Trajectory of Adolescent Cannabis Use on Addiction Vulnerability.
ABSTRACT The adolescent brain is a period of dynamic development making it vulnerable to environmental factors such as drug exposure. Of the illicit drugs, cannabis is most used by teenagers since it is perceived by many to be of little harm. This perception has led to a growing number of states approving its legalization and increased accessibility. Most of the debates and ensuing policies regarding cannabis were done without consideration of its impact on one of the most vulnerable population, namely teens, or without consideration of scientific data. We provide an overview of the endocannabinoid system in relation to adolescent cannabis exposure and provide insights regarding factors such as genetics and behavioral traits that confer risk for subsequent addiction. While it is clear that more systematic scientific studies are needed to understand the long-term impact of adolescent cannabis exposure on brain and behavior, the current evidence suggests that it has a far-reaching influence on adult addictive behaviors particularly for certain subsets of vulnerable individuals.
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ABSTRACT: The racial/ethnic composition of the US population is shifting, with the nonwhite population growing faster than whites. We examined cannabis use disorder (CUD) prevalences and correlates in seven racial/ethnic groups. We included cannabis use (CU) prevalence as a comparison. Data were from the 2005–2011 National Surveys on Drug Use and Health (N = 394,400). Substance use among respondents aged ≥12 years was assessed by computer-assisted, self-interviewing methods. The following were included as control variables: age, sex, family income, government assistance, county type, residential stability, major depressive episode history, arrest history, nicotine dependence, alcohol disorder, and survey year. Past-year CU prevalence increased significantly from 10.45% in 2005 to 11.41–11.54% during 2009–2011. Compared with whites, mixed-race individuals had higher odds of CU; Asian Americans and Hispanics had lower odds of CU. There were no significant yearly changes in CUD prevalence in the sample during 2005–2011 (1.58–1.73%). Compared with whites, individuals who were mixed-race, black, and Native American had higher odds of CUD; Asian Americans had lower odds. In aggregate, 15.35% of past-year cannabis users met criteria for a CUD in the 12-month period. Past-year cannabis users who were black, Native American, Hispanic, or Asian American had higher odds of CUD than white users. In each racial/ethnic group, adolescent cannabis users generally showed greater odds of CUD than adult users. Behavioral health indicators (major depressive episode, arrest history, nicotine dependence, alcohol disorder) were associated with CU and CUD. In conclusion, CUD disproportionally affects nonwhite groups and youth.Journal of Psychiatric Research 01/2014; 50:26–35. · 4.09 Impact Factor
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ABSTRACT: The dose-dependent toxicity of the main psychoactive component of cannabis in brain regions rich with cannabinoid CB1 receptors is well known in animal studies. However, research in humans does not show common findings across studies regarding the brain regions that are affected after long-term exposure to cannabis. In the present study we investigate (using Voxel-based Morphometry) grey matter changes in a group of regular cannabis smokers in comparison to a group of occasional smokers matched by the years of cannabis use. We provide evidence that regular cannabis use is associated with grey matter volume reduction in the medial temporal cortex, temporal pole, parahippocampal gyrus, insula, and orbitofrontal cortex; these regions are rich with cannabinoid CB1 receptors and functionally associated with motivational, emotional, and affective processing. Furthermore, these changes correlate with the frequency of cannabis use in the three months prior to inclusion in the study. The age of onset of drug use also influences the magnitude of these changes. Significant grey matter volume reduction could result either from heavy consumption unrelated to the age of onset or instead from recreational cannabis use initiated at an adolescent age. In contrast, the larger grey matter volume detected in the cerebellum of regular smokers without any correlation with the monthly consumption of cannabis may be related to developmental (ontogenic) processes that occur in adolescence.Neuropsychopharmacology accepted article preview online, 17 March 2014; doi:10.1038/npp.2014.67.Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology 03/2014; · 8.68 Impact Factor
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ABSTRACT: Withdrawal, a diagnostic indicator of cannabis use disorder, is often minimized or ignored as a consequence of cannabis use, particularly among adolescents. This study aims to characterize cannabis withdrawal among adolescents in outpatient treatment for substance use disorder and evaluate the clinical significance of withdrawal as a predictor of substance-related outcomes.Journal of Addiction Medicine 08/2014; · 1.73 Impact Factor
Trajectory of adolescent cannabis use on addiction vulnerability
Yasmin L. Hurda,b,c,*, Michael Michaelidesa,b, Michael L. Millera,b, Didier Jutras-Aswadd,e
aDepartment of Psychiatry, Icahn School of Medicine at Mount Sinai, New York, NY, USA
bDepartment of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA
cJames J. Peters Veterans Administration, Bronx, NY, USA
dDepartment of Psychiatry, Université de Montréal, Montreal, QC, Canada
eCRCHUM, Centre hospitalier de l’Université de Montréal, Montreal, QC, Canada
a r t i c l e i n f o
Received 11 March 2013
Received in revised form
16 July 2013
Accepted 19 July 2013
a b s t r a c t
The adolescent brain is a period of dynamic development making it vulnerable to environmental factors
such as drug exposure. Of the illicit drugs, cannabis is most used by teenagers since it is perceived by
many to be of little harm. This perception has led to a growing number of states approving its legalization
and increased accessibility. Most of the debates and ensuing policies regarding cannabis were done
without consideration of its impact on one of the most vulnerable population, namely teens, or without
consideration of scientific data. We provide an overview of the endocannabinoid system in relation to
adolescent cannabis exposure and provide insights regarding factors such as genetics and behavioral
traits that confer risk for subsequent addiction. While it is clear that more systematic scientific studies
are needed to understand the long-term impact of adolescent cannabis exposure on brain and behavior,
the current evidence suggests that it has a far-reaching influence on adult addictive behaviors particu-
larly for certain subsets of vulnerable individuals.
This article is part of a Special Issue entitled ‘NIDA 40th Anniversary Issue’.
? 2013 Elsevier Ltd. All rights reserved.
Adolescence is an important stage of behavioral maturation and
brain development during which the high degree of neuroplasticity
that occurs in this ontogenetic period places the adolescent brain at
particular risk to environmental factors such as drug exposure.
Marijuana (Cannabis sativa) continues to be the illicit drug most
commonly used by teenagersin the United States as well as in other
Western societies (Johnston et al., 2012; SAMHSA, 2011). Although
cannabis is not as highly addictive as other substances, such as
heroin and cocaine, cannabis-dependent individuals still greatly
outnumber those reporting dependence on other illicit drugs and
the number of people seeking treatment for cannabis dependence
continues to increase yearly (SAMHSA, 2011).
Despite these facts, there is a growing perception, particularly in
adolescents and young adults (Kilmer et al., 2007; Lopez-Quintero
and Neumark, 2010), that cannabis is ‘harmless’ especially when
compared to other abused substances like nicotine (tobacco) and
alcohol that are legal. Reasons cited for this perception include the
consideration that cannabis-associated mortality is lower than to-
bacco and alcohol, which are associated with cancer and overdose/
vehicular accidents, respectively. In addition, cannabinoids provide
medicinal benefits (Hermanson and Marnett, 2011; Hill et al., 2012)
in contrast to tobacco and alcohol, which have no medical in-
dications. These and other considerations have contributed to the
decriminalization, or even legalization, of cannabis in a number of
states within the USA. Economic factors have also been suggested
as a rationalization for legalization as a potential source of tax
revenue for state governments. Despite some cogent arguments in
the current debates regarding legalization and increased avail-
ability of cannabis, most of the discussion and policies have been
made without significant consideration of scientific data.
Growing evidence suggests a differential effect of cannabis
exposure on the human brain based on the age of exposure, but the
question remains as to the potential long-term mental health
consequences of cannabis exposure in teens. Few scientific studies
have systematically investigated the long-term impact of cannabis
use in relation to the developing teenage brain, the population
most crucial to the current debates. Nevertheless, the available data
to date, as discussed in this review, suggest that adolescent
cannabis exposure induces significant protracted effects suggestive
of enhanced vulnerability to addiction and psychiatric disorders in
later life, at least in certain subsets of individuals.
* Corresponding author. Icahn School of Medicine at Mount Sinai, Departments of
Psychiatry, Neuroscience and Pharmacology and Systems Therapeutics, New York,
NY, USA. Tel.: þ1 212 824 9314; fax: þ1 646 537 9598.
E-mail address: Yasmin.email@example.com (Y.L. Hurd).
