Cannabinoids, anxiety, and the periaqueductal gray

Abstract

he use of Cannabis sativa by humans dates back several thousand years, for both its psychotomimetic and potential medicinal properties. As scientific research methods developed, the cannabinoids present in this herb were characterized, as well as their complex interface with the human central nervous system, provided by the activation of specific receptors. The subsequent description of an endogenous cannabinoid system in the mammalian brain shifted the notion of cannabis as a recreational drug to a therapeutic alternative for psychiatric disorders. However, the neuroanatomical sites mediating its effects have remained uncertain. In the present paper, we review recent data suggesting that the midbrain periaqueductal gray may be a structure involved in the anxiolytic-like effects of cannabinoids.

Full text

Psychology & Neuroscience, 2010, 3, 1, 39 - 42
DOI: 10.3922/j.psns.2010.1.004
PSYCHOLOGY
NEUROSCIENCE
Cannabinoids, anxiety, and the periaqueductal gray
Viviane M. Saito and Fabrício A. Moreira
Universidade Federal de Minas Gerais, Brazil
Abstract
The use of Cannabis sativa by humans dates back several thousand years, for both its psychotomimetic and potential medicinal
properties. As scientic research methods developed, the cannabinoids present in this herb were characterized, as well as their
complex interface with the human central nervous system, provided by the activation of specic receptors. The subsequent
description of an endogenous cannabinoid system in the mammalian brain shifted the notion of cannabis as a recreational drug
to a therapeutic alternative for psychiatric disorders. However, the neuroanatomical sites mediating its effects have remained
uncertain. In the present paper, we review recent data suggesting that the midbrain periaqueductal gray may be a structure
involved in the anxiolytic-like effects of cannabinoids. Keywords: cannabis, endocannabinoids, anxiety, periaqueductal gray.
Received 16 February 2010; received in revised form 28 April 2010; accepted 28 April 2010. Available on line 26 June 2010
Viviane M. Saito, Programa de Pós-Graduação em
Neurociências, Universidade Federal de Minas Gerais, Brazil.
Fabrício A. Moreira, Departamento de Farmacologia, Instituto
de Ciências Biológicas, Universidade Federal de Minas Gerais,
Brazil. Correspondence regarding this article should be directed
to: Fabrício A. Moreira, Departamento de Farmacologia, Instituto
de Ciências Biológicas, Universidade Federal de Minas Gerais,
Campus Pampulha, Av. Antônio Carlos, 6627, Belo Horizonte,
MG, CEP 31270-901, Brazil. Tel.: +55-31-3409-2720. E-mail:
fabriciomoreira@icb.ufmg.br
Introduction
The earliest record of Cannabis sativa use by
humans was found approximately 12000 years ago
(Beaulieu & Rice, 2002). Cannabis is known to produce
euphoria, sensory perception imbalance, tachycardia,
antinociception, concentration difculties, memory
impairment, antiemesis, and increased appetite (Ameri,
1999; Di Marzo & Matias, 2005). However, only in
the last half-century have these effects of cannabis
become better understood, attributable to the discovery
and synthesis of its main psychoactive component, Δ9-
tetrahydrocannabinol (THC). The psychotropic effects
of marijuana are attributed to this compound. The plant
has dozens of substances known as phytocannabinoids,
which all share lipophilic properties and are still being
studied for their possible benecial effects in humans.
Characterization of the endocannabinoid system
Cannabinoids are natural or synthetic substances
that resemble or derive from the phytocannabinoids
described above. They induce their pharmacological
effects in vivo by activating two receptor subtypes that
have been identied and cloned: the CB1 receptor, which
is highly expressed in the central nervous system (CNS)
at presynaptic axon terminals (Devane, Dysarz, Johnson,
Melvin, & Howlett, 1988; Matsuda, Lolait, Brownstein,
Young, & Bonner, 1990), and the CB2 receptor, which
is located mainly in peripheral tissues at the level of the
immune system (Munro, Thomas, & Abu-Shaar, 1993).
The brain distribution of CB1 binding sites correlates
with the effects of cannabinoids on emotional states. CB1
receptors are densely expressed in a group of structures
hypothesized to be the neuroanatomical substrate for
stress, aversion, fear, and anxiety, including the prefrontal
cortex, amygdala, hippocampus, periaqueductal gray
(PAG), and hypothalamus (Herkenham et al., 1990).
After the identication of the rst cannabinoid
receptor, the search for an endogenous ligand for this
receptor began. The rst endogenous cannabinoid
ligand, or endocannabinoid, was discovered in
1992 when the arachidonic acid (AA) derivative
arachidonoylethanolamide (AEA; also named
“anandamide” from the Sanskrit “ananda,” meaning
“bliss”) was isolated from porcine brain (Devane
et al., 1992). Anandamide was found to induce
pharmacological actions similar to those of cannabinoids,
such as THC, and to be the principal endogenous ligand
for CB1 receptors. 2-Arachidonoylglycerol (2-AG),
another endocannabinoid and an arachidonic acid
derivative, was then soon discovered by Mechoulam
et al. (1995). These two ligands are considered the
most biologically important endocannabinoids, though
other endocannabinoids have also been described,
including 2-arachidonylglyceryl ether (noladin),
O-arachidonoylethanolamine (virodhamine), and
N-arachidonoyldopamine (De Petrocellis & Di Marzo,

