A rare functional noncoding variant at the GWAS-implicated MIR137/MIR2682 locus might confer risk to schizophrenia and bipolar disorder.

Schizophrenia (SZ) genome-wide association studies (GWASs) have identified common risk variants in >100 susceptibility loci; however, the contribution of rare variants at these loci remains largely unexplored. One of the strongly associated loci spans MIR137 (miR137) and MIR2682 (miR2682), two microRNA genes important for neuronal function. We sequenced ∼6.9 kb MIR137/MIR2682 and upstream regulatory sequences in 2,610 SZ cases and 2,611 controls of European ancestry. We identified 133 rare variants with minor allele frequency (MAF) <0.5%. The rare variant burden in promoters and enhancers, but not insulators, was associated with SZ (p = 0.021 for MAF < 0.5%, p = 0.003 for MAF < 0.1%). A rare enhancer SNP, 1:g.98515539A>T, presented exclusively in 11 SZ cases (nominal p = 4.8 × 10(-4)). We further identified its risk allele T in 2 of 2,434 additional SZ cases, 11 of 4,339 bipolar (BP) cases, and 3 of 3,572 SZ/BP study controls and 1,688 population controls; yielding combined p values of 0.0007, 0.0013, and 0.0001 for SZ, BP, and SZ/BP, respectively. The risk allele T of 1:g.98515539A>T reduced enhancer activity of its flanking sequence by >50% in human neuroblastoma cells, predicting lower expression of MIR137/MIR2682. Both empirical and computational analyses showed weaker transcription factor (YY1) binding by the risk allele. Chromatin conformation capture (3C) assay further indicated that 1:g.98515539A>T influenced MIR137/MIR2682, but not the nearby DPYD or LOC729987. Our results suggest that rare noncoding risk variants are associated with SZ and BP at MIR137/MIR2682 locus, with risk alleles decreasing MIR137/MIR2682 expression.

Genetic variants in or near ADH1B and ADH1C affect susceptibility to alcohol dependence in a British and Irish population.

Certain single nucleotide polymorphisms (SNPs) in genes encoding alcohol dehydrogenase (ADH) enzymes confer a significant protective effect against alcohol dependence syndrome (ADS) in East Asian populations. Recently, attention has focused on the role of these SNPs in determining ADS risk in European populations. To further elucidate these associations, SNPs of interest in ADH1B, ADH1C and the ADH1B/1C intergenic region were genotyped in a British and Irish population (ADS cases n = 1076: controls n = 1027) to assess their relative contribution to ADS risk. A highly significant, protective association was observed between the minor allele of rs1229984 in ADH1B and ADS risk [allelic P = 8.4 × 10(-6) , odds ratio (OR) = 0.26, 95 percent confidence interval, 0.14, 0.49]. Significant associations were also observed between ADS risk and the ADH1B/1C intergenic variant, rs1789891 [allelic P = 7.2 × 10(-5) , OR = 1.4 (1.2, 1.6)] and three non-synonymous SNPs rs698, rs1693482 and rs283413 in ADH1C. However, these associations were not completely independent; thus, while the ADH1B rs1229984 minor allele association was independent of those of the intergenic variant rs1789891 and the three ADH1C variants, the three ADH1C variants were not individually independent. In conclusion, the rare ADH1B rs1229984 mutation provides significant protection against ADS in this British and Irish population; other variants in the ADH gene cluster also alter ADS risk, although the strong linkage disequilibrium between SNPs at this location precluded clear identification of the variant(s) driving the associations.

Genome-wide association study of bipolar disorder in Canadian and UK populations corroborates disease loci including SYNE1 and CSMD1.

