baseline testing then ensued, and then, on the next day,
each animal was injected intraperitoneally with 1.5
mg/kg 9-THC and tested on the brain reward para-
Fig. 3. 9-THC significantly enhanced electrical brain-
stimulation reward (lowered reward thresholds), espe-
cially in Lewis strain rats, a strain long recognized as
sensitive to the reward-enhancing effects of addictive
drugs (97–99).
digm. Two more weeks of daily testing then ensued,
after which each animal was again given 9-THC or
vehicle and again tested on the brain reward para-
Cannabinoid Effects on in Vivo Extracellular
digm. In contrast to the total ineffectiveness of both
DA Overflow in Brain Reward Circuits
saline and the PVP vehicle, 9-THC significantly low-
ered brain reward thresholds in the MFB (92–94). More
As early as 1986, our research group reported that
recently, we have once again examined the effects of
9-THC enhances extracellular DAoverflowin reward-
9-THC on electrical brain-stimulation reward thresh-
relevant brain loci, as measured by in vivo brain
olds, this time using a rate-frequency curve-shift quan-
microdialysis (100). Subsequent work, both by our
titative electrophysiological brain reward threshold
group (93,94,101–104) and by others (105), has con-
paradigm (95,96). Animals were tested in an operant
firmed those original reports. The DA-enhancing effect
chamber equipped with a retractable lever on which
(Fig. 4) is tetrodotoxin-sensitive and calcium-depen-
the rat pressed to obtain rewarding constant current
dent (93,94,104) and is seen not only in the nucleus
electrical brain stimulation. Trains of 0.1-ms cathodal
accumbens (104) but also in other reward-relevant
pulses, of 250 ms duration, were delivered contingent
forebrain DA terminal projection loci, including me-
on a single lever press. A PC-AT microcomputer
dial prefrontal cortex (103) and neostriatum
monitored the rats’ responses, recorded data, and set
(100,101,105). Very recently, this enhancing effect of
appropriate current and pulse frequencies. Animals
cannabinoids on extracellular nucleus accumbens DA
were screened for electrical brain-stimulation behavior
has been once again confirmed by Tanda et al. (106).
using 250-ms trains of 150 Hz stimulation, with the
Additionally, however, those workers determined that
current set at 200 µA. Once animals acquired self-
the DA-enhancing effect occurs selectively within the
stimulation behavior, the current was adjusted to
shell of the accumbens (106), an important advance—
produce moderate rates of responding and the animals
given that the shell subportion of nucleus accumbens
were trained to lever press for a series of 16 different
appearsspecialized for mediating drug-enhanced brain
pulse frequencies, ranging from 25 to 141 Hz, pre-
reward functions (58,107–110). Also, Tanda et al. ex-
sented to the animal in descending order. At each
tended the cannabinoid-induced DA augmentation to
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
508 Gardner and Vorel
FIG. 3. Enhanced brain-stimulation reward following acute sys-
temic administration of 9-THC (1.0 mg/kg, ip). Enhanced brain
reward is experimentally equivalent to a left shift in the mean
rate-frequency electrical brain stimulation reward functions of the
medial forebrain bundle shown above (see text). In Fischer 344 rats,
9-THC did not significantly affect the brain stimulation reward
function curve. In Lewis rats, 9-THC significantly shifted the reward
function curve to the left (enhanced brain reward), as indicated by
significant leftward shifts in both M
50
and
0
points (see text). In
Sprague–Dawley rats, 9-THC enhanced brain stimulation reward
only in terms of a significant leftward shift in the
0
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
point (see text).
Although animals were tested at 16 different pulse frequencies from
25 to 141 Hz, only the central portion of each mean rate-frequency
brain stimulation reward function is depicted, as rewards/30 s were
asymptomatic for all animals below 50 Hz and above 100 Hz.
(Reprinted, with permission of the publisher, from Ref. 96.)
include the synthetic cannabinoid agonist WIN55212-2
and showed that the effects on DAwere blocked by the
selective CB1 antagonist SR141716A. In the present
authors’ laboratory, we have used additional means
for measuring extracellular DA in vivo—voltammetric