Saito and Moreira
40
2009). Endocannabinoids are believed to be produced from
phospholipid precursors and are released upon demand
after cellular depolarization or receptor stimulation in a
calcium-dependent manner. Endocannabinoids are not
stored in vesicles as classical neurotransmitters—they
are promptly diffused from the postsynaptic neuron
to the synaptic cleft. They then bind to presynaptic
CB1 receptors to exert their effects. By activating CB1
receptors, endocannabinoids can inhibit both excitatory
and inhibitory neurotransmission from presynaptic
terminals (Wilson & Nicoll, 2002). Termination of
endocannabinoid signaling occurs through a reuptake
mechanism (possibly mediated by a selective transporter)
and subsequent enzymatic metabolism. AEA is hydrolyzed
by the enzyme fatty acid amide hydrolase (FAAH),
whereas 2-AG undergoes catabolism by a different
enzyme, monoacylglyceride lipase (De Petrocellis &
Di Marzo, 2009). Altogether, the cannabinoid receptors,
their endogenous ligands, and their respective catabolic
enzymes constitute the endocannabinoid system. Despite
Figure 1. Illustration of endocannabinoid synthesis, retrograde signaling, and uptake.
Presynaptic neuron Postsynaptic neuron
1964 Identication of Δ9-THC as the main psychoactive compound of cannabis
1980 Development of synthetic cannabinoids
1988 Identication of the CB1 receptor
1990 Cloning of the CB1 receptor
1992 Identication of the CB2 receptor
Discovery of the endogenous ligand anandamide
1993 Cloning of the CB2 receptor
1994 Development of a CB1 antagonist
1995 Discovery of endocannabinoids 2-AG and palmitoylethanolamide
1997 Development of a CB2 antagonist
1998 Evidence of the analgesic properties of endocannabinoids
Development of knockout mice lacking the gene expressing CB1 receptor
2000 Development of knockout mice lacking the gene expressing CB2 receptor
(Adapted from Beaulieu & Rice, 2002)
Table 1. Timeline of cannabis pharmacology.

