Genetic variation in schizophrenia-risk-gene dysbindin 1 modulates brain activation in anterior cingulate cortex and right temporal gyrus during language production in healthy individuals
Introduction
Schizophrenia is highly heritable (> 80%), but the search for chromosomal loci and genes has been slow and arduous because of small single effects and interactions with epigenetic processes and environmental factors (Owen et al., 2002). Nevertheless, molecular studies using positional genetics identified several candidates including catechol-O-methyltransferase (COMT), dysbindin 1 (DTNBP1), neuregulin 1 (NRG1), regulator of G-protein signalling 4 (RGS4), G72, proline dehydrogenase (PRODH), dopamine D2 receptor (DRD2), disrupted-in-schizophrenia 1 (DISC1) and D-amino acid oxidase (DAAO). Recent studies yielded evidence that some of the genes listed above, such as COMT (for a review see Harrison and Weinberger, 2005), DTNBP1 (Burdick et al., 2006, Burdick et al., 2007, Donohoe et al., 2007, Zinkstok et al., 2007, Luciano et al., 2009), NRG1 (Hall et al., 2006), DISC1 (Burdick et al., 2005), and G72 (Goldberg et al., 2006) have an influence on cognition in patients with schizophrenia and healthy subjects.
Straub et al. (2002) identified and genotyped 17 SNPs from DTNBP1 gene, mostly intronic, which showed association to schizophrenia. A number of subsequent studies could replicate the results for several of these SNPs, including the SNP rs1018381 (P 1578 by Straub et al., 2002), in worldwide populations (Schwab et al., 2003, Tang et al., 2003, Van Den Bogaert et al., 2003, van den Oord et al., 2003, Funke et al., 2004, Kirov et al., 2004, Numakawa et al., 2004). Although one meta-analysis pointed to inconsistency of the positively associated genetic variations at DTNBP1 in the different European samples (Mutsuddi et al., 2006), two meta-analyses supported the more positive than negative association between genetic variations in DTNBP1 and schizophrenia (Owen et al., 2004, Williams et al., 2005).
In the brain, dysbindin 1 binds to β-dystrobrevin, a member of the dystrophin protein complex (DPC), which has been implicated in synaptic structure and signalling of the glutamatergic neuronal system (Benson et al., 2001). The DPC is located at postsynaptic densities throughout various regions of the brain (Blake et al., 1999), and disruption of its function might result in structural and functional abnormalities in cerebral areas that have been associated with schizophrenia (Straub et al., 2002). Furthermore, presynaptic dysbindin involvement was observed in a study by Numakawa et al. (2004), who suggested that dysbindin 1 regulation of the expression of SNAP25 and synapsin 1 proteins in the presynaptic machinery associated with increased glutamate release. Further, a selective reduction in presynaptic dysbindin 1 expression as well as an inverse correlation of dysbindin 1 levels with VGlut-T-1, the primary vesicular glutamate transporter in the hippocampal formation, has been reported from Talbot et al. (2004). To date, numerous studies have reported decreased dysbindin 1 expression in schizophrenia-related brain regions, such as the hippocampus, dorsolateral prefrontal cortex (DLPFC) and midbrain, amongst other regions (Weickert et al., 2004, Numakawa et al., 2004, Talbot et al., 2004, Coyle, 2006, Weickert et al., 2008). Finally, Harrison and Weinberger (2005) proposed that glutamatergic synaptic alterations and aberrant neural circuitry are considered to be crucial for functional abnormalities in schizophrenia. However, the specific role of genetic variation of dysbindin 1 for malfunctioning glutamatergic neuronal system is still unclear.