Contents lists available at ScienceDirect
journal homepage: www.elsevier.com/locate/neuropharm
0028-3908/$ e see front matter ? 2013 Elsevier Ltd. All rights reserved.
Neuropharmacology 76 (2014) 416e424
2. Neurobiology of the endocannabinoid system
The main psychoactive component of cannabis, D9-tetrahydro-
cannabinol (THC), acts primarily via cannabinoid receptors (CBRs)
d CB1R and CB2R (Gerard et al., 1991; Griffin et al., 2000; Matsuda
et al., 1990; Munro et al., 1993). The CB1R is one of the most
abundant G-protein-coupled receptor in the brain (Herkenham
et al., 1990, 1991a) and is Gi/o-coupled, suppressing neurotrans-
mitter release (Howlett et al., 2002). The expression of CB1R is most
pronounced within the basal ganglia, cerebellum, cerebral cortex,
hippocampus and amygdala (Biegon and Kerman, 2001; Glass et al.,
1997; Herkenham et al., 1990, 1991b; Mailleux et al., 1992; Pettit
et al., 1998; Wang et al., 2003) (Fig. 1), consistent with cannabis
exerting significant effects on motor function, cognition, and
emotional regulation. Recent evidence, though initially controver-
sial, suggests that CB2R is also expressed within the central nervous
system in immune cells as well as glia and potentially neurons
(Gong et al., 2006; Lanciego et al., 2011; Onaivi et al., 2006; Van
Sickle et al., 2005). Nevertheless, the broad and abundant expres-
sion of CB1R in neuronal circuits relevant to addiction and psychi-
atric disorders still place a prominent emphasis on cannabis’
modulation of this CBR subtype in relation to psychiatric
Imaging studies of rodents (Verdurand et al., 2011) and human
subjects (Mato et al., 2003) suggest global increases in CB1R
throughout early life into adolescence, at which period adult levels
are generally maintained (Belue et al., 1995; McLaughlin et al.,
1994; Rodriguez de Fonseca et al., 1993), but there are also re-
ports of reduced CB1R expression from juvenile to adulthood that
mirrors developmental changes in CB1R-mediated signaling (Heng
et al., 2011). Some of the inconsistencies regarding the ontogenic
pattern of the CB1R may be due to regional, as opposed to global,
developmental differences in the receptor development in addition
to differences in mRNA, receptor protein or receptor binding being
Fig. 1. Cannabinoid receptor mRNA (CNR1) expression in the human brain emphasizes
this gene’s abundant expression in cerebral cortex e such as insular cortex (I) and
prefrontal cortex (PFC) e as well as the caudate nucleus (CN), putamen (Pu), nucleus
accumbens (NAc), hippocampus (Hipp), amygdala (Amy), and cerebellum (CB). Absent-
to-low mRNA expression is notable in the thalamus (T), basal forebrain (BF), globus
pallidus (GP), and midbrain (Ms).
Fig. 2. Schematic illustration of the striatonigral ‘Go’ andstriatopallidal ‘NoGo’ path-
ways. These medium spiny output neurons are distinguishable based on their targets
and subcellular markers, namely the expression of D1R (purple) and D2R (brown),
respectively. Both cell-types, however, express CB1R (orange). This dissociation is based
mainly on the dorsal striatal circuit, but a similar organization, particularly with
respect to the ‘NoGo’ pathway, exists for the ventral striatal circuit.
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
studied. Most preclinical investigations to date that have examined
the neurodevelopment of CB1R have focused on the striatum and
prefrontal cortical brain regions which are key components of
neuronal circuits implicated in addiction and related psychiatric
disorders. Reward, motivated behavior, decision-making, habit
formation and motor function are mediated by the prefrontal cor-
tex as well as components of the dorsal (associative- and
sensorimotor-related) and ventral (limbic-related) striatal areas
making these brains regions relevant to adolescent cannabis
With respect to cortical development, remodeling of excitatory
connections in the prefrontal cortex is a key feature of adolescent
neurodevelopment. For instance, there is significant pruning of
excitatory synapses that parallels the delayed maturity of cognitive
behaviors such as inhibitory control and working memory. The
CB1R is highlyabundant in the prefrontal cortex, primarily localized
on large cholecystokinin interneurons (Marsicano and Lutz, 1999),
but molecular and functional evidence suggest that CB1R is also
present in a subset of pyramidal neurons (Hill et al., 2007;
Marsicano and Lutz, 1999; Matsuda et al., 1993). Interestingly, the
most pronounced and progressive cortical alteration observed on
CB1R expression and CB1R-mediated functional signaling evident
during adolescent development in rodents is within the medial
prefrontal and other limbic/associative cortices as compared to
sensorimotor cortices (Heng et al., 2011). Whether such develop-
mental fluctuations of the CB1R directly relate to plasticity and the
synaptic remodeling that occurs in the prefrontal cortex during
adolescence is unknown.
In the striatum, CB1R are localized in the medium spiny
GABAergic neurons that constitute the major output pathways d
striatonigral and striatopallidal (Fig. 2). These receptors are
expressed on both striatonigral ‘direct’ and striatopallidal ‘indirect’
pathways which mediate ‘Go’ facilitatory (positive reward/choice)
and ‘NoGo’ (avoidance learning/inhibitory control) behaviors,
respectively, relevant to motor function and decision-making pro-
cesses (Durieux et al., 2009; Frank et al., 2007; Klein et al., 2007;
Sano et al., 2003). While no study thus far has characterized the
pattern and abundance of CB1R in the different output pathways
during development, it is known that CB1R expression is dynamic
during the course of adolescent development in different brain
regions. For example, in vivo (Verdurand et al., 2011) and in vitro
(Belue et al., 1995) imaging of the rat brain that revealed global
enhanced CB1R in the cortex, also showed increased CB1R in other
brain structures including the striatum during the transition from
early adolescence to adulthood. However, other investigators have
provided significant evidence for reduced CB1R expression and
mRNA levels from juvenile to adulthood (Van Waes et al., 2012).
Moreover, examination of CB1R protein expression restricted to the
adolescent developmental window suggests significant CB1R dif-
ferences even within distinct compartments of the nucleus
accumbens (Ellgren et al., 2008). During adolescence, CB1R protein
decreases in the nucleus accumbens shell yet concomitantly in-
creases in the core compartment. This suggests that distinct time
periods during adolescence may have different sensitivity to
cannabis exposure relevant to mesolimbic striatal function.
Given anatomical and functional relationships between the
prefrontal cortex and the striatum, it is not surprising that these
regions are coordinated in regard to development of the endo-
cannabinoid (eCB) system. There appears to be a direct correlation
between the developmental trajectory of CB1R expression in spe-
cific cortical regions with their projection to distinct striatal sub-
regions. For example, striatal subregions with high levels of CB1R
expression (dorsolateral sensorimotor regions) receive input pri-
marily from cortical areas with relatively low CB1R levels (motor,
somatosensory) (Van Waes et al., 2012). In contrast, striatal
subregions with low mRNA expression of Cnr1, such as the asso-
ciative ventromedial dorsal striatum and limbic nucleus accum-
bens, receive afferents from cortical areas with greater CB1R
expression (cingulate, insular). These findings emphasize a strong
inverse relationship between cortical and striatal CB1R expression
in frontostriatal circuits, suggesting a functional orchestration
vulnerable to adolescent cannabis exposure.
In addition to CBRs, other components of the eCB system, such
arachidonoylglycerol (2-AG), have significant developmental im-
plications. Indeed, eCB signaling plays a key role in hardwiring of
the brain during prenatal ontogeny, regulating synaptogenesis and
target selection for the development of neuronal pathways
(Harkany et al., 2008; Mulder et al., 2008). During later develop-
mental stages, the eCBs are well-documented regulators of synaptic
plasticity (Katona and Freund, 2008). Within adolescent ontogeny,
anandamide and 2-AG are dynamically altered in the striatum and
prefrontal cortex, as for instance, 2-AG is reduced from early to late
adolescence in these regions (Ellgren et al., 2008). There is also a
continuous increase of anandamide in the prefrontal cortex over
the course of adolescence. The fact that the eCB system is dynam-
ically altered during adolescence in brain areas central to reward,
decision-making and motivation suggests that cannabis exposure
during this critical developmental phase may have long-term in-
fluence on behaviors linked to the mesocorticolimbic system.
Clearly, however, the limited studies to date are incomplete, so
there still remains a large gap of knowledge regarding the adoles-
cent ontogeny of the eCB system.
