Arrestin3 mediates D 2 dopamine receptor internalization

Molecular Neuropharmacology Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892, USA.
Synapse (Impact Factor: 2.43). 07/2009; 63(7):621-4. DOI: 10.1002/syn.20636
Source: PubMed


Available from: Annika Thorsell, Jun 02, 2015
  • [Show abstract] [Hide abstract]
    ABSTRACT: This review attempts to summarize the current status in relation to the use of positron emission tomography (PET) imaging in the assessment of synaptic concentrations of endogenous mediators in the living brain. Although PET radioligands are now available for more than 40 CNS targets, at the initiation of the Innovative Medicines Initiative (IMI) "Novel Methods leading to New Medications in Depression and Schizophrenia" (NEWMEDS) in 2009, PET radioligands sensitive to an endogenous neurotransmitter were only validated for dopamine. NEWMEDS work-package 5, "Cross-species and neurochemical imaging (PET) methods for drug discovery", commenced with a focus on developing methods enabling assessment of changes in extracellular concentrations of serotonin and noradrenaline in the brain. Sharing the workload across institutions, we utilized in vitro techniques with cells and tissues, in vivo receptor binding and microdialysis techniques in rodents, and in vivo PET imaging in non-human primates and humans. Here, we discuss these efforts and review other recently published reports on the use of radioligands to assess changes in endogenous levels of dopamine, serotonin, noradrenaline, γ-aminobutyric acid, glutamate, acetylcholine, and opioid peptides. The emphasis is on assessment of the availability of appropriate translational tools (PET radioligands, pharmacological challenge agents) and on studies in non-human primates and human subjects, as well as current challenges and future directions. PET imaging directed at investigating changes in endogenous neurochemicals, including the work done in NEWMEDS, have highlighted an opportunity to further extend the capability and application of this technology in drug development.
    Psychopharmacology 04/2015; DOI:10.1007/s00213-015-3938-6 · 3.99 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: A high-throughput screening campaign was conducted to interrogate a 380,000+ small molecule library for novel D2 dopamine receptor modulators using a calcium mobilization assay. Active agonist compounds from the primary screen were examined for orthogonal D2 dopamine receptor signaling activities including cAMP modulation and β-arrestin recruitment. While the majority of the subsequently confirmed hits activated all signaling pathways tested, several compounds showed a diminished ability to stimulate β-arrestin recruitment. One such compound (MLS1547) is a highly efficacious agonist at D2 receptor-mediated G protein-linked signaling, but does not recruit β-arrestin, as demonstrated using two different assays. This compound does, however, antagonize dopamine-stimulated β-arrestin recruitment to the D2 receptor. In an effort to investigate the chemical scaffold of MLS1547 further, we characterized a set of 24 analogs of MLS1547 with respect to their ability to inhibit cAMP accumulation or stimulate β-arrestin recruitment. A number of the analogs were similar to MLS1547 in that they displayed agonist activity for inhibiting cAMP accumulation, but did not stimulate β-arrestin recruitment (i.e., they were highly biased). In contrast, other analogs displayed various degrees of G protein signaling bias. These results provided the basis to use pharmacophore modeling and molecular docking analyses to build a preliminary structure-activity relationship of the functionally-selective properties of this series of compounds. In summary, we have identified and characterized a novel G protein-biased agonist of the D2 dopamine receptor and identified structural features that may contribute to its biased signaling properties.
    Molecular pharmacology 04/2014; 86(1). DOI:10.1124/mol.113.090563 · 4.12 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Arrestins mediate desensitization and internalization of G protein-coupled receptors, and also direct receptor signaling towards heterotrimeric G protein-independent signaling pathways. We previously identified a four-residue segment (residues 212-215) of the dopamine D2 receptor that is necessary for arrestin binding in an in vitro heterologous expression system but that also impairs receptor expression. We now describe the characterization of additional mutations at that arrestin binding site in the third intracellular loop (IL3). Mutating two (residues 214-215) or three (residues 213-215) of the four residues to alanine partially decreased agonist-induced recruitment of arrestin3 without altering activation of a G protein. Arrestin-dependent receptor internalization, which requires arrestin binding to β2-adaptin (the β2 subunit of the clathrin-associated adaptor protein AP2) and clathrin, was disproportionately affected by the three-residue mutation, with no agonist-induced internalization observed even in the presence of overexpressed arrestin or G protein-coupled receptor kinase 2 (GRK2). The disjunction between arrestin recruitment and internalization could not be explained by alterations in the time course of the receptor-arrestin interaction, the recruitment of GRK2, or the receptor-induced interaction between arrestin and β2-adaptin, suggesting that the mutation impairs a property of the internalization complex that has not yet been identified.
    Journal of Biological Chemistry 10/2014; DOI:10.1074/jbc.M114.605378 · 4.60 Impact Factor