Disruption of a dopamine receptor complex amplifies the actions of cocaine

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Abstract

Cocaine-induced increases in dopamine signaling in nucleus accumbens (NAc) play a significant role in cocaine seeking behavior. The majority of cocaine addiction research has focused on neuroanatomically segregated dopamine D1 and D2 receptor-expressing neurons, yet an involvement for those NAc neurons coexpressing D1 and D2 receptors in cocaine addiction has never been explored. In situ proximity ligation assay, confocal fluorescence resonance energy transfer and coimmunoprecipitation were used to show native D1 and D2 receptors formed a heteromeric complex in D1/D2 receptor-coexpressing neurons in rat and non-human primate NAc. D1–D2 heteromer expression was lower in NAc of adolescent rats compared to their adult counterparts. Functional disruption of the dopamine D1–D2 receptor heteromer, using a peptide targeting the site of interaction between the D1 and D2 receptor, induced conditioned place preference and increased NAc expression of ∆FosB. D1–D2 heteromer disruption also resulted in the promotion, exacerbation and acceleration of the locomotor activating and incentive motivational effects of cocaine in the self-administration paradigm. These findings support a model for tonic inhibition of basal and cocaine-induced reward processes by the D1–D2 heteromer thus highlighting its potential value as a novel target for drug discovery in cocaine addiction. Given that adolescents show increased drug abuse susceptibility, an involvement for reduced D1–D2 heteromer function in the heightened sensitivity to the rewarding effects of cocaine in adolescence is also implicated.

Introduction

Cocaine addiction is a chronic disorder associated with uncontrollable craving together with compulsive drug-seeking and drug-taking, which persists despite devastatingly negative health, social and economic consequences. It is well established that increased dopamine signaling and activation of dopamine D1 receptors in nucleus accumbens (NAc) plays a significant role in the development and maintenance of cocaine seeking (Anderson and Pierce, 2005, Nestler, 2005). Yet despite this knowledge, therapeutic targeting of specific neuronal pathways in vivo remains difficult to accomplish and, as such, the mainstay of current addiction therapy remains substitution, which has significant limitations, low success rates and high propensity for relapse. Thus the identification of a specific receptor substrate that regulates addiction processes could completely shift drug discovery paradigms.

In the basal ganglia dopamine D1 and D2 receptors are predominantly segregated to distinct neuronal pathways, the direct striatonigral and indirect striatopallidal pathways respectively, and are thought to be the predominant mediators of dopamine effects in this region. However, a fraction of striatal medium spiny neurons (MSNs) express both receptors with colocalization of D1 and D2 receptors occurring predominantly in NAc (Aubert et al., 2000, Bertran-Gonzalez et al., 2008, Deng et al., 2006, Gangarossa et al., 2013, Perreault et al., 2010). Dopamine signaling in NAc has been widely demonstrated to play a pivotal role in cocaine addiction (Anderson and Pierce, 2005, Lammel et al., 2014), yet the contribution of these D1/D2 receptor-coexpressing MSNs in mediating addiction processes has been ignored. As a therapeutic target with proven efficacy in cocaine addiction has yet to be identified, discerning the physiological relevance of these MSNs is imperative to discover novel avenues for drug discovery.

Up to 17–38% of D1-expressing MSNs also express the D2 receptor in specific NAc subregions of the mouse, rat and monkey (Aubert et al., 2000, Bertran-Gonzalez et al., 2008, Deng et al., 2006, Gangarossa et al., 2013, Perreault et al., 2010) whereas in the rodent caudate nucleus (CN) neuronal cell bodies only 1–6% of such MSNs exhibit D1 and D2 receptor colocalization (Bertran-Gonzalez et al., 2008, Perreault et al., 2010). Within most of the D1/D2 receptor-coexpressing MSNs in NAc the receptors exist within a complex forming the D1–D2 receptor heteromer (Perreault et al., 2010). The D1–D2 heteromer was shown to exhibit distinct pharmacological and functional properties (Lee et al., 2004, Rashid et al., 2007) and to induce the activation or expression of proteins known to be highly involved in addiction such as calcium calmodulin kinase II and brain-derived neurotrophic factor (Hasbi et al., 2009, Ng et al., 2010, Perreault et al., 2012a). The role of the D1–D2 heteromer in mediating the behavioral and biochemical effects of cocaine was evaluated.

Section snippets

Animals

Adult male Sprague-Dawley rats (Charles River, Canada) weighing 300–350 g at the start of each experiment and a total of two adult male Macaca fascicularis primates (body weight 3.8–4.5 kg) were used. Homozygous whole body D1 and D2 receptor gene deleted mice were congenic, backcrossed into a C57BL/6J genetic background.

TAT-D1 and scrambled peptide

The TAT-D1 peptide was generated and its specificity tested as described using different strategies including mutations, truncations, BRET and pharmacological studies (Hasbi et

Dopamine D1–D2 heteromers exist in rodent and monkey striatum

To demonstrate the existence of native D1–D2 heteromer in situ in MSNs in brain two methods were employed, the proximity ligation assay (PLA) and confocal fluorescence resonance energy transfer (FRET). The specificity of the primary antibodies for D1 and D2 receptors was validated by immunochemistry in HEK293T cells individually expressing each of the five dopamine receptor subtypes (Lee et al., 2004) and in striatal sections from D1-/- and D2-/- receptor gene-deleted mice (Appendix A, Figure S1

Discussion

In the present study we showed conclusively that there exists a dopamine receptor complex, the dopamine D1–D2 receptor heteromer, in the NAc of rodents and non-human primates. Pharmacological analysis together with a highly selective interfering peptide further showed that the receptor complex had signaling characteristics distinct from its constituent D1 or D2 receptors, being coupled to the Gq protein with the promotion of intracellular calcium signaling through activation of phospholipase C.