BACKGROUND: Recently, genome-wide association studies (GWAS) for cases versus controls using single nucleotide polymorphism microarray data have shown promising findings for complex neuropsychiatric disorders, including bipolar disorder (BD). METHODS: Here we describe a comprehensive genome-wide study of bipolar disorder (BD), cross-referencing analysis from a family-based study of 229 small families with association analysis from over 950 cases and 950 ethnicity-matched controls from the UK and Canada. Further, loci identified in these analyses were supported by pathways identified through pathway analysis on the samples. RESULTS: Although no genome-wide significant markers were identified, the combined GWAS findings have pointed to several genes of interest that support GWAS findings for BD from other groups or consortia, such as at SYNE1 on 6q25, PPP2R2C on 4p16.1, ZNF659 on 3p24.3, CNTNAP5 (2q14.3), and CDH13 (16q23.3). This apparent corroboration across multiple sites gives much confidence to the likelihood of genetic involvement in BD at these loci. In particular, our two-stage strategy found association in both our combined case/control analysis and the family-based analysis on 1q21.2 (closest gene: sphingosine-1-phosphate receptor 1 gene, S1PR1) and on 1q24.1 near the gene TMCO1, and at CSMD1 on 8p23.2, supporting several previous GWAS reports for BD and for schizophrenia. Pathway analysis suggests association of pathways involved in calcium signalling, neuropathic pain signalling, CREB signalling in neurons, glutamate receptor signalling and axonal guidance signalling. CONCLUSIONS: The findings presented here show support for a number of genes previously implicated genes in the etiology of BD, including CSMD1 and SYNE1, as well as evidence for previously unreported genes such as the brain-expressed genes ADCY2, NCALD, WDR60, SCN7A and SPAG16.

Genetic association of the tachykinin receptor 1 TACR1 gene in bipolar disorder, attention deficit hyperactivity disorder, and the alcohol dependence syndrome.

Single nucleotide polymorphisms (SNPs) in the tachykinin receptor 1 gene (TACR1) are nominally associated with bipolar affective disorder (BPAD) in a genome-wide association study and in several case-control samples of BPAD, alcohol dependence syndrome (ADS) and attention-deficit hyperactivity disorder (ADHD). Eighteen TACR1 SNPs were associated with BPAD in a sample (506 subjects) from University College London (UCL1), the most significant being rs3771829, previously associated with ADHD. To further elucidate the role of TACR1 in affective disorders, rs3771829 was genotyped in a second BPAD sample of 593 subjects (UCL2), in 997 subjects with ADS, and a subsample of 143 individuals diagnosed with BPAD and comorbid alcohol dependence (BPALC). rs3771829 was associated with BPAD (UCL1 and UCL2 combined: P = 2.0 × 10(-3)), ADS (P = 2.0 × 10(-3)) and BPALC (P = 6.0 × 10(-4)) compared with controls screened for the absence of mental illness and alcohol dependence. DNA sequencing in selected cases of BPAD and ADHD who had inherited TACR1-susceptibility haplotypes identified 19 SNPs in the promoter region, 5' UTR, exons, intron/exon junctions and 3' UTR of TACR1 that could increase vulnerability to BPAD, ADS, ADHD, and BPALC. Alternative splicing of TACR1 excludes intron 4 and exon 5, giving rise to two variants of the neurokinin 1 receptor (NK1R) that differ in binding affinity of substance P by 10-fold. A mutation in intron four, rs1106854, was associated with BPAD, although a regulatory role for rs1106854 is unclear. The association with TACR1 and BPAD, ADS, and ADHD suggests a shared molecular pathophysiology between these affective disorders.

Genetic association and sequencing of the insulin-like growth factor 1 gene in bipolar affective disorder.