Cannabinoids, anxiety, and PAG 41
enormous advances in this eld, the endocannabinoid
system has not been fully characterized, and many of its
mechanisms remain unknown.
Remarkable advances in cannabinoid pharmacology
have occurred based on the identication of
endocannabinoid compounds and the development of
potent and selective synthetic cannabinoid agonists,
antagonists, and enzymatic blockers.
Cannabinoids and anxiety
The anxiolytic-like effects of cannabinoids have
been investigated in several studies in humans and
animals. Anatomical, biochemical, and pharmacological
evidence indicates that endocannabinoid signaling
is important in the control of emotional behavior,
although its effects may be contradictory in humans
and animals, depending on such variables as drug
dose, genetic background, and environmental context
(Moreira, Aguiar, & Guimarães, 2009; Viveros, Marco,
& File, 2005). Even under recreational conditions,
marijuana users describe paradoxical effects on anxiety
(Zuardi, Crippa, & Guimarães, 2008). In animal studies,
systemic injections of Δ9-THC or its synthetic analogs
have exerted bidirectional effects on anxiety-related
behaviors, eliciting either anxiogenic- or anxiolytic-like
responses. Lower doses of cannabinoids are apparently
anxiolytic, whereas higher doses may exacerbate anxious
behavior (Viveros et al., 2005; Zuardi et al., 2008).
Some of the possible reasons for these controversial
effects include the recruitment of other neurocircuits,
the specic pharmacological pathway investigated, and
the sensitivity of the model employed.
An integrative hypothesis for the biphasic effects
of cannabinoids on anxiety proposes that they may
occur as a result of the distinct roles of CB1 receptors
in different brain regions, in addition to the drug and
dose range used and the differential sensitivity of
the receptors to these compounds (Viveros et al.,
2005). The biphasic effects may also be related to
the close cooperation between the endocannabinoid
system and other neurotransmitter circuits, including
γ-aminobutyric acid and glutamate systems (Moreira et
al., 2009). In addition to direct agonism of cannabinoid
receptors, pharmacological studies may employ other
strategies such as inhibition of endocannabinoid uptake
or metabolism. Pharmacological blockade of FAAH by
URB597 and URB532 produces anxiolytic-like effects
in the elevated zero-maze in adult rats, suggesting that
increased AEA levels may interfere with the modulation
of emotional states. Therefore, FAAH inhibition may
be a valuable approach for anti-anxiety therapy because
it is not accompanied by any signs of cannabinoid
intoxication (Kathuria et al., 2003). Thus, augmentation
of endocannabinoid signaling has anxiolytic effects,
whereas blockade or genetic deletion of CB1 receptors
has anxiogenic properties (for review, see Patel &
Hillard, 2009).
A functional approach for investigating the role
of the endocannabinoid system as part of the complex
circuitry that regulates anxiety arises from the high
density of CB1 receptors in brain structures presumably
involved in anxiety-related responses, such as the PAG.
This midbrain structure has been proposed to be involved
in the perception of pain, “ght-or-ight” responses, and
behaviors related to fear, anxiety, and panic (Del-Ben
& Graeff, 2009). To thoroughly investigate the overall
interaction between the endocannabinoid system and
the PAG on anxiety-like behaviors, experiments with
intracerebral drug injections have been performed in
rats. The effects of intra-PAG injections of cannabinoids
have been evaluated in diverse animal models.
Moreira et al. (2007) showed that direct AEA
injection into the PAG increased exploration of the
open arms of the elevated plus maze, an animal model
predictive of anxiety-like behavior. This anxiolytic-
like effect was prevented by pretreatment with the CB1
receptor antagonist AM251, whereas the AEA uptake/
metabolism inhibitor AM404 potentiated this effect.
Subsequent studies extended these data to other models.
Local injection of cannabinoids into the PAG also
induced anxiolytic-like effects in the Vogel conict test,
an animal model based on the suppression of punished
responses (Lisboa, Resstel, Aguiar, & Guimarães,
2008). Direct activation of CB1 receptors by intra-PAG
injection of AEA or AM404 also blocked the expression
of conditioned responses; this effect was blocked by
a CB1 antagonist, implicating CB1 receptors in the
modulation of conditioned responses (Resstel, Lisboa,
Aguiar, Corrêa, & Guimarães, 2008).
The bimodal effects on anxiety produced by
pharmacological tools that enhance endocannabinoid
signaling appear to rely also on ne differences between
the animal models employed to investigate its inuence
on behavior (Moreira et al., 2009). Nevertheless,
because of its reliable, stable effects on anxiety,
blockade of endocannabinoid hydrolysis via inactivation
of FAAH appears to be a promising alternative for the
development of anti-anxiety drugs, although many
aspects of the endocannabinoid system and the utility of
other cannabinoids remain uncertain.
Conclusion
The endocannabinoid system is a ubiquitous
signaling system that has important regulatory
functions in the CNS. Extensive biochemical and
neurobiological research has attempted to elucidate
how the brain interacts with the major components
of Cannabis sativa and their endogenous analogs, the
endocannabinoids. The distribution of cannabinoid
receptors in specic structures in the mammalian brain

Saito and Moreira
42
indicates that the endocannabinoid system participates
in the regulation of mood and emotional behaviors. CB1
receptors are broadly distributed throughout the PAG,
which has been considered an important component of
a network of brain structures that process anxiety-like
behavior (Graeff & Del-Ben, 2008). Animal models and
pharmacological manipulation of the endocannabinoid
system, either by stimulating the production and release
of endocannabinoids or by blocking their catabolism,
have suggested the importance of this system in
emotional behavior. Scientic research has explored
the controversial, bimodal effects of cannabinoids on
human anxiety, revealing the delicate balance between
the potential therapeutic use and unique side-effect
prole of cannabis and cannabinoids. Any attempt
to develop endocannabinoid-based therapeutics for
anxiety disorders must weigh the benets against any
undesirable or unpredictable effects.
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