Chronic treatment with cannabinoid receptor agonist, CP 55,940, alters GLT1 expression in adolescent rats

Introduction

Methods

Results

Selected References

Discussion & Future directions

Support

SungHa Lee and George V. RebecProgram in Neuroscience, Indiana University, Bloomington

• 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.

• 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).

• In this study, we tested whether chronic treatment with the non-selective cannabinoid receptor agonist, CP 55,940, in adolescent rats alter the levels of GLT1 in the brain reward circuit.

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.

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 anti-GLT1 antibody as the primary antibody at a dilution 1:1500 and horseradish peroxidase secondary antibody as the secondary

• 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).

• 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).

• 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.

• 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

NIH Grant NIDA R01 DA-02451University Kentucky Foundation Grant, NIH P50 DA-05312

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.

1. VTA 2. NAc

3. STR 4. PFC

5. HIP

Neuronal activityImpulsive behaviorGLT1 expression

Chronic cannabinoid treatment during adolescence

Drug seeking behavior

NAc