Insulin-like growth factor 1 (IGF1) has been shown to have an important role in brain development and function. Studies of IGF1 administration in rodents have shown that it has an anxiolytic and antidepressant effect. A genome-wide association study (GWAS) of the first University College London (UCL) cohort of 506 bipolar affective disorder subjects and 510 controls was carried out. The exons and flanking regions of IGF1 were resequenced, any new polymorphisms found were genotyped in an enlarged UCL sample of 937 cases and 941 controls. GWAS data gave good evidence of allelic and haplotypic association between multiple IGF1 SNP's and bipolar disorder (BD). New polymorphisms were found by resequencing IGF1 region. Data from GWAS and the new markers showed that twelve out of 43 SNPs showed association with BD with the four most significant SNPs having values of 3.7 × 10(-5) , 8.4 × 10(-4) , 2.6 × 10(-4) , and 2.5 × 10(-4) . A 5' promoter microsatellite polymorphism previously correlated with plasma lipoprotein concentration was also associated with BD (P = 0.013). Haplotypic association confirmed association with BD with significance values similar to the single marker SNP values. The marker rs12426318 has also been found to be associated with BD in a second sample. A test of gene wide significance with permutation testing for all markers genotyped at IGF1 was also significant. These data implicate IGF1 as a candidate gene to cause genetic susceptibility to BD.

Three-dimensional textural analysis of brain images reveals distributed grey-matter abnormalities in schizophrenia.

OBJECTIVES: Three-dimensional (3-D) selective- and relative-scale texture analysis (TA) was applied to structural magnetic resonance (MR) brain images to quantify the presence of grey-matter (GM) and white-matter (WM) textural abnormalities associated with schizophrenia. MATERIALS AND METHODS: Brain TA comprised volume filtration using the Laplacian of Gaussian filter to highlight fine, medium and coarse textures within GM and WM, followed by texture quantification. Relative TA (e.g. ratio of fine to medium) was also computed. T1-weighted MR whole-brain images from 32 participants with diagnosis of schizophrenia (n = 10) and healthy controls (n = 22) were examined. Five patients possessed marker alleles (SZ8) associated with schizophrenia on chromosome 8 in the pericentriolar material 1 gene while the remaining five had not inherited any of the alleles (SZ0). RESULTS: Filtered fine GM texture (mean grey-level intensity; MGI) most significantly differentiated schizophrenic patients from controls (P = 0.0058; area under the receiver-operating characteristic curve = 0.809, sensitivity = 90%, specificity = 70%). WM measurements did not distinguish the two groups. Filtered GM and WM textures (MGI) correlated with total GM and WM volume respectively. Medium-to-coarse GM entropy distinguished SZ0 from controls (P = 0.0069) while measures from SZ8 were intermediate between the two. CONCLUSIONS: 3-D TA of brain MR enables detection of subtle distributed morphological features associated with schizophrenia, determined partly by susceptibility genes.

Support of association between BRD1 and both schizophrenia and bipolar affective disorder.

A recent study published by our group implicated the bromodomain containing protein 1 (BRD1) gene located at chromosome 22q13.33 with schizophrenia (SZ) and bipolar affective disorder (BPD) susceptibility and provided evidence suggesting a possible role for BRD1 in neurodevelopment. The present study reports an association analysis of BRD1 and the neighboring gene ZBED4 using a Caucasian case-control sample from Denmark and England (UK/DK sample: 490 patients with BPD, 527 patients with SZ, and 601 control individuals), and genotypes obtained from a BPD genome wide association (GWA) study of an overlapping English sample comprising 506 patients with BPD and 510 control individuals (UCL sample). In the UK/DK sample we genotyped 11 SNPs in the BRD1 region, of which six showed association with SZ (minimal single marker P-values of 0.0014), including two SNPs that previously showed association in a Scottish population [Severinsen et al. (2006); Mol Psychiatry 11(12): 1126-1138]. Haplotype analysis revealed specific risk as well as protective haplotypes with a minimal P-value of 0.0027. None of the 11 SNPs showed association with BPD. However, analyzing seven BRD1 SNPs obtained from the BPD GWA study, positive associations with BPD was observed with all markers (minimal P-value of 0.0014). The associations reported add further support for the implication of BRD1 with SZ and BPD susceptibility.

Genetic association and brain morphology studies and the chromosome 8p22 pericentriolar material 1 (PCM1) gene in susceptibility to schizophrenia.

