Neuropsychopharmacology (2013) 38, 341–349 & 2013 American College of Neuropsychopharmacology. All rights reserved 0893-133X/13 www.neuropsychopharmacology.org Effect of Schizophrenia Risk-Associated Alleles in SREB2 (GPR85) on Functional MRI Phenotypes in Healthy Volunteers 1,3 1,4 1,5 1 1,5 Eugenia Radulescu , Fabio Sambataro , Venkata S Mattay , Joseph H Callicott , Richard E Straub , 2 1,5 ,1 Mitsuyuki Matsumoto , Daniel R Weinberger and Stefano Marenco* 1 2 Clinical Brain Disorders Branch, GCAP, IRP, NIMH, Bethesda, MD, USA; CNS, Astellas Research Institute of America LLC, Skokie, IL, USA Genetic variants in GPR85 (SREB2: rs56080411 and rs56039557) have been associated with risk for schizophrenia. Here, we test the hypothesis that these variants impact on brain function in normal subjects, measured with functional magnetic resonance imaging (fMRI) paradigms that target regions with greatest SREB2 expression (hippocampal formation and amygdaloid complex). During a facial emotion recognition paradigm, a significant interaction of rs56080411 genotype by sex was found in the left amygdaloid complex (male risk allele carriers showed less activation than male homozygotes for the non-risk allele, while females showed the opposite pattern). During aversive encoding of an emotional memory paradigm, we found that risk allele carriers for rs56080411 had greater activation in the right inferior frontal gyrus. Trends in the same direction were present for rs56039557 in the right occipital cortex and right fusiform gyrus. During a working memory paradigm, a significant sex-by-genotype interaction was found with male risk allele carriers of rs56080411 having inefficient activation within the left dorsolateral prefrontal cortex (DLPFC), compared with same sex non-risk carriers, while females revealed an opposite pattern, despite similar levels of performance. These data suggest that risk-associated variants in SREB2 are associated with phenotypes similar to those found in patients with schizophrenia in the DLPFC and the amygdala of males, while the pattern is opposite in females. The findings in females and during the emotional memory paradigm are consistent with modulation by SREB2 of brain circuitries implicated in mood regulation and may be relevant to neuropsychiatric conditions other than schizophrenia. Neuropsychopharmacology (2013) 38, 341–349; doi:10.1038/npp.2012.184; published online 12 September 2012 Keywords: emotion; hippocampal formation; amygdala; dorsolateral prefrontal cortex; single-nucleotide polymorphisms INTRODUCTION GPCR subfamilies (Matsumoto et al, 2000). The unique characteristic of the SREB family is the high conservation of G protein-coupled receptors (GPCRs) are the largest family the amino-acid sequence across species: for example, SREB2 of membrane surface receptors, essential for signal trans- has an identical primary amino-acid sequence in duction in central nervous system, and a preferred humans, rats, and mice, which suggests a critical function therapeutic target for psychiatric and neurological disorders of the protein. To date, an endogenous ligand of SREB (Gilchrist, 2007; Kobilka and Deupi, 2007). The SREB has not been identified. Nevertheless, SREB2 has been (Super Conserved Receptor Expressed in Brain) family— implicated in schizophrenia both because of statistical SREB1 (GPR27), SREB2 (GPR85), and SREB3 (GPR173)—is association of gene variants with risk for illness and because a particular subfamily of the GPCRs. They share 52–63% transgenic mice overexpressing SREB2 display behaviors amino-acid identity and have lower homology to other that are compatible with some models of schizophrenia (Matsumoto et al, 2008). *Correspondence: Dr S Marenco, Clinical Brain Disorders Branch, SREB2, located on Chromosome 7q31, is an orphan GPCR GCAP, IRP, NIMH, 10 Center Drive, Building 10, Room 3C103, of 7.5 kB that generates nine transcripts encoding for four Bethesda, MD 20892, USA, Tel: +1 301 4358964, Fax: +1 0301 proteins (ECgene; http://genome.ewha.ac.kr/) through alter- 4807795, E-mail: [email protected] native splicing. Our laboratory recently demonstrated that 3Current address: Clinical Imaging Sciences Centre (CISC), Rm 006, two single-nucleotide polymorphisms (SNPs) in SREB2 Brighton and Sussex Medical School, Brighton BN1 9RR, UK 4 were nominally associated with schizophrenia risk Current address: Brain Center for Motor and Social Cognition, Italian (Matsumoto et al, 2008). Specifically, the minor alleles of Institute of Technology, Via Volturno 39/E, 43100 Parma, Italy 0 5 SNP rs56080411 (3 untranslated region—of the third exon) Current address: Lieber Institute for Brain Development, Johns and SNP rs56039557 (second intron) were overtransmitted Hopkins University Medical Campus, 