Expression of Neuronal Trace Amine-associated Receptor (Taar) mRNAs in Leukocytes

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Abstract

Trace amines such as tyramine, octopamine and β-phenylethylamine bind with high affinity to the mammalian trace amine-associated receptor 1 (Taar1), potentially activating G-proteins in the synaptic membranes of target neurons. Recently there has been significant interest in Taar1, since this receptor can bind certain psychoactive drugs of abuse such as Ecstasy (3,4-methylenedioxymethamphetamine). Surprisingly, Ecstasy has been shown to alter responses of immune cells, and we questioned whether Taar receptors might be responsible for this effect. Using sensitive and quantitative RT-PCR assays, we found no detectable expression of Taar mRNA in bone marrow, or in primary cultures of mouse macrophages and dendritic cells whether quiescent or activated by exposure to lipopolysaccharide or the mouse gamma herpesvirus-68 (γHV-68). Mouse B cells and NK cells isolated from spleen, however, showed expression of several Taar mRNA species. Taar mRNA expression was also upregulated in human peripheral blood lymphocytes following in vitro stimulation with PHA. These studies represent the first to define expression of the mRNAs encoding these trace amine receptors in leukocytes.

Introduction

Trace amines such as tyramine, octopamine and β-phenylethylamine are present at low concentrations in the mammalian central nervous system (Berry, 2004, Branchek and Blackburn, 2003, Burchett and Hicks, 2006, Lindemann and Hoener, 2005, Premont et al., 2001) and in peripheral tissues (Zucchi et al., 2006), and bind with high affinity to Taar1 (Borowsky et al., 2001, Bunzow et al., 2001), a member of the trace amine-associated receptor (Taar) family of genes and proteins (Lindemann et al., 2005, Lindemann and Hoener, 2005). Binding to the receptor is thought to activate G-proteins in the synaptic membranes of target neurons, and when expressed in HEK293 cells, a variety of drugs, including Ecstasy (3,4-methylenedioxymethamphetamine), stimulate the production of cAMP. Ecstasy's EC50 of 1.7 μM (Bunzow et al., 2001) suggests that the typical micromolar serum levels reached by drug abusers could activate cells expressing the Taar1 protein, providing a mechanism for the direct action of Ecstasy on cells and tissues.

While Ecstasy abuse has been shown to affect immune responses (Connor, 2004), the exact mechanisms responsible for such effects are not altogether clear. This lack of understanding has limited our ability to define targets of immunomodulation for this drug of abuse. Teenagers and young adults are the age group most likely to abuse Ecstasy (Landry, 2002), as this drug leads to elevated moods, extroversion, and enhancement of sensual experiences (Downing, 1986, Freese et al., 2002). Thus the use of Ecstasy at parties and “raves” appears to promote close-contact behaviors between individuals (Freese et al., 2002, Gahlinger, 2004, Ropero-Miller and Goldberger, 1998, Smith et al., 2002). Such behaviors, by themselves, would appear to increase transmission of a variety of infectious diseases. However if Ecstasy abuse also limits a protective immune response, then such infections might be more numerous or more severe in these teenagers and young adults. It is difficult to make such inferences without a more defined understanding of the mechanisms by which Ecstasy can alter immunity to microbial infections.

The high affinity of Ecstasy for binding to Taar1 (Bunzow et al., 2001) makes this receptor an attractive candidate for mediating the immunomodulatory effects of this drug of abuse. In fact, Taar1 mRNA has been reported to be expressed in a variety of peripheral tissues including liver, kidney, spleen, pancreas, heart and the gastrointestinal tract (Bunzow et al., 2001). Unfortunately, there have been no studies to determine whether Taar1, or any Taar proteins, are expressed within leukocyte subpopulations. We initiated this study to clearly define Taar mRNA expression in cultured macrophages and dendritic cells, prior to, and during pathogen activation. Although we were unable to detect Taar mRNA expression in macrophages or dendritic cells, an extension of this study demonstrated that mouse B lymphocytes and NK cells express Taar mRNAs. Taar mRNA expression was also upregulated in human peripheral blood lymphocytes following in vitro stimulation with PHA. These studies represent the first to define expression of the mRNAs encoding these trace amine receptors in leukocytes.

Section snippets

Mice

Six to eight week old female C57BL/6 mice (18–22 g) were purchased from Jackson Laboratories (Bar Harbor, ME) and housed in filter top cages containing sterile bedding. Mice were fed chow and water ad libitum and housed for at least 5 days after arrival before being used.