3. Cannabis and ‘gateway’ effects
A major aspect of the debate regarding adolescent cannabis use
is whether it increases the use of other addictive substances such as
heroin and cocaine later in life, a phenomenon known as the
gateway hypothesis. Clinical and epidemiological studies have
documented a significant link between repeated early cannabis
exposure and an increased risk of other illicit drug use (Agrawal
et al., 2004; Brook et al., 1999a; Fergusson and Boden, 2008;
Fergusson and Horwood, 2000; Hall and Lynskey, 2005; Kandel,
1975; Yamaguchi and Kandel, 1984). Altogether, the data suggest
that use of ‘heavy’ drugs is almost systematically preceded by
cannabis use, and that risk is correlated with the intensity of
cannabis exposure. Moreover, cannabis use also appears more
deleterious when its onset occurs in younger versus older adoles-
cents in regard to adjustment in transition from adolescence to
young adulthood, education attainment, employment, delinquency
and ability to conform to adult role (Brook et al., 1999a, 1999b;
Fergusson and Boden, 2008; Fergusson et al., 2002; Lynskey et al.,
2003; Tucker et al., 2006). Despite these findings, a major caveat
of human studies is the difficulty of demonstrating a causal rela-
tionship between adolescent cannabis use and subsequent behav-
ioral disturbances, especially when considering the influence of
genetic and environmental factors alongside other aspects such as
polysubstance use (Cleveland and Wiebe, 2008; Fergusson et al.,
2006; Kandel et al., 2006; Lessem et al., 2006; Maccoun, 2006;
Tarter et al., 2006). Given these complexities, animal models are a
valuable tool to obtain direct insights about the relationship be-
tween early cannabis exposure and behavioral disruptions.
Many rodent investigations exploring the potential gateway
effects of cannabis have primarily studied synthetic cannabinoid
agonists that differ in pharmacological properties to THC. Never-
theless, studies examining adolescent exposure to cannabinoid
agonists or THC provide evidence of enhanced intake and sensi-
tivity later in life to opiate drugs (Biscaia et al., 2008; Ellgren et al.,
2007; Tomasiewicz et al., 2012). In our experimental rat model that
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
mimics the more periodic use of most adolescent cannabis users,
adult male rats with low-to-moderate THC exposure during
adolescence exhibit enhanced heroin self-administration behavior
(Ellgren et al., 2007). Cannabinoid-opioid interactions have also
been documented by studies showing that developmental expo-
sure to cannabinoid agonists increases heroin-induced conditioned
place preference (Biscaia et al., 2008; Singh et al., 2006). Short
adolescentexposureto cannabinoid agonist,WIN 55,212-2, has also
been reported to instead induce tolerance to morphine (Pistis et al.,
Animal models make it possible to identify neuroadaptations
that may contribute to the behavioral vulnerability related to
adolescent cannabis. Intriguingly, many experimental animal
studies to date have implicated the striatopallidal circuit in asso-
ciation with developmental cannabinoid exposure (Corchero et al.,
1998, 1999; Ellgren et al., 2007; Morel et al., 2009; Perez-Rosado
et al., 2000; Spano et al., 2007; Valverde et al., 2001). This theory
is based on consistent alterations of striatal dopamine D2 receptors
(Drd2) and proenkephalin (Penk) mRNA expression both of which
are preferentially co-expressed within striatopallidal medium
spiny neurons (Gerfen et al., 1990; Gerfen and Young III, 1988; Le
Moine et al., 1990). In our model of adolescent THC exposure,
reduced Drd2 and Penk (Ellgren et al., 2007; Tomasiewicz et al.,
2012) mRNA levels were observed within the nucleus accumbens
of adult animals (Fig. 3). Reduced D2R, the protein encoded by Drd2,
has long been a characteristic neurobiological feature of addiction
vulnerability. In vivo positron emission tomography (PET) evidence
has consistently demonstrated that subjects with substance abuse
have less available D2R in the striatum (Heinz et al., 2004; Volkow
et al., 2001, 2004, 1999; Wang et al., 1997), findings that animal
models have shown to be linked to enhanced drug self-
administration vulnerability (Morgan et al., 2002; Nader et al.,
2006). Over-expression of Drd2 in the ventral striatum attenuates
cocaine intake (Thanos et al., 2008), and D2R binding in this region
in cocaine-naïve rats negatively predicts future cocaine-seeking
behavior (Michaelides et al., 2012). In addition to adolescent THC
exposure, prenatal THC also leads to dysregulation of the Drd2 gene
in adulthood (DiNieri et al., 2011). That developmental THC expo-
sure reduces Drd2 mRNA expression in the striatum, and affects
related behavioral traits, support the hypothesis that develop-
mental cannabis may induce a neurobiological state of addiction
The finding of impaired Penk gene expression in cannabis-
exposed subjects is perhaps not surprising given the tight neuro-
biological interactions between the opioid and eCB systems. Opioid
neuropeptide receptors and CB1R are coexpressed on similar
neurons in the striatum, share similar G-protein coupled signaling
mechanisms and appear functionally interdependent (Blume et al.,
2013; Canals and Milligan, 2008). Of the opioid neuropeptides,
enkephalin, encoded by the Penk gene, directly regulates hedonic
states (Kelley et al., 2002; Skoubis et al., 2005). We recently docu-
mented a direct causal link between regulation of ventral striatal
Penk mRNA expression and heroin self-administration behavior.
Overexpression of the Penk gene in the nucleus accumbens shell by
useof viral-mediated manipulation
administration and heroin-seeking behavior in animals naïve to
THC, whereas in contrast, knocking down the Penk gene in THC-
exposed rats reduced heroin intake behavior (Tomasiewicz et al.,
2012). Altogether, these and other findings (Spano et al., 2010;
Vigano et al., 2005) suggest the tight interaction between canna-
binoids and the opioid system could contribute to the development
of opiate abuse in adults with previous exposure to THC during
In contrast to the effects noted for opiates, the impact of early
cannabinoid exposure on the subsequent sensitivity to stimulant
drugs have yielded inconsistent findings. While some studies failed
to find significant behavioral differences in response to amphet-
amine later in life (Ellgren et al., 2004), others report that adoles-
cent cannabinoids enhance cocaine-induced motor behavior (Dow-
Edwards and Izenwasser,2012).
(Higuera-Matas et al., 2008) and deceased self-administration of
cocaine (Panlilio et al., 2007) have been reported in adult animals
with adolescent cannabinoid exposure. Differences in experimental
factors, such as the duration and frequency of exposure, dose and
formulation of the cannabinoid, and even gender likely contribute
to these inconsistencies. For example, adolescent administration of
the CB1R agonist CP 55,940 was reported to increase cocaine self-
administration primarily in females, not males (Higuera-Matas
et al., 2008). These findings emphasize the need to systematically
probe factors such as the magnitude and duration of cannabinoid
exposure, adolescent period of exposure and gender in order to
help expand insights about individual risk factors contributing to
the gateway effects of adolescent cannabis.
4. Genetic and behavioral traits contribute to individual
Although animal studies demonstrate protracted behavioral and
neurobiological effects of adolescent THC exposure into adulthood,
there remains the fact that not all teenage cannabis users develop
future addictions or psychiatric disorders. In fact, despite its com-
mon use, only a subset of teens (w25%) and young adults (w19%)
using cannabis progress to abuse or dependence (SAMHSA, 2011).
Indeed, for most teenagers, cannabis is a terminus with no further
use of that or other illicit drugs as they mature into full adulthood,
suggesting that there are differences in individual vulnerability.
Humans vary tremendously for instance in regards to environment,
behavioral traits, genetics, and cultural norms. While these and
other factors play significant roles in complex disorders as addic-
tion, understanding the contribution of each factor is as much a
challenge as determining their interactions to risk.
4.1. Behavioral traits and personality
Cannabis users are generally characterized by apathy, loss of
goal-motivated behavior and negative mood states, and dependent
subjects report more negative affect, neuroticism, aggressivity and
impulsivity (Dorard et al., 2008; Hyman and Sinha, 2009; Jutras-
Aswad et al., 2012; Zvolensky et al., 2007). Negative affect in
cannabis-dependent subjects is also related to the severity of
dependence insomuch that this personality trait correlates with
Fig. 3. Periodic low-to-moderate THC exposure during adolescence (1.5 mg/kg every
third day between postnatal days 28 and 49) alters Drd2 and Penk mRNA expression in
the adult nucleus accumbens. These genes are strongly enriched in striatopallidal
neurons of the nucleus accumbens (*p < 0.05).