CONTEXT: There is evidence of linkage to a schizophrenia susceptibility locus on chromosome 8p21-22 found by several family linkage studies. OBJECTIVES: To fine map and identify a susceptibility gene for schizophrenia on chromosome 8p22 and to investigate the effect of this genetic susceptibility on an endophenotype of abnormal brain structure using magnetic resonance imaging. DESIGN: Fine mapping and identification of a chromosome 8p22 susceptibility gene was carried out by finding linkage disequilibrium between genetic markers and schizophrenia in multiply affected families, a case-control sample, and a trio sample. Variation in brain morphology associated with pericentriolar material 1 (PCM1) alleles was examined using voxel-based morphometry and statistical parametric mapping with magnetic resonance imaging. Setting and Patients A family sample of 13 large families multiply affected with schizophrenia, 2 schizophrenia case-control samples from the United Kingdom and Scotland, and a sample of schizophrenic trios from the United States containing parents and 1 affected child with schizophrenia. MAIN OUTCOME MEASURES: Tests of transmission disequilibrium between PCM1 locus polymorphisms and schizophrenia using a family sample and tests of allelic association in case-control and trio samples. Voxel-based morphometry using statistical parametric mapping. RESULTS: The family and trio samples both showed significant transmission disequilibrium between marker D85261 in the PCM1 gene locus and schizophrenia. The case-control sample from the United Kingdom also found significant allelic association between PCM1 gene markers and schizophrenia. Voxel-based morphometry of cases who had inherited a PCM1 genetic susceptibility showed a significant relative reduction in the volume of orbitofrontal cortex gray matter in comparison with patients with non-PCM1-associated schizophrenia, who, by contrast, showed gray matter volume reduction in the temporal pole, hippocampus, and inferior temporal cortex. CONCLUSIONS: The PCM1 gene is implicated in susceptibility to schizophrenia and is associated with orbitofrontal gray matter volumetric deficits.

Failure to confirm allelic association between markers at the CAPON gene locus and schizophrenia in a British sample.

BACKGROUND: Linkage studies have confirmed that chromosome 1q23.3 is a susceptibility locus for schizophrenia. It was then claimed that markers at the carboxyl-terminal PDZ ligand of neuronal nitric oxide synthase (CAPON) gene showed allelic association with schizophrenia in Canadian families. A second Chinese study found a base pair polymorphism at the CAPON gene also associated with schizophrenia. METHODS: We attempted replication using eight markers from the Canadian study in a UK based sample of 450 cases and 450 supernormal controls. RESULTS: We found no evidence for allelic or haplotypic association with schizophrenia for any of the markers found to be associated in the Canadian sample. CONCLUSIONS: The negative results might reflect genetic heterogeneity between the Canadian, Chinese and UK samples or be due to methodological problems. The present finding weakens the evidence that mutations or variation in the CAPON gene are causing genetic susceptibility to schizophrenia in European populations.

Genetic linkage analysis of the X chromosome in autism, with emphasis on the fragile X region.

The higher prevalence of autism in males than in females suggests the possible involvement of the X chromosome. To test the hypothesis that there are mutations increasing susceptibility to autism on the X chromosome, and in particular the distal portion of the long arm that encompasses the FMRI and MECP2 loci, a genetic linkage study was performed. Twenty-two fragile X-negative families multiplex for autism and related disorders were used for the study. Linkage analysis, for markers in the Xq27-q28 region, using model-free likelihood-based analysis, produced a maximum MLOD of 1.7 for the narrowest diagnostic category of the typical autism/severe autism spectrum, and nonparametric analysis produced a maximum non-parametric lod (NPL) score of 2.1 for a broad phenotype diagnostic model. Thus, this study offers modest support for a susceptibility locus for autism within the Xq27-q28 region. Further genetic investigations of this region are warranted.

Genetic linkage analysis supports the presence of two susceptibility loci for alcoholism and heavy drinking on chromosome 1p22.1-11.2 and 1q21.3-24.2.