Whilst a series of linkage and association studies have implicated the dysbindin 1 SNP rs1018381 with genetic susceptibility to schizophrenia (Straub et al., 2002, Schwab et al., 2003, Tang et al., 2003, Van Den Bogaert et al., 2003, Funke et al., 2004), there is also evidence for no significant results to this marker (Stefanis et al., 2007, Tosato et al., 2007). Although the allele frequencies of SNP rs1018381 differ between HapMap populations (see also Genetic analysis part), associations with the minor T-allele of this SNP were found in a study by Funke et al. (2004) in three independent samples of different ethnic origin (White, Hispanic and African American). In their study the SNP rs1018381 was conceived as a tagging SNP for the risk six-locus haplotype (CTCTAC) that was significantly over-represented in the Caucasian patients when compared with Caucasian controls (Funke et al., 2004). It was the only SNP of this risk-haplotype that showed a significant effect of genotype on cognitive abilities in patients with schizophrenia and healthy volunteers (Burdick et al., 2006). Minor T-allele carriers performed significantly worse on diverse cognitive tests than non-carriers.
Nonetheless, this SNP tags a large number of SNPs, and it is not clear which of these SNPs is responsible for the observed effect (see also Genetic analysis part). Using the web tool Haplotter (http://pritch.bsd.uchicago.edu/data.html) to investigate any marker of DTNBP1 in HapMap in the CEU population (for a method of DTNBP1 selection in the HapMap see Voight et al., 2006), it could be shown that the SNP rs1018381 is in perfect LD (r2 = 1.0) with the majority of SNPs (rs11755055, rs9296984, rs2619535, rs2743550, and rs9476860) of top scoring markers in DTNBP1. However, in a single SNP query, the integrated haplotype score (iHS) for marker rs1018381 itself (standardized iHS = − 1.957) just failed to approach the positive/negative iHS scores of > 2/< − 2, which were used as a threshold to identify the strongest signals of selection in the study of Voight et al. (2006). Thus, the assumed SNP cannot serve as functional but its role as a significant marker in DTNBP1 remains crucial.
A series of current studies supported the influence of this and other SNPs of this risk-haplotype on cognitive abilities in healthy individuals. Luciano et al. (2009) showed significant associations between the SNP rs1018381 and verbal ability, the SNPs rs2619528 and rs1011313 and executive function, and the SNP rs2619522 and memory, speed and executive function in two cohorts. Furthermore, the SNPs rs2619522 and rs760761 were associated with lower sustained attention in healthy volunteers (Stefanis et al., 2007). Finally, only the SNP rs1018381 implemented in the risk six-locus haplotype (CTCTAC) reported by Burdick et al., 2006, Funke et al., 2004 seemed to be characteristic for cognitive deficits in patients with schizophrenia and healthy individuals.
Deficient language production is one of the core symptoms in patients with schizophrenia. Results from meta-analyses (DeLisi, 2001, Sitskoorn et al., 2004, Szöke et al., 2004, Snitz et al., 2006) have provided evidence that this deficiency has a strong genetic component. Verbal fluency is a widely employed measure to assess alteration in language production in schizophrenia patients (for a review see Bokat and Goldberg, 2003). Two meta-analyses have shown that patients with schizophrenia were more impaired on semantic (the word generation is constrained by a specified category, e.g. “tools”) relative to lexical (the participants are asked to generate as many words as possible beginning with a specified letter, e.g. “S”) fluency tasks, which has been taken as evidence for a selective semantic deficit (Bokat and Goldberg, 2003, Henry and Crawford, 2005). Further, Chen et al. (2000) have found that semantic verbal fluency abnormality can serve as a familial trait marker for schizophrenia.