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
years of cannabis use, implying that long-term use of cannabis also
worsens negative affect. Such findings are consistent with the hy-
pothesis that in attempts to ‘self-medicate’, a user’s continued
consumption of cannabis itself exacerbates underlying negative
traits (Arendt et al., 2007). Rat studies also substantiate these hu-
man findings by documenting that exposure to CB1R agonists
during adolescence induces long-term increases in anxiety- and
Ciccocioppo et al., 2002). Indeed, a growing body of evidence
suggests that cannabis exposure in humans during adolescence is
linked to the development of symptoms characteristic of mood and
anxiety disorders (Fergusson et al., 2002; Hayatbakhsh et al., 2007;
Patton et al., 2002).
While the use of cannabis itself appears to lead to negative
affect, which could contribute to subsequent drug abuse as in-
dividuals try to self-medicate, even young cannabis dependent
subjects without a long history of drug use show high neuroticism/
anxiety and depression traits, implying a preexisting negative
emotional affect in these users (Dorard et al., 2008). As such, there
may be subsets of individuals with at-risk behavioral traits that
contribute to self-medication ultimately leading to dependence.
Self-medication due to a preexisting vulnerable state is also evident
in regard to psychosis risk. Cannabis use is high among people with
psychosis (Koskinen et al., 2010; van Gastel et al., 2013) and it has
been documented that psychotic symptoms are evident in subjects
who have never used cannabis before the onset of psychotic
symptoms, which also predicts future cannabis use (Ferdinand
et al., 2005). This suggests that current cannabis-dependent sub-
jects may have underlying psychiatric disorders that contributed to
self-medication and that through repeated use, led to dependence.
Thus, while cannabis may itself increase drug addiction and psy-
chiatric vulnerability, pre-existing prodromal states (or disease
vulnerability) may initially promote the initiation and continuation
of cannabis use.
Fig. 4. PENK SNPs (rs2609997 and rs2576573) are associated with PENK expression in the human brain. High-risk alleles for cannabis dependence of the rs2609997 and rs2576573
PENK SNPs associate with elevated mRNA expression and met-enkephalin peptide (met-enk) levels in the human striatum (A). Similarly, these alleles associate with elevated mRNA
expression in central amygdala nucleus (B). (High-risk genotypes ¼ C/C þ C/T for rs2609997 or A/A þ A/G for rs2576573; low-risk genotypes ¼ T/T for rs2609997 or G/G for
rs2576573. *p < 0.05.)
Fig. 5. Synergistic Contribution of negative affect trait and PENK variants to cannabis
dependence vulnerability. (High-risk genotype ¼ C/C þ C/T for rs2609997 or A/A þ A/G
for rs2576573; low-risk genotype ¼ T/T for rs2609997 or G/G for rs2576573.
***p < 0.01; ****p < 0.001. Modified from Jutras-Aswad et al., 2012.)
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
4.2. Heritable genetic factors
A growing number of family, twin and adoption studies have
shown that cannabis use disorder is strongly heritable (30e80%)
(Agrawal and Lynskey, 2009; Kendler et al., 2000; Kendler and
Prescott, 1998; Maes et al., 1999; McGue et al., 2000; Miles et al.,
2001; Rhee et al., 2003; Tsuang et al., 1998; van den Bree et al.,
1998). Most studies examining cannabis dependence and genetic
risk have used a candidate gene approach. Thus far, genes encoding
CB1R (CNR1) and the fatty acid amide hydrolase (FAAH), an enzyme
responsible for the hydrolysis of the eCB anadamide, have been
shown to modulate cannabis dependence risk (Agrawal et al., 2009;
Hopfer et al., 2006; Tyndale et al., 2007).
Given the effects of adolescent THC on striatopallidal-related
genes, namely Drd2 and Penk expression, in the rodent models,
(Corchero et al., 1998, 1999; Ellgren et al., 2007; Morel et al., 2009;
Perez-Rosado et al., 2000; Spano et al., 2007; Valverde et al., 2001;
Wanget al., 2006), it was of interest toexploreDRD2 andPENK SNPs
in humans in relation to cannabis dependence. The DRD2 Taq1A
polymorphism, which has been studied in multiple addiction dis-
orders, did not associate with cannabis use (Creemers et al., 2011;
Jutras-Aswad et al., 2012; Sakai et al., 2007). However, consid-
ering the link between DRD2 and inhibitory control, we probed
their interacting association with cannabis dependence risk. A
number of neurocognitive studies have documented that DRD2
SNPs predict avoidance-based decisions in healthy subjects, in line
with this gene’s association with the striatopallidal NoGo pathway
(Frank and Hutchison, 2009; Frank et al., 2007; Klein et al., 2007).
Our results confirmed that negative reinforcement learning, linked
with the ability to avoid maladaptative choices in a probabilistic
learning task, was indeed associated with the DRD2 rs6277 SNP
(Jutras-Aswad et al., 2012). For this DRD2 SNP, however, both
cannabis users and controls exhibited the same association with
negative reinforcement. Interestingly, negative affect (high anxiety/
neuroticism trait) which was prominent in cannabis users signifi-
cantly modulated the association between DRD2 and PENK geno-
types with cannabis dependence. This interaction was most
apparent for PENK SNPs (rs2609997 and rs2576573), with neurot-
icism/anxiety trait explaining approximately 15%e20% of the as-
sociation between genotype and cannabis dependence (Jutras-
Aswad et al., 2012).
Recent evidence from animal models have demonstrated a
direct role of the nucleus accumbens Penk striatopallidal pathway
in mediating behavioral responses associated with aversive
behavior (Hikida et al., 2010). In the human brain, there is a sig-
nificant association of PENK SNPs (rs2609997 and rs2576573) with
mRNA expression of this gene in the nucleus accumbens and dorsal
striatum as well as with striatal met-enkephalin peptide levels
(Jutras-Aswad et al., 2012). These findings emphasize the tran-
scriptional and translational functional relationships of poly-
morphisms of the PENK gene (Fig. 4A). In addition to the striatum,
PENK mRNA amygdala expression is also related to PENK genotype
which is interesting given that the amygdala plays a prominent role
in negative mood states and enkephalinergic neurons in the central
amygdala are critically involved in anxiety and stress responsivity
(Kang et al., 2000; Kung et al., 2010) (Fig. 4B).
4.3. Interactions between genetics and behavioral traits
It is clear that multiple factors can converge to contribute to
vulnerability. For example, while PENK genotype and anxiety/
neuroticismtrait areindividually associated
dependence, there was a strong synergism between high-risk ge-
notypes and the negative affect trait that enhanced cannabis
dependence risk 8e9-fold (Jutras-Aswad et al., 2012) (Fig. 5). This
synergistic interaction was also evident in another population
(homogenous Caucasian Greek armyconscripts) inwhich aspects of
cigarette use could be explored separately from cannabis use
(Jutras-Aswad et al., 2012). The finding that cannabis dependence is
significantly enhanced in individuals with both high neuroticism/
anxiety and risk genotypes emphasizes the important synergistic
contribution of negative emotional traits and genetics to vulnera-
bility. Thus, developmental cannabis exposure may confer suscep-
tibility primarily in those individuals with underlying genetic and
behavioral trait (Fig. 6). Both clinical reports and research studies
suggest that coping with stress and negative mood states is a
common motive for use among heavy abusers (Hyman and Sinha,
2009), which would be consistent with self-medicating even sub-
threshold anxietyand negative affect induced by PENK dysfunction.
Cannabis exposure and negative affect may thus interact in a
complex way such that cannabis is used to cope with subthreshold
symptoms, but paradoxically further increases these symptoms in
the long term.
Different lines of evidence suggest a link between adolescent
THC and subsequent vulnerability to addiction and psychiatric risk.
Yet, it is clear that more scientific evidence is critically needed to
fully understand this relationship considering the multiple factors
that appear to influence this trajectory. While some neurobiological
insights have been obtained, it is clear that additional information
is needed to fully understand the dynamic neurodevelopment of
distinct components of the eCB and related neuronal systems, and
the impact of cannabis upon these systems during adolescent
ontogeny. The mechanisms by which cannabis may disrupt the
functional organization of brain structures such as the striatum and
prefrontal cortex during adolescent development as well as specific
behavioral phenotypes are still unknown. Aside from the direct
pharmacological effects of the drug on brain development, indi-
vidual factors contribute tremendously to the complexity of the
relationship between adolescent cannabis exposure and addiction
risk (Fig. 6). The apparent synergistic interactions of genetics and
negative affective trait suggest that genetic screens should begin to
Fig. 6. Schematic overview of the interaction between environmental factors, genetics and behavioral traits that together contribute to complex neuropsychiatric disorders like
addiction. Vulnerability involves a delicate balance between factors that promote and protect against disease, and adolescent THC, an environmental factor, may tip this balance in
teens with high-risk genotypes and behavioral traits.