BACKGROUND: In order to confirm a previous finding of linkage to alcoholism on chromosome 1 we have carried out a genetic linkage study. METHODS: DNA from eighteen families, densely affected by alcoholism, was used to genotype a set of polymorphic microsatellite markers at loci approximately 10 centimorgans apart spanning the short arm and part of the long arm of chromosome 1. Linkage analyses were performed using the classical lod score and a model-free method. Three different definitions of affection status were defined, these were 1. Heavy Drinking (HD) where affected subjects drank more than the Royal College of Psychiatrists recommended weekly amount. 2. The Research Diagnostic Criteria for alcoholism (RDCA) 3. Alcohol Dependence Syndrome (ADS) as defined by Edwards and Gross (1976) and now incorporated into ICD10 and DSMIV. RESULTS: Linkage analyses with the markers D1S1588, D1S2134, D1S1675 covering the cytogenetic region 1p22.1-11.2 all gave positive two point and multipoint lods with a maximum lod of 1.8 at D1S1588 (1p22.1) for the RDCA definition of alcoholism. Another lod of 1.8 was found with D1S1653 in the region 1q21.3-24.2 using the HD affection model. CONCLUSION: These results both support the presence of linkage in the 1p22.1-11.2 region which was previously implicated by the USA Collaborative Study of the Genetics of Alcoholism (COGA) study and also suggest the presence of another susceptibility locus at 1q21.3-24.2.

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

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 ß-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.

Genetic association, mutation screening, and functional analysis of a Kozak sequence variant in the metabotropic glutamate receptor 3 gene in bipolar disorder.

IMPORTANCE: Genetic markers at the gene encoding the metabotropic glutamate receptor 3 (GRM3) showed allelic association with bipolar disorder. OBJECTIVE: To screen the GRM3 gene and adjacent control regions of genomic DNA in volunteers with bipolar affective disorder for mutations increasing susceptibility to bipolar disorder. DESIGN: Sequencing and high-resolution melting curve analysis of DNA followed by genotyping was carried out in 1099 patients with bipolar affective disorder and 1152 healthy comparator individuals. SETTING: Participants with bipolar disorder were recruited from National Health Service psychiatric services and from patient organizations. PARTICIPANTS: Individuals were included if they had Research Diagnostic Criteria diagnoses of bipolar I and bipolar II disorder and were of British or Irish ancestry. MAIN OUTCOMES AND MEASURES: Identification of base pair changes in the GRM3 gene that affected expression or function of the GRM3 receptor that also showed an allelic association with bipolar disorder. RESULTS: A base pair variant (rs148754219) was found in the Kozak sequence of exon 1 of the GRM3 gene, 2 bases before the translation start codon of one of the receptor isoforms, in 23 of 2251 people who were screened and genotyped. Nineteen of the 1099 bipolar cases (1.7%) were mutation carriers compared with 4 of 1152 healthy comparators (0.3%). The variant was associated with bipolar disorder (P = .005; odds ratio, 4.20). Bioinformatic, electrophoretic mobility shift assay, and gene expression analysis found that the variant created a new transcription factor protein binding site and had a strong effect on gene transcription and translation. CONCLUSIONS AND RELEVANCE: Confirmation of these findings is needed before the Kozak sequence variant can be accepted as a potential marker for personalized treatment of affective disorders with drugs targeting the metabotropic glutamate receptor 3.

Mutations in the Gabrb1 gene promote alcohol consumption through increased tonic inhibition.

Alcohol dependence is a common, complex and debilitating disorder with genetic and environmental influences. Here we show that alcohol consumption increases following mutations to the γ-aminobutyric acidA receptor (GABAAR) ß1 subunit gene (Gabrb1). Using N-ethyl-N-nitrosourea mutagenesis on an alcohol-averse background (F1 BALB/cAnN x C3H/HeH), we develop a mouse model exhibiting strong heritable preference for ethanol resulting from a dominant mutation (L285R) in Gabrb1. The mutation causes spontaneous GABA ion channel opening and increases GABA sensitivity of recombinant GABAARs, coupled to increased tonic currents in the nucleus accumbens, a region long-associated with alcohol reward. Mutant mice work harder to obtain ethanol, and are more sensitive to alcohol intoxication. Another spontaneous mutation (P228H) in Gabrb1 also causes high ethanol consumption accompanied by spontaneous GABA ion channel opening and increased accumbal tonic current. Our results provide a new and important link between GABAAR function and increased alcohol consumption that could underlie some forms of alcohol abuse.