Isolation of bone marrow cells

Mice were anesthetized with isoflurane and sacrificed by cervical dislocation. Bone marrow cells were isolated by flushing femurs with RPMI 1640 tissue culture medium supplemented with 10% heat-inactivated and filtered FBS,

No detectable Taar1 mRNA expression in mouse macrophages and dendritic cells

To assess the ability of immune cells to express Taar1 mRNA, we initially followed our standard procedures for preparing total RNA and cDNA from macrophages and dendritic cells. Fig. 1 shows representative results following PCR amplification of cDNA samples using primers specific for Taar1, IL-6, and GADPH. In un-stimulated and LPS stimulated macrophage and dendritic cell cultures, Taar1 gene amplification was observed (Fig. 1). Control PCR reactions demonstrated the LPS-induced activation of

Discussion

A survey of Ecstasy abusers suggested that there might be a correlation between abuse of this drug and increased numbers of infections (Parrott et al., 2002). Surveying novice, moderate, and heavy users of Ecstasy, Parrott et al. (2002) found that the perceived number of infections reported by these individuals increased with increasing usage of the drug. Specifically, 35% of heavy Ecstasy users and approximately 10% of moderate users reported increased problems with infections. Despite this

Acknowledgements

This work was supported by the National Institute of Drug Abuse (NIDA) grant DA021833.

References (54)

  • LindemannL. et al.

    Trace amine-associated receptors form structurally and functionally distinct subfamilies of novel G protein-coupled receptors

    Genomics

    (2005)
  • MossnerR. et al.

    Role of serotonin in the immune system and in neuroimmune interactions

    Brain Behav. Immun.

    (1998)
  • NashJ.F. et al.

    Effect of the R(−) and S(+) isomers of MDA and MDMA on phosphatidyl inositol turnover in cultured cells expressing 5-HT2A or 5-HT2C receptors

    Neurosci. Lett.

    (1994)
  • NelsonD.A. et al.

    Expression of hemokinin 1 mRNA by murine dendritic cells

    J. Neuroimmunol.

    (2004)
  • PacificiR. et al.

    Effects of repeated doses of MDMA (“ecstasy”) on cell-mediated immune response in humans

    Life Sci.

    (2001)
  • Ropero-MillerJ.D. et al.

    Recreational drugs. Current trends in the 90s

    Clin. Lab. Med.

    (1998)
  • ScheicherC. et al.

    Dendritic cells from mouse bone marrow: in vitro differentiation using low doses of recombinant granulocyte-macrophage colony-stimulating factor

    J. Immunol. Methods

    (1992)
  • StefuljJ. et al.

    mRNA expression of serotonin receptors in cells of the immune tissues of the rat

    Brain Behav. Immun.

    (2000)
  • ThomasD.M. et al.

    Microglial activation is a pharmacologically specific marker for the neurotoxic amphetamines

    Neurosci. Lett.

    (2004)
  • BerryM.D.

    Mammalian central nervous system trace amines. Pharmacologic amphetamines, physiologic neuromodulators

    J. Neurochem.

    (2004)
  • BorowskyB. et al.

    Trace amines: identification of a family of mammalian G protein-coupled receptors

    Proc. Natl. Acad. Sci. U. S. A.

    (2001)
  • BowmanC.C. et al.

    Cyclooxygenase-2-mediated prostaglandin E2 production in mesenteric lymph nodes and in cultured macrophages and dendritic cells after infection with Salmonella

    J. Immunol.

    (2004)
  • BunzowJ.R. et al.

    Amphetamine, 3,4-methylenedioxymethamphetamine, lysergic acid diethylamide, and metabolites of the catecholamine neurotransmitters are agonists of a rat trace amine receptor

    Mol. Pharmacol.

    (2001)
  • Cloez-TayaraniI. et al.

    Differential effect of serotonin on cytokine production in lipopolysaccharide-stimulated human peripheral blood mononuclear cells: involvement of 5-hydroxytryptamine2A receptors

    Int. Immunol.

    (2003)
  • ConnorT.J.

    Methylenedioxymethamphetamine (MDMA, ‘Ecstasy’): a stressor on the immune system

    Immunology

    (2004)
  • ConnorT.J. et al.

    Methylenedioxymethamphetamine suppresses production of the proinflammatory cytokine tumor necrosis factor-alpha independent of a beta-adrenoceptor-mediated increase in interleukin-10

    J. Pharmacol. Exp. Ther.

    (2005)
  • DowningJ.

    The psychological and physiological effects of MDMA on normal volunteers

    J. Psychoact. Drugs

    (1986)
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