electrochemistry—in forebrain reward-related loci and


reported enhancement of extracellular DA by 9-THC
(Fig. 5) (101). Thus, despite one negative report (111),
the overwhelming evidence from several different
laboratoriesand using two different in vivo neurochemi-
cal techniques is that cannabinoids enhance extracellu-
lar DA in the reward-relevant forebrain.
Cannabinoid Effects on in Vitro DA Indices
in Brain Reward Circuits
An extensive literature exists on cannabinoid effects
on DA function in forebrain reward-related loci as
measured by in vitro biochemical techniques. Although
this literature spans more than 20 years and involves
many different laboratories, a considerable consistency
emerges from it—cannabinoid administration (at low
or moderate doses that appear relevant to human
cannabinoid use) enhances DA synthesis, release, and
turnover and inhibits DA reuptake in reward-relevant
brain loci (112–123). A great deal of the work has been
done by the Spanish research group of Rodrı´guez de
Fonseca, Navarro, Romero, Bonnin, Ferna´ndez-Ruiz,
Cebeira, de Miguel, Herna´ndez, Ramos, and their
colleagues (124–129). This group has reported that
9-THC enhances DA synthesis, release, and turnover
in reward-relevant brain loci. The effects on DA recep-
tor binding are more mixed, with both decreases in D
1
and D
2
receptor densities (without changes in receptor
affinities) (124–126) and increases in D
1
receptor density
(127) being reported. Interestingly and surprisingly,
this group finds that anandamide—the endogenous
cannabinoid—decreases DAsynthesis, release, and turn-
over (128,129), although these effects are in neostria-
tum rather than nucleus accumbens.
Cannabinoid Effects on Single-Neuron Electrical
Activity in Reward-Relevant Brain Loci
Given the above-cited evidence for cannabinoid
enhancement of DAfunctions in reward-relevant (espe-
cially mesolimbic) DA loci, the question arises as to the
underlying mechanism(s) of such enhancement. Some
compounds (e.g., amphetamines, cocaine) which en-
hance DA functions in reward-relevant DA loci do so
by enhancing DA release or inhibiting DA reuptake in
DA axon terminal areas such as nucleus accumbens
(58). Other compounds (e.g., opioids, nicotine) act by
stimulating the firing rate of reward-relevant DA
neurons that project from the ventral tegmental area to

Rewards/30 sec
Log Frequency (Hz) . I Ls* 1.9.
Fischer 344
O Vehicle I THC 1.0 mg/kg
I | 1 | I Lewis
I|1II
40 50 60 70 80 90
Frequency (Hz)
100
Cannabinoids and Brain Reward 509
FIG. 4. Enhanced extracellular DA overflow produced by 9-THC as measured by in vivo brain microdialysis in nucleus
accumbens
(brain-reward-relevant mesolimbic DA terminal region) and attenuation of resulting enhancement of extracellular DA overflow
by acute
naloxone. Each bar represents the peak amount of DA measured (shown as percentage change from predrug basal DA efflux)
following acute
systemic administration of 9-THC (0.5 or 1.0 mg/kg, ip) or vehicle. Probability values shown are for the specific comparisons
indicated.
(Reprinted, with permission of the publisher, from Ref. 94.)
the nucleus accumbens (58,130,131). Only two studies
appear to have addressed the question of whether
cannabinoids augment DA function in the mesolimbic
DAreward/reinforcement system by augmenting neu-
ronal firing within that system (132,133). Our research
group, using microelectrodes to record the firing pat-
terns of clearly identified single DA neurons in the
ventral tegmental area, could find no evidence for such
an effect (132). On the other hand, French (133) recently
reported finding that 9-THC does augment the firing
of single DAneurons in the ventral tegmental area and
that this effect is blocked by the selective CB1 antago-
nist SR141716A but not by the opioid antagonist
naloxone. Given the extremely similar methods used
in the two studies, the dissimilar findings are not
readily explicable. It is of particular note and interest
that French found cannabinoid augmentation of meso-
limbic DA single neuron firing to be antagonized by
SR141716A but not naloxone (133), because naloxone
antagonism of cannabinoid-induced DA enhancement
in forebrain reward-related loci has been a consistent
(albeit counterintuitive) finding (58,93,94,104,106,114—
see also discussion below).
Two other groups have used single-neuron record-
ing techniques to study cannabinoid effects on the
neostriatal outflow neurons that synapse into the re-
ward-relevant DAcell fields of the ventral mesencepha-
lon (134,135). These neurons are highly relevant for
study, as they appear to be extraordinarly enriched
with cannabinoid receptors (15,136–139). Both groups
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.