Case-case genome-wide association analysis shows markers differentially associated with schizophrenia and bipolar disorder and implicates calcium channel genes.

OBJECTIVE: There are theoretical reasons why comparing marker allele frequencies between cases of different diseases, rather than with controls, may offer benefits. The samples may be better matched, especially for background risk factors common to both diseases. Genetic loci may also be detected which influence which of the two diseases occurs if common risk factors are present. METHOD: We used samples of UK bipolar and schizophrenic cases that had earlier been subject to genome-wide association studies and compared marker allele frequencies between the two samples. When these differed for a marker, we compared the case sample allele frequencies with those of a control sample. RESULTS: Eight markers were significant at P value of less than 10(-5). Of these, the most interesting finding was for rs17645023, which was significant at P value of less than 10(-6) and which lies 36 kb from CACNG5. Control allele frequencies for this marker were intermediate between those for bipolar and schizophrenic cases. CONCLUSION: The application of this approach suggests that it does have some merits. The finding for CACNG5, taken together with the earlier implication of CACNA1C and CACNA1B, strongly suggests a key role for voltage-dependent calcium channel genes in the susceptibility to bipolar disorder and/or schizophrenia.

Genetics of attention-deficit hyperactivity disorder (ADHD).

Attention-deficit hyperactivity disorder (ADHD) is a clinically and genetically heterogeneous syndrome which is comorbid with childhood conduct disorder, alcoholism, substance abuse, dis-social personality disorder, and affective disorders. A small but consistent overlap with autistic symptoms has also been established. Twin and family studies of ADHD show a substantial genetic heritability with little or no family environmental effect. Linkage and association studies have conclusively implicated the dopamine transporter gene (DAT1). DAT1 has also been confirmed as being associated with bipolar disorder. Remarkably, and for the first time in psychiatry, genetic markers at the DAT1 locus appear to be able to predict clinical heterogeneity because the non-conduct disordered subgroup of ADHD is associated with DAT1 whereas other subgroups do not appear to be associated. The second most well replicated susceptibility gene encodes the DRD4 dopamine receptor and many other dopamine related genes appear to be implicated. It is becoming increasingly clear that genes causing bipolar mania overlap with genes for a subtype of ADHD. The key to understanding the genetics of ADHD is to accept very considerable heterogeneity with different genes having effects in different families and in different individuals. It is too early to interpret the new wave of genome-wide association and copy number variant studies but preliminary data support the overlap with affective disorder genes and also with CNS connectivity genes likely to be involved in autism and affective disorders.

No evidence for excess runs of homozygosity in bipolar disorder.

BACKGROUND: Recent studies have reported large common regions of homozygosity (ROHs) that are the result of autozygosity, that is, the cooccurrence within individuals of long haplotypes that have a high frequency in the population. A recent study reports that such regions are found more commonly in individuals with schizophrenia compared with controls, and identified nine 'risk ROHs' that were individually more common in cases. Of these, four contained or neighboured genes associated with schizophrenia (NOS1AP/UHMK1, ATF2, NSF and PIK3C3). METHODS: We have applied the same methodology to a UK sample of 506 cases with bipolar disorder and 510 controls. RESULTS: There was no overall excess of common ROHs among bipolar individuals. With one exception, the haplotypes accounting for the ROHs appeared to be distributed according to the Hardy-Weinberg equilibrium. One ROH was individually more common among cases (uncorrected P = 0.0003). This ROH spanned the chromosome 2p23.3 gene ITSN2 (the gene for intersectin 2 isoform 2). However, inspection of the homozygous haplotypes and haplotype-based tests for association failed to provide a clearer understanding of why this ROH was occurring more commonly. CONCLUSION: Overall, we conclude that, in contrast with schizophrenia, common ROHs are rarely associated with susceptibility to bipolar disorder. This supports the idea that predominantly different genes are increasing susceptibility to schizophrenia and bipolar affective disorders.

