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. 2010 Jun 9;5(6):e11038.
doi: 10.1371/journal.pone.0011038.

Altered ratio of D1 and D2 dopamine receptors in mouse striatum is associated with behavioral sensitization to cocaine

Dawn Thompson  1 , Lene Martini, Jennifer L Whistler
Affiliations

Altered ratio of D1 and D2 dopamine receptors in mouse striatum is associated with behavioral sensitization to cocaine

Dawn Thompson et al. PLoS One. .

Abstract

Background: Drugs of abuse elevate brain dopamine levels, and, in vivo, chronic drug use is accompanied by a selective decrease in dopamine D2 receptor (D2R) availability in the brain. Such a decrease consequently alters the ratio of D1R:D2R signaling towards the D1R. Despite a plethora of behavioral studies dedicated to the understanding of the role of dopamine in addiction, a molecular mechanism responsible for the downregulation of the D2R, in vivo, in response to chronic drug use has yet to be identified.

Methods and findings: ETHICS STATEMENT: All animal work was approved by the Gallo Center IACUC committee and was performed in our AAALAC approved facility. In this study, we used wild type (WT) and G protein coupled receptor associated sorting protein-1 (GASP-1) knock out (KO) mice to assess molecular changes that accompany cocaine sensitization. Here, we show that downregulation of D2Rs or upregulation of D1Rs is associated with a sensitized locomotor response to an acute injection of cocaine. Furthermore, we demonstrate that disruption of GASP-1, that targets D2Rs for degradation after endocytosis, prevents cocaine-induced downregulation of D2Rs. As a consequence, mice with a GASP-1 disruption show a reduction in the sensitized locomotor response to cocaine.

