Transcript Chronic treatment with cannabinoid receptor

Chronic treatment with cannabinoid receptor agonist, CP 55,940,
alters GLT1 expression in adolescent rats
SungHa Lee and George V. Rebec
Program in Neuroscience, Indiana University, Bloomington
• In this study, we tested whether chronic treatment with the nonselective cannabinoid receptor agonist, CP 55,940, in adolescent
rats alter the levels of GLT1 in the brain reward circuit.
• Chronic exposure to CP 55,940 (n=3) increased GLT1 expression
in the nucleus accumbens (NAc), prefrontal cortex (PFC), striatum
(STR), and hippocampus (HIP), but down-regulated its expression
in the ventral tegmental area (VTA) relative to control (n=3).
1. VTA
optical density (% of
control)
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Control
3. STR
4. PFC
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CP 55,940
CP 55,940
Optical density (%of control)
Control
CP 55,940
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• These preliminary data suggest that chronic exposure of CP
55,940 during adolescence may alter glutamate transmission in the
forebrain motive circuit via changes in GLT1 expression.
However, such changes seem reversible if there is no additional
treatment.
• Along with changes in glutamate transmission, the effects of
chronic adolescent cannabinoids exposure on neurobehavioral
changes also need to be examined. We have been investigating
whether a cannabinoid receptor agonist increases impulsive
behavior, a risk factor for drug addiction. In the future, we will use
several measurements of impulsive behavior such as Go/No-Go
task in responding to test behavioral differences between rats
treated with CP 55,940 and vehicle. Moreover, single-unit activity
can be measured simultaneously while animals perform such
behavioral tasks. Specially, neurons in the orbital frontal cortex,
which has been implicated in impulsivity, is of special interest.
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Control
CP 55,940
5. HIP
Chronic cannabinoid treatment
during adolescence
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GLT1 expression
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Impulsive behavior
Neuronal activity
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Control
CP 55,940
Drug seeking behavior
• However, 20 days after the last injection, the CP 55,940-induced
increase in GLT1 expression returned to the baseline, suggesting
that the effect of CP 55,940 is reversible (n=4; control=2 vs. CP
55,940=2).
Optical density (% of control)
150
Western blot : Brain tissue were collected on pd 39 (24 hour after the
last injection) and pd 58 (20 days after) to test the change of GLT1 is
reversible. Extracted proteins were separated in 4-20% glycine gel and
then transferred onto a nitrocelluose membrane electrophysiology 20
V for 2h 30min. Immunodetection analysis was performed on the
blotted membrane with SNAP i.d. (Millipore) using guinea pig antiGLT1 antibody as the primary antibody at a dilution 1:1500 and
horseradish peroxidase secondary antibody as the secondary
Discussion & Future directions
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Methods
Drug treatment : Sprague Dawley rats received daily injections of CP
55,940 (intraperitoneal injection, 0.4mg/kg) or vehicle between
postnatal (pd) day 28 and 38.
2. NAc
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Optical density (% of control)
• Among various neurotransmitters, glutamate, an excitatory amino
acid, appears to play a critical role in drug-seeking behavior. Once
glutamate is released, it is removed by active uptake. Glutamate
transporter 1 (GLT1) is responsible for most glutamate uptake. We
recently found that increased expression of GLT1 in the nucleus
accumbens, a key component of the forebrain motive circuit,
prevents cocaine relapse (Sari, Smith et al., 2009).
Results
opitical density (% of
control)
• Cannabis, also known as marijuana, is the most widely used illicit
drug, especially prevalent among adolescents and young adults.
Cannabis has been suggested as a gateway drug in that its use
usually precedes the use of more addictive drugs such as cocaine
and heroin. The debate whether cannabis use causes the
progression to the use of other drugs has lasted for decades. In
spite of widely spreading use of cannabis among adolescents, little
is known about its neurobiological effects. Because adolescent
brains undergo a critical period of neural development, it is
necessary to investigate how cannabis exposure affects the brain
reward circuit in this early period and its potential role in increasing
the propensity to use other illicit drugs.
antibody. Equivalent protein loading was assessed by GAPDH as a
loading control. After incubation with HRP kit, membranes were
exposed to Kodak Biomax MR film. Digitized images were
quantified using an image analysis system.
Opitical density (% of
control)
Introduction
Selected References
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Control
CP 55,940
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VTA
NAc
Nac
STR
PFC
HIP
• Sari and Smith et al. (2009), J. of Neurosci., 29(29); 9239-9243
• LaLumiere and Kalivas (2008), J. of Neurosci., 28(23);6046-6053
• Kreek et al. (2005), Nature, 8, 1450-1457
Support
NIH Grant NIDA R01 DA-02451
University Kentucky Foundation Grant, NIH P50 DA-05312