title: The atypical antipsychotics clozapine and olanzapine promote down-regulation and display functional selectivity at human 5-HT7 receptors.
abstract: OBJECTIVE
  Classically, ligands of GPCRs have been classified primarily upon their affinity and efficacy to activate a signal transduction pathway. Recent reports indicate that the efficacy of a particular ligand can vary depending on the receptor-mediated response measured (e.g. activating G proteins, other downstream responses, internalization). Previously, we reported that inverse agonists induce both homo- and heterologous desensitization, similar to agonist stimulation, at the Gs -coupled 5-HT7 receptor. The primary objective of this study was to determine whether different inverse agonists at the 5-HT7 receptor also induce internalization and/or degradation of 5-HT7 receptors.
  
  
  METHODS
  HEK293 cells expressing 5-HT7(a, b or d) receptors were pre-incubated with 5-HT, clozapine, olanzapine, mesulergine or SB269970 and their effects upon receptor density, AC activity, internalization, recruitment of β-arrestins and lysosomal trafficking were measured.
  
  
  RESULTS
  The agonist 5-HT and three out of four inverse agonists tested increased internalization independently of β-arrestin recruitment. Among these, only the atypical antipsychotics clozapine and olanzapine promoted lysosomal sorting and reduced 5-HT7 receptor density (∼60% reduction within 24 h). Inhibition of lysosomal degradation with chloroquine blocked the clozapine- and olanzapine-induced down-regulation of 5-HT7 receptors. Incubation with SB269970 decreased both 5-HT7(b) constitutive internalization and receptor density but increased 5-HT7(d) receptor density, indicating differential ligand regulation among the 5-HT7 splice variants.
  
  
  CONCLUSIONS
  Taken together, we found that various ligands differentially activate regulatory processes governing receptor internalization and degradation in addition to signal transduction. Thus, these data extend our understanding of functional selectivity at the 5-HT7 receptor.
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.|K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: K W
authors.initials: KW
authors.lastname: Andressen
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.|K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: O
authors.initials: O
authors.lastname: Manfra
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.
authors.forename: C H
authors.initials: CH
authors.lastname: Brevik
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.|K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: A H
authors.initials: AH
authors.lastname: Ulsund
authors.affiliation: Laboratory of Eukaryotic Gene Expression and Signal Transduction, Ghent University, Ghent, Belgium.|Intrexon ActoBiotics N.V., Technologiepark 4, Zwijnaarde, Belgium.
authors.forename: P
authors.initials: P
authors.lastname: Vanhoenacker
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.|K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: F O
authors.initials: FO
authors.lastname: Levy
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway.|K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: K A
authors.initials: KA
authors.lastname: Krobert
mesh_terms: D014150:Antipsychotic Agents / Q000494:pharmacology*; D019390:Arrestins / Q000378:metabolism; D001569:Benzodiazepines / Q000494:pharmacology*; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology; D003024:Clozapine / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects*; D054314:Drug Inverse Agonism; D004873:Ergolines / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D000077152:Olanzapine; D010636:Phenols / Q000494:pharmacology; D011869:Radioligand Assay; D011985:Receptors, Serotonin / Q000235:genetics / Q000378:metabolism*; D012701:Serotonin / Q000494:pharmacology; D012702:Serotonin Antagonists / Q000494:pharmacology; D017366:Serotonin Receptor Agonists / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology; D000071557:beta-Arrestins
chemical_list: D014150:Antipsychotic Agents; D019390:Arrestins; D004873:Ergolines; D008024:Ligands; D010636:Phenols; D011985:Receptors, Serotonin; C404922:SB 269970; D012702:Serotonin Antagonists; D017366:Serotonin Receptor Agonists; D013449:Sulfonamides; D000071557:beta-Arrestins; C104425:serotonin 7 receptor; D001569:Benzodiazepines; D012701:Serotonin; D002738:Chloroquine; D003024:Clozapine; D000077152:Olanzapine; C031649:mesulergine