855N Wolfe Street, Baltimore, MD 21205, USA to affected individuals in families (Matsumoto et al, 2008). This work was presented in part at the 2007 ACNP meeting and at the SNP rs56039557 was also associated with risk for schizo- 2008 meeting of the Society for Neuroscience. phrenia in the independent National Institute of Mental Received 13 March 2012; revised 6 August 2012; accepted 7 August Health Genetics Initiative (NIMHGI) cohort (Wood et al, 2012 2002). Furthermore, voxel-based morphometry—a commonly Genetic variation in SREB2 affects fMRI phenotypes E Radulescu et al 342 used MRI method for detecting global and regional volu- and emotional memory (Andreano and Cahill, 2009). metric changes in the brain—revealed an association Moreover, sex modulates the expression and possibly the between the same risk alleles (rs56080411 and rs56039557) frequency of psychiatric disorders, including depression and reduced hippocampal volume in schizophrenia subjects and schizophrenia (Abel et al, 2010). but not in normal volunteers (Matsumoto et al, 2008). More recently, a possible role of SREB2 in autism (Trikalinos et al, 2006; Voineagu et al, 2011) and possibly MATERIALS AND METHODS attention deficit hyperactivity disorder (Anney et al, 2008), Subjects Tourette’s syndrome and intellectual disability (Patel et al, 2011) has emerged. Thus, it is conceivable that genetic Healthy volunteers were recruited from the National variation in SREB2 might contribute to a spectrum of Institute of Mental Health Genetic Study of Schizophrenia neurodevelopmental psychiatric disorders, analogous to other (NCT00001486: the Clinical Brain Disorders Branch ‘Sibling genetic risk factors associated with risk for schizophrenia. Study’), a study of neurobiological abnormalities related to Previous work in our laboratory showed that SREB2 genetic risk for schizophrenia (Dr Weinberger, PI). The transcripts are expressed virtually in all neurons but most participants included in this study had available genotypes abundantly in hippocampal dentate gyrus and subventri- for two SREB2 risk-associated SNPs (rs56080411 and cular zone of the amygdala (Matsumoto et al, 2005). This rs56039557) and quality controlled neuroimaging data from expression pattern is consistent with the behavioral profile three fMRI studies: an emotionally variable picture encod- of the SREB2 transgenic mice that show alterations in ing task (PICT: 114 subjects), an emotional face matching contextual fear conditioning, ie, the behavioral response to task (FMT: 354 subjects) and a working memory task a stressful cage environment, which is mediated by (n-back: 100 subjects). The 3 subsamples partly overlapped amygdala and hippocampal formation (HF) in rodents. (45 subjects participated in all 3 studies). More recently, Chen et al (2012) showed that SREB2 affects Only subjects of self-described European ancestry were adult neurogenesis and neurogenesis-dependent learning in included in the study, to minimize the potential for a transgenic mouse model. Given these data, a reasonable population stratification artifacts. The demographic data are prediction is that the function of the amygdalar– presented in Table 1. Subjects were screened with extensive hippocampal system would be preferentially affected in clinical interviews and examinations, including the SCID individuals carrying risk-associated alleles for rs56080411 interview for DSM-IV diagnoses. Inclusion criteria were no and rs56039557 as compared with non-carriers, even in history of alcohol or drug abuse, no history of psychiatric normal subjects. In this paper, we use the blood oxygena- (Axis I and Cluster A Axis II), neurological or medical tion level-dependent (BOLD) signal during fMRI as an conditions, and no current psychotropic medication. All the index of brain function during the execution of tasks subjects gave written informed consent before the study, specifically designed to engage the amygdala and HF. according to the guidelines of National Institute of Mental Predicting the directionality of this effect is not straightfor- Health Institutional Review Board, who approved the study. ward given the lack of knowledge of SREB2’s endogenous ligand and function and the effect of specific alleles on Genotyping. Standard methods to extract DNA from SREB2 processing in human brain. Moreover, a review of white blood cells with the PUREGENE DNA purification the literature linking amygdalar and hippocampal BOLD to kit (Gentra Systems; Minneapolis, MN, USA) were used. The the genetics of schizophrenia reveals conflicting informa- two SNPs (rs56080411 and rs56039557) were genotyped tion (see Supplementary Material). using the Taq Man 50-exonuclease allelic discrimination In contrast to work on fMRI in hippocampus and assay