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
consider behavioral endophenotypes since the gene-addiction risk
relationship is not direct. The possibility to identify vulnerable
adolescents is essential for early intervention of cannabis depen-
dence and related psychiatric disorders. Overall, it is impossible to
ignore the evidence that cannabis/THC is not harmless to the
developing brain, but there remain large gaps of knowledge that
need to be filled in order to help inform public policy, thereby
enhancing teenagers’ well-being and their mental health later in
This work was funded by NIDA DA030359 and DA023214.
Agrawal, A., Lynskey, M.T., 2009. Candidate genes for cannabis use disorders:
findings, challenges and directions. Addiction 104, 518e532.
Agrawal, A., Neale, M.C., Prescott, C.A., Kendler, K.S., 2004. Cannabis and other illicit
drugs: comorbid use and abuse/dependence in males and females. Behav.
Genet. 34, 217e228.
Agrawal, A., Wetherill, L., Dick, D.M., Xuei, X., Hinrichs, A., Hesselbrock, V.,
Kramer, J., Nurnberger Jr., J.I., Schuckit, M., Bierut, L.J., Edenberg, H.J., Foroud, T.,
2009. Evidence for association between polymorphisms in the cannabinoid
receptor 1 (CNR1) gene and cannabis dependence. Am. J. Med. Genet. B Neu-
ropsychiatr. Genet. 150B, 736e740.
Arendt, M., Rosenberg, R., Fjordback, L., Brandholdt, J., Foldager, L., Sher, L., Munk-
Jorgensen, P., 2007. Testing the self-medication hypothesis of depression and
aggression in cannabis-dependent subjects. Psychol. Med. 37, 935e945.
Belue, R.C., Howlett, A.C., Westlake, T.M., Hutchings, D.E., 1995. The ontogeny of
cannabinoid receptors in the brain of postnatal and aging rats. Neurotoxicol.
Teratol. 17, 25e30.
Biegon, A., Kerman, I.A., 2001. Autoradiographic study of pre- and postnatal dis-
tribution of cannabinoid receptors in human brain. Neuroimage 14,1463e1468.
Biscaia, M., Fernandez, B., Higuera-Matas, A., Miguens, M., Viveros, M.P., Garcia-
Lecumberri, C., Ambrosio, E., 2008. Sex-dependent effects of periadolescent
exposure to thecannabinoidagonist
administration behaviour and the endogenous opioid system. Neuropharma-
cology 54, 863e873.
Biscaia, M., Marin, S., Fernandez, B., Marco, E.M., Rubio, M., Guaza, C., Ambrosio, E.,
Viveros, M.P., 2003. Chronic treatment with CP 55,940 during the peri-
adolescent period differentially affects the behavioural responses of male and
female rats in adulthood. Psychopharmacology (Berl) 170, 301e308.
Blume, L.C., Bass, C.E., Childers, S.R., Dalton, G.D., Roberts, D.C., Richardson, J.M.,
Xiao, R., Selley, D.E., Howlett, A.C., 2013. Striatal CB1 and D2 receptors regulate
expression of each other, CRIP1A and delta opioid systems. J. Neurochem. 124,
Brook, J.S., Balka, E.B., Whiteman, M., 1999a. The risks for late adolescence of early
adolescent marijuana use. Am. J. Public Health 89, 1549e1554.
Brook, J.S., Richter, L., Whiteman, M., Cohen, P., 1999b. Consequences of adolescent
marijuana use: incompatibility with the assumption of adult roles. Genet. Soc.
Gen. Psychol. Monogr. 125, 193e207.
Canals, M., Milligan, G., 2008. Constitutive activity of the cannabinoid CB1 receptor
regulates the function of co-expressed Mu opioid receptors. J. Biol. Chem. 283,
Ciccocioppo, R., Antonelli, L., Biondini, M., Perfumi, M., Pompei, P., Massi, M., 2002.
tetrahydrocannabinol and ethanol in rats. Eur. J. Pharmacol. 449, 245e252.
Cleveland, H.H., Wiebe, R.P., 2008. Understanding the association between
adolescent marijuana use and later serious drug use: gateway effect or devel-
opmental trajectory? Dev. Psychopathol. 20, 615e632.
Corchero, J., Garcia-Gil, L., Manzanares, J., Fernandez-Ruiz, J.J., Fuentes, J.A.,
Ramos, J.A., 1998. Perinatal delta9-tetrahydrocannabinol exposure reduces
proenkephalin gene expression in the caudate-putamen of adult female rats.
Life Sci. 63, 843e850.
Corchero, J., Romero, J., Berrendero, F., Fernandez-Ruiz, J., Ramos, J.A., Fuentes, J.A.,
Manzanares, J., 1999. Time-dependent differences of repeated administration
with Delta9-tetrahydrocannabinol in proenkephalin and cannabinoid receptor
gene expression and G-protein activation by mu-opioid and CB1-cannabinoid
receptors in the caudate-putamen. Brain Res. Mol. Brain Res. 67, 148e157.
Creemers, H.E., Harakeh, Z., Dick, D.M., Meyers, J., Vollebergh, W.A., Ormel, J.,
Verhulst, F.C., Huizink, A.C., 2011. DRD2 and DRD4 in relation to regular alcohol
and cannabis use among adolescents: does parenting modify the impact of
genetic vulnerability? the TRAILS study. Drug Alcohol Depend. 115, 35e42.
DiNieri, J.A., Wang, X., Szutorisz, H., Spano, S.M., Kaur, J., Casaccia, P., Dow-
Edwards, D., Hurd, Y.L., 2011. Maternal cannabis use alters ventral striatal
dopamine D2 gene regulation in the offspring. Biol. Psychiatry 70, 763e769.
Dorard, G., Berthoz, S., Phan, O., Corcos, M., Bungener, C., 2008. Affect dysregulation
in cannabis abusers: a study in adolescents and young adults. Eur. Child
Adolescent Psychiatry 17, 274e282.
tetrahydrocannabinol (THC) increases cocaine-stimulated activity in adoles-
cent but not adult male rats. Pharmacol. Biochem. Behav. 100, 587e591.
Durieux, P.F., Bearzatto, B., Guiducci, S., Buch, T., Waisman, A., Zoli, M.,
Schiffmann, S.N., de Kerchove d’Exaerde, A., 2009. D2R striatopallidal neurons
inhibit both locomotor and drug reward processes. Nat. Neurosci. 12, 393e395.
Ellgren, M., Artmann, A., Tkalych, O., Gupta, A., Hansen, H.S., Hansen, S.H., Devi, L.A.,
Hurd, Y.L., 2008. Dynamic changes of the endogenous cannabinoid and opioid
mesocorticolimbic systems during adolescence: THC effects. Eur. Neuro-
psychopharmacol. 18, 826e834.
Ellgren, M., Hurd, Y.L., Franck, J., 2004. Amphetamine effects on dopamine levels
and behavior following cannabinoid exposure during adolescence. Eur. J.
Pharmacol. 497, 205e213.
Ellgren, M., Spano, S.M., Hurd, Y.L., 2007. Adolescent cannabis exposure alters opiate
intakeand opioidlimbic neuronal populations
psychopharmacology 32, 607e615.
Ferdinand, R.F., Sondeijker, F., van der Ende, J., Selten, J.P., Huizink, A., Verhulst, F.C.,
2005. Cannabis use predicts future psychotic symptoms, and vice versa.
Addiction 100, 612e618.
Fergusson, D.M., Boden, J.M., 2008. Cannabis use and later life outcomes. Addiction
103, 969e976. Discussion 977e968.
Fergusson, D.M., Boden, J.M., Horwood, L.J., 2006. Cannabis use and other illicit drug
use: testing the cannabis gateway hypothesis. Addiction 101, 556e569.
Fergusson, D.M., Horwood, L.J., 2000. Does cannabis use encourage other forms of
illicit drug use? Addiction 95, 505e520.
