Olanzapine Increases RGS7 Protein Expression via Stimulation of the Janus Tyrosine Kinase-Signal Transducer and Activator of Transcription Signaling Cascade Rakesh K. Singh, Ju Shi, Bozena W. Zemaitaitis, and Nancy A. Muma Department of Pharmacology and Experimental Therapeutics, Loyola University Chicago, Stritch School of Medicine, Maywood, Illinois Received January 26, 2007; accepted March 27, 2007 ABSTRACT Atypical antipsychotics such as olanzapine have high affinity for multiple monoamine neurotransmitter receptors and are the mainstay of pharmacological therapy for treatment of schizo- phrenia. In addition to blocking monoamine receptors, these drugs also affect intracellular signaling cascades. We now re- port that 24-h treatment with 300 nM olanzapine causes de- sensitization of serotonin (5-HT) 2A receptors in A1A1v cells, a rat cortical cell line, as indicated by a reduction in inositol phosphate accumulation following stimulation with a 5-HT 2A/2C receptor agonist (-)-1-(2,5-dimethoxy-4-lodophenyl)-2-amino- propane HCl. Olanzapine treatment for 24 h increased the levels of 5-HT 2A receptors in both cytosol (234  34% of control level) and membrane fractions (206  14% of control levels) and RGS7 proteins in both cytosol (193  32% of control levels) and membrane fractions (160  18% of control levels) as measured on Western blots. Increased phosphoryla- tion of Janus tyrosine kinase (JAK) 2 and increased phosphor- ylation and nuclear translocation of signal transducer and ac- tivator of transcription (STAT) 3 with 24-h olanzapine treatment demonstrate activation of the JAK-STAT signaling cascade. Pretreatment with a JAK inhibitor, AG490 [-cyano-(3,4-dihy- droxy)-N-benzylcinnamide], prevented the olanzapine-induced increase in membrane RGS7 protein levels; AG490 alone had no effect on RGS7 protein levels. We verified that treatment with AG490 reduced phosphorylation of JAK2 and inhibited the nuclear localization of phospho-STAT3. Interestingly, treatment with the JAK inhibitor had no effect on 5-HT 2A receptor protein levels. These data suggest that olanzapine-induced activation of the JAK-STAT signaling cascade causes increased expres- sion of RGS7 protein, which in turn could mediate desensitiza- tion of 5-HT 2A receptor signaling caused by olanzapine be- cause RGS7 binds to G q protein and accelerates GTP hydrolysis. Atypical antipsychotics are widely prescribed for the treat- ment of schizophrenia. They are classified as atypical be- cause of their ability to achieve antipsychotic effects with lower rates of extrapyramidal side effects compared with first generation antipsychotics such as haloperidol. In addition, selected atypical antipsychotics also improve certain aspects of cognitive function in schizophrenic patients, whereas typ- ical antipsychotics may worsen cognition (Meltzer et al., 1999). Atypical antipsychotics improved side effects, and ef- ficacy has been attributed to the high-affinity interaction with 5-HT 2A receptors (Kasper et al., 1999). Atypical anti- psychotics have also been shown to block other 5-HT 2 recep- tor subtypes, mainly 5-HT 2B and 5-HT 2C (Lucaites et al., 1996). However, only 5-HT 2C receptor antagonism is sug- gested in contributing to the atypical antipsychotic effects (Herrick-Davis et al., 2000; Rauser et al., 2001). The 5-HT 2A receptor subtype has been implicated in vari- ous psychiatric disorders including depression, anxiety, and schizophrenia (Glennon et al., 1984). Olanzapine is an atyp- ical antipsychotic, approved for the treatment of schizophre- nia and bipolar disorder. Olanzapine has been also studied in the treatment of disorders such as substance abuse, aggres- sion/violence, borderline personality disorder, and obsessive- compulsive disorder (Littrell et al., 2006). Atypical antipsychotics as well as a specific 5-HT 2A recep- tor antagonist, MDL 100,907, desensitize 5-HT 2A -mediated responses (Willins et al., 1999). However, the molecular This study was supported by United States Public Health Service Grant MH068612. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.107.120386. ABBREVIATIONS: 5-HT, serotonin; MDL 100,907, (+)--(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidinemethanol; ERK, extracel- lular receptor kinase; JAK, Janus tyrosine kinase; STAT, signal transducers and activators of transcription; RGS, regulator of G protein signaling; AG490, -cyano-(3,4-dihydroxy)-N-benzylcinnamide; DOI, (-)-1-(2,5-dimethoxy-4-lodophenyl)-2-aminopropane HCl; PBS, phosphate-buffered saline; IP, inositol phosphate. 0022-3565/07/3221-133–140$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 322, No. 1 Copyright © 2007 by The American Society for Pharmacology and Experimental Therapeutics 120386/3214214 JPET 322:133–140, 2007 Printed in U.S.A. 133