found that systemic injections or local microinjections


of cannabinoid agonists (WIN55212-2 and CP55940)
produced increased firing rates in substantia nigra
pars reticulata neurons. Tersigni and Rosenberg (135)
also found that local microinjections of the selective
CB1 antagonist SR141716A produced a decrease in the
spontaneous firing of those neurons, implying that they
are normally under tonic cannabinoid regulatory con-
trol. Miller and Walker (134) found that the cannabi-
noid agonist WIN55212-2 inhibited the activation of
substantia nigra pars reticulata neurons produced by
electrical stimulation of the neostriatum and that this
effect was reversed by the GABAantagonist bicuculline.
They posit that cannabinoid receptors on axon termi-
nals in the ventral mesencephalon produce neuronal
disinhibition by inhibiting GABA release into the ven-
tral mesencephalic cell fields. This is an interesting
hypothesis, because a similar mechanism could well
underlie the cannabinoid activation of reward-relevant
mesolimbic DA neurons reported by French (133).
Prenatal and Perinatal Cannabinoid Exposure
Effects on Brain Reward Substrates
The Spanish research group cited above has also
been extraordinarily active in studying prenatal and
perinatal cannabinoid exposure effects on brain re-
ward substrates (140–149). These studies of pre- and
perinatal cannabinoid exposure on developing DA

PERCENT CHANGE FROM BASAL DA


N RAT NUCLEUS ACCUMBENS
NAL
510 Gardner and Vorel
FIG. 5. Enhanced presynaptic DA efflux produced by 9-THC and
by the presynaptic DA reuptake blocker nomifensine, as measured
by in vivo voltammetric brain electrochemistry. (A) Effects of 9-THC
(0.5 mg/kg, ip) on K -evoked voltammetric electrochemical signals
corresponding to presynaptic DA efflux. The arrow indicates the
timing of the localized intracerebral micropressure K applications.
(B) Effects of nomifensine (5.0 mg/kg, ip) on K -evoked voltammet-
ric electrochemical signals corresponding to presynaptic DA efflux.
The arrow indicates the timing of the localized intracerebral micro-
pressure K applications. The time dynamics of the 9-THC-
enhanced presynaptic DA efflux signal are identical to those of the
nomifensine-enhanced presynaptic DA efflux signal, raising the
possibility that 9-THC may act on reward-relevant DA neurons by
inhibiting presynaptic DA reuptake. (Reprinted, with permission of
the publisher, from Ref. 101.)
neurons provide clear evidence that cannabinoids
produce marked effects on several biochemical and
molecular neurobiological indices of DA activity mea-
sured at various perinatal and adult ages. Strikingly,
these effects show considerable (indeed, in some cases,
extreme) ontogenetic, neuroanatomic, and sexually
dimorphic variation. Some of these effects are very
long-lasting, even into adulthood—presumably result-
ing in long-term or permanent alterations in the
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.