Fergusson, D.M., Horwood, L.J., Swain-Campbell, N., 2002. Cannabis use and psy-
chosocial adjustment in adolescence and young adulthood. Addiction 97,1123e
Frank, M.J., Hutchison, K., 2009. Genetic contributions to avoidance-based de-
cisions: striatal D2 receptor polymorphisms. Neuroscience 164, 131e140.
Frank, M.J., Moustafa, A.A., Haughey, H.M., Curran, T., Hutchison, K.E., 2007. Genetic
triple dissociation reveals multiple roles for dopamine in reinforcement
learning. Proc. Natl. Acad. Sci. U. S. A. 104, 16311e16316.
Gerard, C.M., Mollereau, C., Vassart, G., Parmentier, M., 1991. Molecular cloning of a
human cannabinoid receptor which is also expressed in testis. Biochem. J. 279
(Pt 1), 129e134.
Gerfen, C.R., Enber, T.M., Susel, Z., Chase, T.N., Monsma, F.J., Mahan, L.C., Sibley, D.R.,
1990. D1 and D2 dopamine receptor regulated gene expression of striatonigral
and striatopallidal neurons. Science 250, 1429e1432.
Gerfen, C.R., Young III, W.S., 1988. Distribution of striatonigral and striatopallidal
peptidergic neurons in both patch and matrix compartments: an in situ hy-
bridization histochemistry and fluorescent retrograde tracing study. Brain Res.
Glass, M., Dragnunow, M., Faull, R.L., 1997. Cannabinoid receptors in the human
brain: a detailed anatomical quantitative autoradiographic study in the fetal,
neonatal and adult human brain. Neurosci 77, 299e318.
Gong, J.P., Onaivi, E.S., Ishiguro, H., Liu, Q.R., Tagliaferro, P.A., Brusco, A., Uhl, G.R.,
2006. Cannabinoid CB2 receptors: immunohistochemical localization in rat
brain. Brain Res. 1071, 10e23.
Griffin, G., Tao, Q., Abood, M.E., 2000. Cloning and pharmacological characterization
of the rat CB(2) cannabinoid receptor. J. Pharmacol. Exp. Ther. 292, 886e894.
Hall, W.D., Lynskey, M., 2005. Is cannabis a gateway drug? Testing hypotheses about
the relationship between cannabis use and the use of other illicit drugs. Drug
Alcohol Rev. 24, 39e48.
Harkany, T., Keimpema, E., Barabas, K., Mulder, J., 2008. Endocannabinoid functions
controlling neuronal specification during brain development. Mol. Cell Endo-
crinol. 286, S84eS90.
Hayatbakhsh, M.R., Najman, J.M., Jamrozik, K., Mamun, A.A., Alati, R., Bor, W., 2007.
Cannabis and anxiety and depression in young adults: a large prospective study.
J. Am. Acad. Child Adolesc. Psychiatry 46, 408e417.
Heinz, A., Siessmeier, T., Wrase, J., Hermann, D., Klein, S., Grusser, S.M., Flor, H.,
Braus, D.F., Buchholz, H.G., Grunder, G., Schreckenberger, M., Smolka, M.N.,
Rosch, F., Mann, K., Bartenstein, P., 2004. Correlation between dopamine D(2)
receptors in the ventral striatum and central processing of alcohol cues and
craving. Am. J. Psychiatry 161, 1783e1789.
Heng, L., Beverley, J.A., Steiner, H., Tseng, K.Y., 2011. Differential developmental
trajectories for CB1 cannabinoid receptor expression in limbic/associative and
sensorimotor cortical areas. Synapse 65, 278e286.
Herkenham, M., Lynn, A., Little, M., Johnson, M., Melvin, L., De Costa, B., Rice, K.,
1990. Cannabinoid receptor localization in brain. Proc. Natl. Acad. Sci. U. S. A. 87,
Herkenham, M., Lynn, A.B., de Costa, B.R., Richfield, E.K., 1991a. Neuronal localiza-
tion of cannabinoid receptors in the basal ganglia of the rat. Brain Res. 547,
Herkenham, M., Lynn, A.B., Johnson, M.R., Melvin, L.S., de Costa, B.R., Rice, K.C.,
1991b. Characterization and localization of cannabinoid receptors in rat brain: a
quantitative in vitro autoradiographic study. J. Neurosci. The Off. J. Soc. Neu-
rosci. 11, 563e583.
Hermanson, D.J., Marnett, L.J., 2011. Cannabinoids, endocannabinoids, and cancer.
Cancer Metastasis Rev. 30, 599e612.
Higuera-Matas, A., Soto-Montenegro, M.L., del Olmo, N., Miguens, M., Torres, I.,
Vaquero, J.J., Sanchez, J., Garcia-Lecumberri, C., Desco, M., Ambrosio, E., 2008.
Augmented acquisition of cocaine self-administration and altered brain glucose
metabolism in adult female but not male rats exposed to a cannabinoid agonist
during adolescence. Neuropsychopharmacology 33, 806e813.
inadult rats. Neuro-
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
Hikida, T., Kimura, K., Wada, N., Funabiki, K., Nakanishi, S., 2010. Distinct roles of
synaptic transmission in direct and indirect striatal pathways to reward and
aversive behavior. Neuron 66, 896e907.
Hill, A.J., Williams, C.M., Whalley, B.J., Stephens, G.J., 2012. Phytocannabinoids as
novel therapeutic agents in CNS disorders. Pharmacol. Ther. 133, 79e97.
Hill, E.L., Gallopin, T., Ferezou, I., Cauli, B., Rossier, J., Schweitzer, P., Lambolez, B.,
2007. Functional CB1 receptors are broadly expressed in neocortical GABAergic
and glutamatergic neurons. J. Neurophysiol. 97, 2580e2589.
Hopfer, C.J., Young, S.E., Purcell, S., Crowley, T.J., Stallings, M.C., Corley, R.P.,
Rhee, S.H., Smolen, A., Krauter, K., Hewitt, J.K., Ehringer, M.A., 2006. Cannabis
receptor haplotype associated with fewer cannabis dependence symptoms in
adolescents. Am. J. Med. Genet. B Neuropsychiatr. Genet. 141B, 895e901.
Howlett, A.C., Barth, F., Bonner, T.I., Cabral, G., Casellas, P., Devane, W.A., Felder, C.C.,
Herkenham, M., Mackie, K., Martin, B.R., Mechoulam, R., Pertwee, R.G., 2002.
International Union of Pharmacology. XXVII. Classification of cannabinoid re-
ceptors. Pharmacol. Rev. 54, 161e202.
Hyman, S.M., Sinha, R., 2009. Stress-related factors in cannabis use and misuse:
implications for prevention and treatment. J. Subst. Abuse Treat 36, 400e413.
Johnston, L.D., O’Malley, P.M., Bachman, J.G., Schulenberg, J.E., 2012. Monitoring the
Future National Results on Adolescent Drug Use: Overview of Key Findings,
2011. Institute for Social Research, The University of Michigan., Ann Arbor.
Jutras-Aswad, D., Jacobs, M.M., Yiannoulos, G., Roussos, P., Bitsios, P., Nomura, Y.,
Liu, X., Hurd, Y.L., 2012. Cannabis-dependence risk relates to synergism be-
tween neuroticism and proenkephalin SNPs associated with amygdala gene
expression: case-control study. PloS One 7, e39243.
Kandel, D.B., Yamaguchi, K., Klein, L.C., 2006. Testing the gateway hypothesis.
Addiction 101, 470e472. Discussion 474e476.
Kang, W., Wilson, S.P., Wilson, M.A., 2000. Overexpression of proenkephalin in the
amygdala potentiates the anxiolytic effects of benzodiazepines. Neuro-
psychopharmacol. Off. Publ. Am. Coll. Neuropsychopharmacol. 22, 77e88.
Katona, I., Freund, T.F., 2008. Endocannabinoid signaling as a synaptic circuit
breaker in neurological disease. Nat. Med. 14, 923e930.
Kelley, A.E., Bakshi, V.P., Haber, S.N., Steininger, T.L., Will, M.J., Zhang, M., 2002.
Opioid modulation of taste hedonics within the ventral striatum. Physiol. Behav.
Kendler, K.S., Karkowski, L.M., Neale, M.C., Prescott, C.A., 2000. Illicit psychoactive
substance use, heavy use, abuse, and dependence in a US population-based
sample of male twins. Arch. Gen. Psychiatry 57, 261e269.
Kendler, K.S., Prescott, C.A., 1998. Cannabis use, abuse, and dependence in a
population-based sample of female twins. Am. J. Psychiatry 155, 1016e1022.
