The effect of clozapine on mRNA expression for genes encoding G protein-coupled receptors and the protein components of clathrin-mediated endocytosis

Sharp, Sally I.a; Hu, Yingc; Weymer, Jon F.a; Rizig, Miea; McQuillin, Andrewa; Hunt, Stephen P.b; Gurling, Hugh M.D.a

Psychiatric Genetics:
doi: 10.1097/YPG.0b013e32835fe51d
Original Articles
Abstract

Objectives: Clathrin-mediated endocytosis (CME) is an intracellular trafficking mechanism for packaging cargo, including G protein-coupled receptors (GPCRs), into clathrin-coated vesicles (CCVs). The antipsychotic chlorpromazine inhibits CCV assembly of adaptor protein AP2 whereas clozapine increases serotonin2A receptor internalization. We hypothesized that clozapine alters the expression of CME genes modulating vesicle turnover and GPCR internalization.

Materials and methods: SH-SY5Y human neuroblastoma cells were incubated with clozapine (1–20 µmol/l) for 24–72 h. GPCR and CME-related gene mRNA expression was measured using RT-PCR. We quantified changes in the same genes using expression data from a microarray study of mice brains after 12 weeks of treatment with 12 mg/kg/day clozapine.

Results: The expression of genes encoding adaptor and clathrin assembly proteins, AP2A2, AP2B1, AP180, CLINT1, HIP1, ITSN2, and PICALM, increased relative to the control in SH-SY5Y cells incubated with 5–10 µmol/l clozapine for 24–72 h. The microarray study showed significantly altered expression of the above CME-related genes, with a marked 641-fold and 17-fold increase in AP180 and the serotonin1A GPCR, respectively. The expression of three serotonergic receptor and lysophosphatidic acid receptor 2 (EDG4) GPCR genes was upregulated in SH-SY5Y cells incubated with 5 µmol/l clozapine for 24 h. EDG4 expression was also increased with 10–20 µmol/l clozapine treatment at 48–72 h. Clozapine significantly decreased the expression of β-arrestin, involved in GPCR desensitization, both in vitro and vivo.

Conclusion: The changes we report in CME and GPCR mRNAs implicate CCV-mediated internalization of GPCRs and the serotonergic system in clozapine’s mechanism of action, which may be useful in the design of more effective and less toxic antipsychotic therapies.

Author Information

aMolecular Psychiatry Laboratory, UCL Mental Health Sciences Unit

bDepartment of Anatomy & Developmental Biology, University College London, London, UK

cDepartment of Stomatology, Beijing Institute of Dental Research, Capital University of Medical Sciences, Beijing, People’s Republic of China

Correspondence to Sally I. Sharp, PhD, Molecular Psychiatry Laboratory, UCL Mental Health Sciences Unit, University College London, Rockefeller Building, 21 University Street, London WC1E 6JJ, UK Tel: +44 20 31082342; fax: +44 20 31082194; e-mail: s.sharp@ucl.ac.uk

This is an open access article distributed under the Creative Commons Attribution License 4.0 (CCBY), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.http://creativecommons.org/licenses/by-nc-nd/4.0.

Received March 2, 2012

Received in revised form October 9, 2012

Accepted October 31, 2012

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