Contributors

MLP, AH, PJF, JLL and SRG designed research; MLP, AH, TF, MS, GN, JLL and RF performed research; MLP, AH, MLP, RF, JLL and SRG analyzed data; and MLP, AH and SRG wrote the paper.

Conflict of interest

The authors declare no conflict of interests.

Acknowledgments

This work was supported by a Grant from the National Institute on Drug Abuse (DA-007223 to S.R.G). S.R.G. holds a Canada Research Chair in Molecular Neuroscience. M.L.P was supported by a NARSAD Young Investigator Award (Brain and Behavior Research Foundation).

Each funding source had no further role in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decision to submit the paper for publication.

References (64)

  • E.J. Nestler et al.

    The mesolimbic dopamine reward circuit in depression

    Biol. Psychiatry

    (2006)
  • J.L. Neumeyer et al.

    Receptor affinities of dopamine D1 receptor-selective novel phenylbenzazepines

    Eur. J. Pharmacol.

    (2003)
  • J. Ng et al.

    Activation of calcium/calmodulin-dependent protein kinase IIalpha in the striatum by the heteromeric D1-D2 dopamine receptor complex

    Neuroscience

    (2010)
  • B.F. O׳Dowd et al.

    Two amino acids in each of D1 and D2 dopamine receptor cytoplasmic regions are involved in D1-D2 heteromer formation

    Biochem Biophys. Res. Commun.

    (2012)
  • G.J. O׳Sullivan et al.

    SK&F 83822 distinguishes adenylyl cyclase from phospholipase C-coupled dopamine D1-like receptors: behavioural topography

    Eur. J. Pharmacol.

    (2004)
  • M.L. Perreault et al.

    Regulation of c-fos expression by the dopamine D1-D2 receptor heteromer

    Neuroscience

    (2015)
  • M.L. Perreault et al.

    Reduced striatal dopamine D1-D2 receptor heteromer expression and behavioural subsensitivity in juvenile rats

    Neuroscience

    (2012)
  • M.L. Perreault et al.

    The dopamine D1-D2 receptor heteromer localizes in dynorphin/enkephalin neurons: increased high affinity state following amphetamine and in schizophrenia

    J. Biol. Chem.

    (2010)
  • M.Y. Shen et al.

    Rapid anti-depressant and anxiolytic actions following dopamine D1-D2 receptor heteromer inactivation

    Eur. Neuropsychopharmacol.

    (2015)
  • E. Zakharova et al.

    Sensitivity to cocaine conditioned reward depends on sex and age

    Pharmacol. Biochem. Behav.

    (2009)
  • E. Zakharova et al.

    Differential effects of methamphetamine and cocaine on conditioned place preference and locomotor activity in adult and adolescent male rats

    Behav. Brain Res.

    (2009)
  • O. Aizman et al.

    Anatomical and physiological evidence for D1 and D2 dopamine receptor colocalization in neostriatal neurons

    Nat. Neurosci.

    (2000)
  • J.J. Anker et al.

    Reinstatement of cocaine seeking induced by drugs, cues, and stress in adolescent and adult rats

    Psychopharmacology

    (2010)
  • I. Aubert et al.

    Phenotypical characterization of the neurons expressing the D1 and D2 dopamine receptors in the monkey striatum

    J. Comp. Neurol.

    (2000)
  • J. Bertran-Gonzalez et al.

    Opposing patterns of signaling activation in dopamine D1 and D2 receptor-expressing striatal neurons in response to cocaine and haloperidol

    J. Neurosci.

    (2008)
  • D.K. Biezonski et al.

    Evidence for limited D1 and D2 receptor coexpression and colocalization within the dorsal striatum of the neonatal mouse

    J. Comp. Neurol.

    (2015)
  • H.C. Brenhouse et al.

    Delayed extinction and stronger reinstatement of cocaine conditioned place preference in adolescent rats, compared to adults

    Behav. Neurosci.

    (2008)
  • H.M. Cates et al.

    Threonine 149 phosphorylation enhances DeltaFosB transcriptional activity to control psychomotor responses to cocaine

    J. Neurosci.

    (2014)
  • S.Z. Chao et al.

    D(1) dopamine receptor stimulation increases GluR1 phosphorylation in postnatal nucleus accumbens cultures

    J. Neurochem.

    (2002)
  • J. Chen et al.

    Chronic Fos-related antigens: stable variants of deltaFosB induced in brain by chronic treatments

    J. Neurosci.

    (1997)
  • L.S. Chun et al.

    D1-D2 dopamine receptor synergy promotes calcium signaling via multiple mechanisms

    Mol. Pharmacol.

    (2013)
  • A.M. Deveney et al.

    Pharmacological characterization of behavioural responses to SK&F 83959 in relation to ׳D1-like׳ dopamine receptors not linked to adenylyl cyclase

    Br. J. Pharmacol.

    (1995)
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