In healthy populations, cerebral correlates of semantic verbal fluency comprise a left-lateralized fronto-temporal network (for a review on word production see Indefrey and Levelt, 2004). Due to the demand on organizing and controlling the semantic search, validation and retrieval processes, the most consistent activations have been found in the lateral and medial prefrontal cortex including the inferior frontal gyrus and anterior cingulate cortex, complimented by lateral and polar temporal areas, the thalamus or hippocampal region (Markowitsch, 1995, Cabeza and Nyberg, 2000, Crespo-Facorro et al., 2004, Demonet et al., 2005, Basho et al., 2007, Kircher et al., 2008, Ragland et al., 2008, Whitney et al., 2009). Functional imaging studies of verbal fluency in patients with schizophrenia report aberrant activation patterns in prefrontal (Yurgelun-Todd et al., 1996, Boksman et al., 2005, Schaufelberger et al., 2005), anterior cingulate (Fletcher et al., 1996) and temporal structures (Yurgelun-Todd et al., 1996, Kuperberg et al., 2007). Such neural abnormalities have been interpreted within the fronto-temporal dysconnectivity hypothesis (Friston and Frith, 1995). Further, when patients with schizophrenia are compared with healthy controls during semantic verbal fluency tasks, stronger activation has been found in the left prefrontal and motor cortex, right anterior cingulate gyrus, bilateral insula, right putamen, bilateral middle and STG, left inferior and superior parietal lobule, left lingual gyrus, left cuneus and precuneus, and cerebellum (Ragland et al., 2008).
The aim of the current study was to investigate the influence of the SNP rs1018381 of the DTNBP1 gene on neural correlates of verbal fluency processes in healthy volunteers. Consistent with the findings in the literature (Markowitsch, 1995, Cabeza and Nyberg, 2000, Crespo-Facorro et al., 2004, Demonet et al., 2005, Basho et al., 2007, Kircher et al., 2008, Ragland et al., 2008, Whitney et al., 2009), differential activation for semantic but not for lexical verbal fluency in the lateral and medial prefrontal cortex, right anterior cingulate gyrus, bilateral insula, bilateral middle and STG, left inferior and superior parietal lobule, and cerebellum was expected. Stronger brain activations in some or all of these cerebral regions in risk-allele carriers compared with non-carriers and correlations between them and verbal fluency task performance were hypothesized.
Section snippets
Participants
A sample of 521 subjects (268 men, 253 women) was recruited from students of the RWTH Aachen University. Advertisements were made by word of mouth, posted flyers in institutes and canteens, and an e-mail was sent to all students of the University. All subjects gave written informed consent, and the study was approved by the local Ethics Committee according to the declaration of Helsinki. All 521 subjects were genotyped for SNP rs1018381 carrier status and classified into groups of heterozygous
fMRI behavioural data
All 46 participants followed the instructions and completed both tasks in a satisfactory manner. Mean number of errors was 3.96 (SD = 3.28) for non-carriers (n = 26) and 2.9 (SD = 3.87) for risk-allele carriers (n = 20). No difference in the number of errors was evident between the groups (two-sample t-test, F = 1.01, df = 44, p = .50).
fMRI data
The main contrast (n = 46) “semantic verbal fluency > baseline condition (reading aloud)” revealed activations in bilateral caudate nuclei, the left parahippocampal gyrus (BA 19),
Discussion
The current study demonstrates that genetic variation in schizophrenia-risk DTNBP1 gene has an influence on brain activity in regions that are related to semantic word retrieval processes. During semantic verbal fluency, the group of risk-allele carriers revealed stronger activation in the rostral part of the right ACC (BA 24), right STG (BA 22, 38), and right MTG (BA 21) than the group of non-carriers, in the absence of performance differences.
Stronger signal changes in the group of
Conclusion
Our results provide evidence for an influence of genetic variation of the schizophrenia-risk DTNBP1 gene onto brain activation in the right rostral anterior cingulate and right lateral temporal cortex during semantic verbal fluency. These regions have previously been related to semantic verbal fluency processes and are further core neuropathological areas in schizophrenia. This alteration of brain activity in risk-allele carriers occurred in the absence of performance differences between
Acknowledgments
This work was supported by grants from the Federal Ministry of Education and Research (Brain Imaging Center West, 01GO0204). This work is a part of the thesis of Mr. Markov. We thank Anna Weiss, Jane Horn, Petra Engels, Veronika Ermer and Dorothe Krug for their assistance and support as well as all the participants of this study.
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