set-point or responsiveness of DA brain reward sub-


strates. Thus, this group has reported that prenatal
exposure to hashish extracts produces decreased DA
and 3,4-dihydroxyphenylacetic acid (DOPAC, a DA
metabolite) content in the prosencephalon of male rats
on the day before birth but no changes in females
(142,145). Combined prenatal and perinatal hashish
exposure (from gestational day 5 to postnatal day 24)
produces decreased DA content and tyrosine hydroxy-
lase (TH, the rate-limiting synthesizing enzyme for
neuronal DA) activity in the prosencephalon of male
rats on postnatal day 5 but no changes in females
(142,145), while the same exposure produces increased
TH activity in the prosencephalon at postnatal day 10
in males and increased DA content in females (142,145).
In the neostriatum, combined prenatal and perinatal
hashish exposure (again, from gestational day 5 to
postnatal day 24) produces increased numbers of DA
D1 and D2 receptors in males on postnatal day 15 but
no changes in females (140,141,145), increased numbers
of D2receptors(with decreasedreceptor affinity)coupled
with decreased TH activity in males on postnatal day 20
and decreased numbers of D1 receptors coupled with
increased DOPAC/DA ratios (suggesting increased pre-
synaptic DAneuronal activity) in females(140,141,145),
increased numbers of D1 receptors coupled with de-
creased TH activity in males on postnatal day 30 but no
changes in females (140,141,145), and increased num-
bers of D1 and D2 receptors (with decreased D2 receptor
affinity) coupled with decreased TH activity in males on
postnatal day 40 and decreased numbers of D1 receptors
in females (140,141,145). In the limbic forebrain, com-
bined prenatal and perinatal hashish exposure (gesta-
tional day 5 to postnatal day 24) produces no changes
in DA substrates in males on postnatal day 15 but
decreased numbers of D1 receptors together with de-
creased DA and DOPAC in females (140,141,145), no
changes in DA substrates in males on postnatal day 20
and increased presynaptic DA neuronal activity (in-
creased DOPAC and DOPAC/DA ratios) in females
(140,141,145), increased TH activity in males on postna-
tal day 30 with decreased DA in females on postnatal
day 30 (140,141,145), and increased presynaptic DA
neuronal activity (increased DOPAC and DOPAC/DA
ratios) in males on postnatal day 40 and no changes in
females (140,141,145). In all these studies, postnatal
days 15 and 20 were within the period of cannabinoid
exposure, while postnatal days 30 and 40 were after
termination of cannabinoid exposure. In adult animals
exposed to combined prenatal and perinatal hashish
(gestational day 5 to postnatal day 24), males have
increased numbers of neostriatal D2 receptors while