Conclusions: Together, our data suggests that changes in the ratio of the D1:D2R could contribute to cocaine-induced behavioral plasticity and demonstrates a role of GASP-1 in regulating both the levels of the D2R and cocaine sensitization.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Generation of GASP-1 knockout mice.
A. Targeting vector design for generating GASP-1 KO mice. A cassette expressing the G418 resistance gene flanked by lox P sites was inserted into the intron upstream of the GASP-1 open reading frame (ORF) (intron 4) and a third loxP site was inserted in the intron downstream of the GASP ORF (intron 5). ES cells from C57/Bl6 mice were transfected with this vector. Properly targeted clones (see B, C) were transfected with Cre-recombinase and blastocysts from clones in which the GASP-1 ORF was disrupted were implanted into C57/BL6 females. B, C. Southern blotting analysis identified homologous recombination and single insertion using B. 5′ and C. Neo probes. D. Immuno-detection of GASP-1 in KO mice. Whole brain lysates from WT, HET and KO mice were prepared, separated by SDS-PAGE, transferred to nitrocellulose and immunoblotted for GASP-1 using a GASP specific antibody (upper panel) or β-Actin (lower panel) to control for protein loading.
Figure 2
Figure 2. GASP-1 knockout mice exhibit reduced cocaine sensitization.
A-D. Cocaine sensitization in WT and GASP-1 KO mice. Mice were injected daily (IP) with saline (WT n = 27, GASP-1 KO n = 25) or cocaine (15 mg/kg, WT n = 29, GASP-1 KO n = 30) and the distance traveled reported for A. 15 min or B. 60 min post-injection. WT mice exhibited a sensitized locomotor response to cocaine by day 3 (WT cocaine, day 3 versus day 1, ##p<0.01 as analyzed by one-way ANOVA followed by Bonferroni multiple comparison tests between corresponding days), while GASP-1 KOs developed a significantly lower sensitized response within this first 15 min by day 4 (GASP-KO cocaine, day 4 versus day 1 ##p<0.01 as analyzed by one-way ANOVA followed by Bonferroni multiple comparison tests between corresponding days). GASP-1 KO mice showed significantly reduced sensitized locomotor response to cocaine compared to WT mice on all days (GASP-1 KO cocaine versus WT cocaine where *p<0.05, **p<0.01 and ***p<0.001 as analyzed by two-way ANOVA followed by Bonferroni multiple comparison tests between corresponding days). GASP-1 KO mice showed reduced locomotor responsiveness to cocaine after repeated treatment both 15 min (C. ***p<0.001) and 60 min post-injection (D. *p<0.05). E, F. Stereotypic behaviors of WT and GASP-1 KO mice after 1 (E) and 5 (F) days of cocaine exposure. On day 5 there were significant differences in stereotypy at 5, 10 and 15 min (GASP-1 KO cocaine versus WT cocaine where ***p<0.001, **p<0.01 and *p<0.05 respectively as analyzed by two-way ANOVA followed by Bonferroni multiple comparison tests between corresponding days) post-injection between cocaine treated genotypes. Data is presented as mean ± S.E.M.
Figure 3
Figure 3. Reduced D2 receptor number in WT mice treated with cocaine.
A. D1R number in striatal membranes was not affected by cocaine treatment in either genotype when assessed by saturation binding using [3H]-SCH23390. B. Cocaine treatment reduced D2R number in WT mice (**p<0.01 WT cocaine versus WT saline). Saline treated GASP-1 KO showed reduced D2R number (GASP-1 KO saline versus WT saline where *p<0.05) and D2R number was increased in cocaine treated GASP-1 KO mice (#p<0.05 GASP-1 KO cocaine versus GASP-1 KO saline). D2R number was assessed by saturation binding using [3H]-raclopride. Data shown are normalized to saline treated groups and are from three independent radioligand binding experiments, performed in duplicate, with at least eight mice per group.
Figure 4
Figure 4. Repeated quinpirole treatment augmented cocaine-induced behavioral responding.
A, Dose response of quinpirole pretreatment. WT C57/BL6 mice were injected daily (IP) with saline or one of three doses of quinirole (0.5, 1 or 5 mg/kg n = 8 per group) for 7 days. On day 8 all mice were injected (IP) with 20 mg/kg cocaine and their locomotor activity recorded. B. Mice become tolerant to the locomotor-suppressing effects of quinpirole. WT C57/BL6 mice were injected daily (IP) with saline (n = 24) or quinpirole (5 mg/kg, n = 24) and their locomotor activity recorded once per day for 7 days. On days 1 and 2, quinpirole reduced locomotor activity when compared to saline treated animals (***p<0.001 and *p<0.05 respectively). WT mice became tolerant to the locomotor inhibitory effects of quinpirole on day 4 and remained so for days 6 and 7 compared to day 1 (#p<0.05 and ###p<0.001 respectively). C, D. Repeated quinpirole pre-treatment augments locomotor response to acute cocaine. On day 8, mice from (B) were injected with cocaine (IP, 20 mg/kg) and their locomotor activity (D) and stereotypy (E) recorded. Mice pretreated with repeated quinpirole showed enhanced locomotor and stereotypic responses to acute cocaine compared to saline treated mice (***p<0.001, **p<0.01 and *p<0.05). Data was analyzed by either one or two-way ANOVA with Bonferroni multiple comparison tests and presented as mean ± S.E.M.
Figure 5
Figure 5. Repeated aripiprazole treatment augmented cocaine-induced behavioral responding.
A. Aripiprazole does not induce endocytosis of D2Rs. HEK 293 cells stably expressing N-terminal FLAG-tagged D2Rs were incubated with M1 antibody to the extracellular tag for 20 mins then stimulated with quinpirole or aripiprazole (10 µM), or left untreated for 30 mins. Cells were fixed, blocked, permeabilised and receptors visualized with fluorescent secondary antibody Alexa 488. Quinpirole but not aripiprazole induced endocytosis of the D2R. B. Acute aripiprazole inhibits locomotion. WT C57/BL6 mice were injected daily (IP) with vehicle (n = 24) or aripiprazole (5 mg/kg, n = 24) and their locomotor activity recorded once per day for 7 days. Aripiprazole reduced locomotor activity when compared to vehicle treated animals across all 7 days (*p<0.05, ***p<0.001) and there was no significant change in the effect of aripiprazole and any day compared to day 1. C, D. Repeated aripiprazole pretreatment augments locomotor response to acute cocaine. On day 8, mice from (B) were injected with cocaine (IP, 20 mg/kg) and their locomotor activity (D) and stereotypy (E) recorded. Mice pretreated with repeated aripiprazole showed enhanced locomotor and stereotypic responses to acute cocaine compared to saline treated mice (***p<0.001, **p<0.01 and *p<0.05). Data was analyzed by either one or two-way ANOVA with Bonferroni multiple comparison tests and presented as mean ± S.E.M.
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References

    1. Fehr C, Yakushev I, Hohmann N, Buchholz HG, Landvogt C, et al. Association of low striatal dopamine d2 receptor availability with nicotine dependence similar to that seen with other drugs of abuse. Am J Psychiatry. 2008;165:507–514. - PubMed
    1. Wang GJ, Volkow ND, Fowler JS, Logan J, Abumrad NN, et al. Dopamine D2 receptor availability in opiate-dependent subjects before and after naloxone-precipitated withdrawal. Neuropsychopharmacology. 1997;16:174–182. - PubMed
    1. Hietala J, West C, Syvalahti E, Nagren K, Lehikoinen P, et al. Striatal D2 dopamine receptor binding characteristics in vivo in patients with alcohol dependence. Psychopharmacology (Berl) 1994;116:285–290. - PubMed
    1. Volkow ND, Wang GJ, Maynard L, Fowler JS, Jayne B, et al. Effects of alcohol detoxification on dopamine D2 receptors in alcoholics: a preliminary study. Psychiatry Res. 2002;116:163–172. - PubMed
    1. Volkow ND, Chang L, Wang GJ, Fowler JS, Ding YS, et al. Low level of brain dopamine D2 receptors in methamphetamine abusers: association with metabolism in the orbitofrontal cortex. Am J Psychiatry. 2001;158:2015–2021. - PubMed

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