Kilmer, J.R., Hunt, S.B., Lee, C.M., Neighbors, C., 2007. Marijuana use, risk perception,
and consequences: is perceived risk congruent with reality? Addict Behav. 32,
Klein, T.A., Neumann, J., Reuter, M., Hennig, J., von Cramon, D.Y., Ullsperger, M.,
2007. Genetically determined differences in learning from errors. Science 318,
Koskinen, J., Lohonen, J., Koponen, H., Isohanni, M., Miettunen, J., 2010. Rate of
cannabis use disorders in clinical samples of patients with schizophrenia: a
meta-analysis. Schizophr. Bull. 36, 1115e1130.
Kung, J.C., Chen, T.C., Shyu, B.C., Hsiao, S., Huang, A.C., 2010. Anxiety- and
depressive-like responses and c-fos activity in preproenkephalin knockout
mice: oversensitivity hypothesis of enkephalin deficit-induced posttraumatic
stress disorder. J. Biomed. Sci. 17, 29.
Lanciego, J.L., Barroso-Chinea, P., Rico, A.J., Conte-Perales, L., Callen, L., Roda, E.,
Gomez-Bautista, V., Lopez, I.P., Lluis, C., Labandeira-Garcia, J.L., Franco, R., 2011.
Expression of the mRNA coding the cannabinoid receptor 2 in the pallidal
complex of Macaca fascicularis. J. Psychopharmacol. 25, 97e104.
Le Moine, C., Normand, E., Guitteny, A.F., Fouque, B., Teoule, R., Bloch, B., 1990.
Dopamine receptor gene expression by enkephalin neurons in rat forebrain.
Proc. Natl. Acad. Sci. U. S. A. 87, 230e234.
Lessem, J.M., Hopfer, C.J., Haberstick, B.C., Timberlake, D., Ehringer, M.A., Smolen, A.,
Hewitt, J.K., 2006. Relationship between adolescent marijuana use and young
adult illicit drug use. Behav. Genet. 36, 498e506.
Lopez-Quintero, C., Neumark, Y., 2010. Effects of risk perception of marijuana use on
marijuana use and intentions to use among adolescents in Bogota, Colombia.
Drug Alcohol Depend. 109, 65e72.
Lynskey, M.T., Coffey, C., Degenhardt, L., Carlin, J.B., Patton, G., 2003. A longitudinal
study of the effects of adolescent cannabis use on high school completion.
Addiction 98, 685e692.
Maccoun, R.J., 2006. Competing accounts of the gateway effect: the field thins, but
still no clear winner. Addiction 101, 473e474. Discussion 474e476.
Maes, H.H., Woodard, C.E., Murrelle, L., Meyer, J.M., Silberg, J.L., Hewitt, J.K.,
Rutter, M., Simonoff, E., Pickles, A., Carbonneau, R., Neale, M.C., Eaves, L.J., 1999.
Tobacco, alcohol and drug use in eight- to sixteen-year-old twins: the Virginia
twin study of adolescent behavioral development. J. Stud. Alcohol 60, 293e305.
Mailleux, P., Parmentier, M., Vanderhaeghen, J.J., 1992. Distribution of cannabinoid
receptor messenger RNA in the human brain: an in situ hybridization histo-
chemistry with oligonucleotides. Neurosci. Lett. 143, 200e204.
Marsicano, G., Lutz, B., 1999. Expression of the cannabinoid receptor CB1 in distinct
neuronal subpopulations in the adult mouse forebrain. Eur. J. Neurosci. 11,
Mato, S., Del Olmo, E., Pazos, A., 2003. Ontogenetic development of cannabinoid
receptor expression and signal transduction functionality in the human brain.
Eur. J. Neurosci. 17, 1747e1754.
Matsuda, L.A., Bonner, T.I., Lolait, S.J., 1993. Localization of cannabinoid receptor
mRNA in rat brain. J. Comp. Neurol. 327, 535e550.
Matsuda, L.A., Lolait, S.J., Brownstein, M.J., Young, A.C., Bonner, T.I., 1990. Structure
of a cannabinoid receptor and functional expression of the cloned cDNA. Nature
McGue, M., Elkins, I., Iacono, W.G., 2000. Genetic and environmental influences on
adolescent substance use and abuse. Am. J. Med. Genet. 96, 671e677.
McLaughlin, C.R., Martin, B.R., Compton, D.R., Abood, M.E., 1994. Cannabinoid re-
ceptors in developing rats: detection of mRNA and receptor binding. Drug and
Alcohol Depend. 36, 27e31.
Michaelides, M., Thanos, P.K., Kim, R., Cho, J., Ananth, M., Wang, G.J., Volkow, N.D.,
2012. PET imaging predicts future body weight and cocaine preference. Neu-
roimage 59, 1508e1513.
Miles, D.R., van den Bree, M.B., Gupman, A.E., Newlin, D.B., Glantz, M.D.,
Pickens, R.W., 2001. A twin study on sensation seeking, risk taking behavior and
marijuana use. Drug Alcohol Depend. 62, 57e68.
Morel, L.J., Giros, B., Dauge, V., 2009. Adolescent exposure to chronic delta-9-
tetrahydrocannabinol blocks opiate dependence in maternally deprived rats.
Neuropsychopharmacology 34, 2469e2476.
Morgan, D., Grant, K.A., Gage, H.D., Mach, R.H., Kaplan, J.R., Prioleau, O., Nader, S.H.,
Buchheimer, N., Ehrenkaufer, R.L., Nader, M.A., 2002. Social dominance in
monkeys: dopamine D2 receptors and cocaine self-administration. Nat. Neu-
rosci. 5, 169e174.
Mulder, J., Aguado, T., Keimpema, E., Barabas, K., Ballester Rosado, C.J., Nguyen, L.,
Monory, K., Marsicano, G., Di Marzo, V., Hurd, Y.L., Guillemot, F., Mackie, K.,
Lutz, B., Guzman, M., Lu, H.C., Galve-Roperh, I., Harkany, T., 2008. Endocanna-
binoid signaling controls pyramidal cell specification and long-range axon
patterning. Proc. Natl. Acad. Sci. U. S. A. 105, 8760e8765.
Munro, S., Thomas, K.L., Abu-Shaar, M., 1993. Molecular characterization of a pe-
ripheral receptor for cannabinoids. Nature 365, 61e65.
Nader, M.A., Morgan, D., Gage, H.D., Nader, S.H., Calhoun, T.L., Buchheimer, N.,
Ehrenkaufer, R., Mach, R.H., 2006. PET imaging of dopamine D2 receptors
during chronic cocaine self-administration in monkeys. Nat. Neurosci. 9, 1050e
Onaivi, E.S., Ishiguro, H., Gong, J.P., Patel, S., Perchuk, A., Meozzi, P.A., Myers, L.,
Mora, Z., Tagliaferro, P., Gardner, E., Brusco, A., Akinshola, B.E., Liu, Q.R., Hope, B.,
Iwasaki, S., Arinami, T., Teasenfitz, L., Uhl, G.R., 2006. Discovery of the presence
and functional expression of cannabinoid CB2 receptors in brain. Ann. N. Y.
Acad. Sci. 1074, 514e536.
Panlilio, L.V., Solinas, M., Matthews, S.A., Goldberg, S.R., 2007. Previous exposure to
THC alters the reinforcing efficacy and anxiety-related effects of cocaine in rats.
Neuropsychopharmacology 32, 646e657.
use and mental health in young people: cohort study. BMJ 325,1195e1198.
Perez-Rosado, A., Manzanares, J., Fernandez-Ruiz, J., Ramos, J.A., 2000. Prenatal
Delta(9)-tetrahydrocannabinol exposure modifies proenkephalin gene expres-
sion in the fetal rat brain: sex-dependent differences. Brain Res. Dev. Brain Res.
Pettit, D.A., Harrison, M.P., Olson, J.M., Spencer, R.F., Cabral, G.A., 1998. Immuno-
histochemical localization of the neural cannabinoid receptor in rat brain.
J. Neurosci. Res. 51, 391e402.
Pistis, M., Perra, S., Pillolla, G., Melis, M., Muntoni, A.L., Gessa, G.L., 2004. Adolescent
exposure to cannabinoids induces long-lasting changes in the response to drugs
of abuse of rat midbrain dopamine neurons. Biol. Psychiatry 56, 86e94.
Rhee, S.H., Hewitt, J.K., Young, S.E., Corley, R.P., Crowley, T.J., Stallings, M.C., 2003.