,uM CHANGE FROM BASELINE


Cannabinoids and Brain Reward 511
females have decreased numbers of D1 receptors to-
number, which are not seen in males. Taken all to-
gether with increased DOPAC/DA ratios, indicating
gether, this impressive body of work suggests that
increased neostriatal presynaptic DA neuronal activity
prenatal and perinatal cannabinoid exposure affects DA
and in vivo DA synthesis (143,144,147). At the molecu-
brain reward substrates during fetal development, during
lar neurobiological level, combined prenatal and peri-
early-to-mid postnatal life, and during adulthood—albeit
natal hashish exposure of male rats (gestational day 5
with significant ontogenetic, neuroanatomic, and sexually
to postnatal day 24) produces increased amounts of TH
dimorphic variations. Additional work from other labora-
mRNA and TH protein in the mesencephalon (but no
tories—including the pioneering studies of Walters and
differences in striatum) at postnatal days 15 and 20
Carr (150)—iscongruentwith thatofthe Spanish group.
(during the hashish exposure), no changes in TH mRNA
amountin the mesencephalon at postnatal days 30 and 40
(after cessation of hashish exposure), and a decrease in TH
mRNAin adult(postnatal day70) animals(148).
These same workers have also addressed the ques-
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.
Cannabinoid Withdrawal Effects on
Reward-Related Brain Mechanisms
tion of whether pre- and perinatal cannabinoid expo-
sure affects TH gene expression during fetal and early
neonatal periods (149). They found that exposure to
9-THC produces increased amounts of TH mRNA and
TH protein, together with increased TH activity, in the
fetal rat brain at gestational day 14 (149). However, a
marked sexual dimorphism in the response of the TH
gene to 9-THC exposure appeared at later gestational
periods (gestational days 18 and 21), when TH mRNA
amounts increased in the developing female brains but
decreased in the developing male brains. These data are
summarized in Table 1. Clearly, there are ontogenetic,
As noted above, administration of drugs with recre-
ational properties and abuse potential enhances brain-
stimulation reward and the mesotelencephalic DA
neurochemical substrates underlying such reward. Con-
versely, withdrawal from such dosing regimens produces
depletion of DA in brain reward loci. For example, with-
drawal from cocaine administration produces nucleus
accumbens DA depletion (151–153), as does opiate with-
drawal(154–158).Withdrawalalsoproducesclearreward-
related functional sequelae. For example, withdrawal from
cocaine or amphetamine produces elevations in brain-
stimulation reward thresholds (159–166), as does with-
neuroanatomic, and sexually dimorphic variations in
drawal from opiates (167,168). Also, opiate withdrawal
cannabinoid effects on in vitro brain DA indices. How-
produces conditioned cue aversion (168–172). Signifi-
ever, to the present authors’ eyes, the following consis-
cantly, the neural mechanisms underlying this with-
tencies emerge: (a) prenatal cannabinoid exposure
drawal-produced presumptive negative hedonic tone
appears to enhance TH mRNA, TH protein, and TH
or dysphoria may involve the nucleus accumbens
activity in fetuses at gestational day 14, but this
(169,170), just as the drug-induced positive hedonic
disappears by gestational day 16; (b) prenatal cannabi-
tone may (see above). Congruent with these findings
noid exposure appears to inhibit DA functions (TH
from a variety of paradigms, DA depletion in the nucleus
mRNA, DA, DOPAC) in male fetuses at gestational
accumbens and elevations in brain reward thresholds
day 21, but this is not seen in females; (c) a similar
havebeenproposedasneuralsubstratesforpost-drug-use
inhibition of TH mRNA appears in male fetuses at
anhedonia and drug craving (169,173,174). As noted by
postnatal day 1, which may be a continuation of the
Wise and Munn (165), ‘‘dopamine depletion and . . .
effects seen at gestational day 21; (d) in males assayed
attendant subsensitivity of the reward system offers a
at postnatal days 20, 30, and 40, there appear to be
withdrawal symptom that may be more significant for
relatively consistent decreases in TH activity and D2
drug self-administration than classic [physical with-
DA receptor affinity and increases in D1 and D2 DA
drawal] . . . symptoms’’ and ‘‘subsensitivity of the
receptor numbersin the neostriatum, butthese changes
reward system . . . is more obviously linked to the
are not observed in female animals; (e) in males
habit-forming property of drugs rather than to corre-
assayed at postnatal days 15 and 20, enhanced TH
lated side effects.’’ Importantly, since elevations in
mRNA and TH protein levels are seen in the mesen-
brain reward thresholds and correlated DA depletion
cephalon; (f) in females assayed at postnatal days 1, 5,
in forebrain reward loci, unlike other physical with-
and 10, there appear to be some reasonably consistent
drawal symptoms, offer a set of withdrawal symptoms
increases in DAand DOPAC levels in the prosencepha-
common to psychostimulants, opiates, and ethanol,
lon which are not seen in males; and (g) in females
they may constitute the long-sought common denomi-
assayed at postnatal days 20–70, there appear to be
nator for addiction (165; see also discussions of the
reasonably consistent decreases in D1 DA receptor
search for a common neurobiological denominator for