A microarray gene expression study of the molecular pharmacology of lithium carbonate on mouse brain mRNA to understand the neurobiology of mood stabilization and treatment of bipolar affective disorder.

OBJECTIVES: Lithium is the most widely prescribed and effective mood-stabilizing drug used for the treatment of bipolar affective disorder. To understand how lithium produces changes in the brain, we studied brain mRNA from 10 mice after treatment with lithium and compared them with 10 untreated controls. METHODS: We used the MAS 5.0, Smudge miner, GC-RMA and FDR-AME packages of software (Bioconductor, Seattle, Washington, USA) to determine gene expression changes using Affymetrix MOE430E 2.0 microarrays after 2 weeks of lithium treatment. RESULTS: We used both a false discovery rate (FDR-AME) assessment of significance and the Bonferroni method to correct for the possibility of false-positive changes in gene expression among the 39,000 genes present in each array. Our primary method of analysis was to use t-tests on normalized gene expression intensities. By using a Bonferroni correction of P<1.28x10(-6), we found that 121 genes showed significant changes in expression. The three genes with the most changed mRNA expression were alanine-glyoxylate aminotransferase 2-like 1 (Agxt2l1), c-mer proto-oncogene tyrosine kinase (Mertk) and sulfotransferase family 1A phenol-preferring member 1 (Sult1a1). Also among the group of 121 genes with significant changes in gene expression that survived Bonferroni correction () were the genes encoding the Per2 period gene (Per2 P=1.33x10(-8), 2.47-fold change), the metabotropic glutamate receptor (Grm3, P=9.48x10(-7), 0.7-fold change) and secretogranin II (Scg2, P=9.48x10(-7), 1.28-fold change) as well as several myelin-related genes and protein phosphatases. By taking a significance value of P<0.05 without Bonferroni or FDR-AME correction, we identified a total of 4474 genes showing changed mRNA expression in response to lithium. FDR-AME analysis showed that 1027 out of these 4474 genes were significantly changed in expression. Among the mRNAs that were significantly changed with t-tests and FDR-AME were several that had already been implicated in response to lithium such as increased brain-derived neurotrophic factor mRNA ( t-test P=0.0008-0.0005, FDR-AME P=0.0396-0.0393, 1.44-fold change) beta-phosphatidylinositol transfer protein (Pitpnb, t-test P<0.0000, FDR-AME P=0.003, 1.26-fold change) and inositol (myo)-1(or 4)-monophosphatase 1(Impa1, t test P<0.0000, FDR-AME P=0.004, 1.22-fold change). Of interest in relation to the side effect of hypothyroidism, which is caused by long-term lithium treatment was the fact that we observed changes in mRNA expression in five genes related to thyroxine metabolism. These included deiodinase (Dio2 t-test P=0.000003-0.004, FDR-AME P=0.0048-0.061, 1.53-fold change) and thyroid hormone receptor interactor 12 (Trip12, t-test P=0.003, FDR-AME P=0.075, 1.19-fold change). Of relevance to multiple sclerosis was the observed upregulation of the long isoform of myelin basic protein (t-test P=0.00013, FDR-AME P=0.0169). Changes in mRNA expression were found in 45 genes related to phosphatidylinositol metabolism using uncorrected t-tests but only 13 genes after FDR-AME. Thus, our work confirms the considerable previous research implicating this system. Gene ontology analysis showed that lithium significantly affected a cluster of processes associated with nucleotide and nucleoside metabolism. The analysis showed that there were 170 genes expressing RNA described as having ATP-binding or ATPase activity that had changed mRNA expression. The changes found have been discussed in relation to previous experimental work on the pharmacology of lithium.