Genetic and environmental influences on substance initiation, use, and prob-
lem use in adolescents. Arch. Gen. Psychiatry 60, 1256e1264.
Rodriguez de Fonseca, F., Ramos, J.A., Bonnin, A., Fernández-Ruiz, J.J.,1993. Presence
of cannabinoid binding sites in the brain from early postnatal ages. Neuroreport
Sakai, J.T., Hopfer, C.J., Hartman, C., Haberstick, B.C., Smolen, A., Corley, R.P.,
Stallings, M.C., Young, S.E., Timberlake, D., Hewitt, J.K., Crowley, T.J., 2007. Test of
association between TaqIA A1 allele and alcohol use disorder phenotypes in a
sample of adolescent patients with serious substance and behavioral problems.
Drug Alcohol Depend. 88, 130e137.
SAMHSA, 2011. Results from the 2010 National Survey on Drug Use and Health:
National Findings. Office of Applied Studies, NSDUH Series H-41, HHS Publica-
tion No. (SMA) 11e4658, Rockville, MD.
Sano, H., Yasoshima, Y., Matsushita, N., Kaneko, T., Kohno, K., Pastan, I.,
Kobayashi, K., 2003. Conditional ablation of striatal neuronal types containing
dopamine D2 receptor disturbs coordination of basal ganglia function.
J. Neurosci. 23, 9078e9088.
Singh, M.E., McGregor, I.S., Mallet, P.E., 2006. Perinatal exposure to delta(9)-
tetrahydrocannabinol alters heroin-induced place conditioning and fos-
psychopharmacol. 31, 58e69.
Skoubis, P.D., Lam, H.A., Shoblock, J., Narayanan, S., Maidment, N.T., 2005. Endog-
enous enkephalins, not endorphins, modulate basal hedonic state in mice. Eur.
J. Neurosci. 21, 1379e1384.
Spano, M.S., Ellgren, M., Wang, X., Hurd, Y.L., 2007. Prenatal cannabis exposure
increases heroin seeking with allostatic changes in limbic enkephalin systems
in adulthood. Biol. Psychiatry 61, 554e563.
Spano, M.S., Fadda, P., Fratta, W., Fattore, L., 2010. Cannabinoid-opioid interactions
in drug discrimination and self-administration: effect of maternal, postnatal,
adolescent and adult exposure to the drugs. Curr. Drug Targets 11, 450e461.
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424
Tarter, R.E., Vanyukov, M., Kirisci, L., Reynolds, M., Clark, D.B., 2006. Predictors of
marijuana use in adolescents before and after licit drug use: examination of the
gateway hypothesis. Am. J. Psychiatry 163, 2134e2140.
Thanos, P.K., Michaelides, M., Benveniste, H., Wang, G.J., Volkow, N.D., 2008. The
effects of cocaine on regional brain glucose metabolism is attenuated in
dopamine transporter knockout mice. Synapse 62, 319e324.
Tomasiewicz, H.C., Jacobs, M.M., Wilkinson, M.B., Wilson, S.P., Nestler, E.J., Hurd, Y.L.,
2012. Proenkephalin mediates the enduring effects of adolescent cannabis
exposure associated with adult opiate vulnerability. Biol. Psychiatry 72, 803e
Tsuang, M.T., Lyons, M.J., Meyer, J.M., Doyle, T., Eisen, S.A., Goldberg, J., True, W.,
Lin, N., Toomey, R., Eaves, L., 1998. Co-occurrence of abuse of different drugs in
men: the role of drug-specific and shared vulnerabilities. Arch. Gen. Psychiatry
Tucker, J.S., Ellickson, P.L., Collins, R.L., Klein, D.J., 2006. Are drug experimenters
better adjusted than abstainers and users?: a longitudinal study of adolescent
marijuana use. J. Adolesc. Health 39, 488e494.
(P129T) missense variant in cannabis users: studies of drug use and dependence in
Caucasians. Am. J. Med. Genet. B Neuropsychiatr. Genet.144B, 660e666.
Valverde, O., Noble, F., Beslot, F., Dauge, V., Fournie-Zaluski, M.C., Roques, B.P., 2001.
Delta9-tetrahydrocannabinol releases and facilitates the effects of endogenous
enkephalins: reduction in morphine withdrawal syndrome without change in
rewarding effect. Eur. J. Neurosci. 13, 1816e1824.
van den Bree, M.B., Johnson, E.O., Neale, M.C., Pickens, R.W., 1998. Genetic and
environmental influences on drug use and abuse/dependence in male and fe-
male twins. Drug Alcohol Depend. 52, 231e241.
van Gastel, W.A., Maccabe, J.H., Schubart, C.D., Vreeker, A., Tempelaar, W., Kahn, R.S.,
Boks, M.P., 2013. Cigarette smoking and cannabis use are equally strongly
associated with psychotic-like experiences: a cross-sectional study in 1929
young adults. Psychol. Med., 1e9.
Van Sickle, M.D., Duncan, M., Kingsley, P.J., Mouihate, A., Urbani, P., Mackie, K.,
Stella, N., Makriyannis, A., Piomelli, D., Davison, J.S., Marnett, L.J., Di
Marzo, V., Pittman, Q.J., Patel, K.D., Sharkey, K.A., 2005. Identification and
functional characterization of brainstem cannabinoid CB2 receptors. Science
Van Waes, V., Beverley, J.A., Siman, H., Tseng, K.Y., Steiner, H., 2012. CB1 cannabinoid
receptor expression in the striatum: association with corticostriatal circuits and
developmental regulation. Front Pharmacol. 3, 21.
Verdurand, M., Nguyen, V., Stark, D., Zahra, D., Gregoire, M.C., Greguric, I.,
Zavitsanou, K., 2011. Comparison of cannabinoid CB(1) receptor binding in
adolescent and adult rats: a positron emission tomography study using [F]MK-
9470. Int. J. Mol. Imaging 2011, 548123.
Vigano, D., Rubino, T., Parolaro, D., 2005. Molecular and cellular basis of cannabi-
noid and opioid interactions. Pharmacol. Biochem. Behav. 81, 360e368.
Volkow, N.D., Chang, L., Wang, G.J., Fowler, J.S., Ding, Y.S., Sedler, M., Logan, J.,
Franceschi, D., Gatley, J., Hitzemann, R., Gifford, A., Wong, C., Pappas, N., 2001.
Low level of brain dopamine D2 receptors in methamphetamine abusers: as-
sociation with metabolism in the orbitofrontal cortex. Am. J. Psychiatry 158,
Volkow, N.D., Fowler, J.S., Wang, G.J., Swanson, J.M., 2004. Dopamine in drug abuse
and addiction: results from imaging studies and treatment implications. Mol.
Psychiatry 9, 557e569.
Volkow, N.D., Wang, G.J., Fowler, J.S., Logan, J., Gatley, S.J., Gifford, A., Hitzemann, R.,
Ding, Y.S., Pappas, N., 1999. Prediction of reinforcing responses to psychosti-
mulants in humans by brain dopamine D2 receptor levels. Am. J. Psychiatry 156,
Wang, G.J., Volkow, N.D., Fowler, J.S., Logan, J., Abumrad, N.N., Hitzemann, R.J.,
Pappas, N.S., Pascani, K., 1997. Dopamine D2 receptor availability in opiate-
dependent subjects before and after naloxone-precipitated withdrawal. Neu-
ropsychopharmacol. Off. Publ. Am. Coll. Neuropsychopharmacol. 16, 174e182.
Wang, X., Dow-Edwards, D., Andersen, V., Minkoff, H., Hurd, Y.L., 2006. Discrete
opioid gene expression impairment in the human fetal brain associated with
maternal marijuana use. Pharmacogenomic. J. 6.
Wang, X., Dow-Edwards, D., Keller, E., Hurd, Y.L., 2003. Preferential limbic expres-
sion of the cannabinoid receptor mRNA in the human fetal brain. Neuroscience
Yamaguchi, K., Kandel, D.B., 1984. Patterns of drug use from adolescence to young
adulthood: III. Predictors of progression. Am. J. Public Health 74, 673e681.
Zvolensky, M.J., Vujanovic, A.A., Bernstein, A., Bonn-Miller, M.O., Marshall, E.C.,
Leyro, T.M., 2007. Marijuana use motives: a confirmatory test and evaluation
among young adult marijuana users. Addict. Behav. 32, 3122e3130.
Y.L. Hurd et al. / Neuropharmacology 76 (2014) 416e424