Screening of chromosomal region 21q22.3 for mutations in genes associated with neuronal Ca2+ signalling in bipolar affective disorder

The therapeutic effect of lithium in bipolar affective disorder may be connected with decreasing intracellular Ca(2+) concentrations. Several linkage studies have identified a potential bipolar affective disorder susceptibility locus within chromosomal region 21q22.3. This locus contains two genes expressed in the brain - ADARB1 and TRPM2 - involved in regulating intracellular Ca(2+) concentrations. The aim of this study was an identification of mutations in the coding sequences of ADARB1 and TRPM2 and their association with bipolar affective disorder. For that purpose we screened 60 patients with bipolar affective disorder and a control group of 66 subjects using single strand conformation polymorphism and sequence analysis. For rapid screening we performed restriction fragment length polymorphism analysis. Screening of bipolar affective disorder patients for mutations in TRPM2 led to identification of three novel and four known transitions. Two transitions resulted in the substitutions: R755C and A890V. Screening of the coding sequence of ADARB1 did not reveal any mutations except one already known transition. A comparison of the transition frequency in patients and controls does not support association of the detected mutations with bipolar affective disorder. According to our results, bipolar affective disorder may not be caused by mutations in ADARB1. However, this study does not exclude TRPM2 as a candidate gene since we have screened only about 30 per cent of the entire coding sequence of this large gene.     

Association of the HTR2A gene with alcohol and heroin abuse

Positive genetic associations of rs6313 (102T/C at exon 1) and rs6311 (?1438A/G) on the 5-hydroxytryptamine (serotonin) 2A receptor gene (HTR2A or 5-HT2A) were reported for alcohol and drug abuse; however, other association studies failed to produce consistent results supporting the susceptibility of the two single nucleotide polymorphisms (SNPs). To clarify the associations of the HTR2A gene with substance use disorders, we performed a meta-analysis based on the genotypes from the available candidate gene association studies of the two SNPs with alcohol and drug abuse from multiple populations. Evidence of association was found for HTR2A rs6313 in all the combined studies (e.g., allelic P = 0.0048 and OR 0.86, 95 % CI 0.77–0.95) and also in the combined studies of alcohol dependence (abuse) (e.g., allelic P = 0.0001 and OR 0.71, 95 % CI 0.59–0.85). The same association trend was also observed in the Study of Addiction: Genetics and Environment datasets. The meta-analysis supports a contribution of the HTR2A gene to the susceptibility to substance use disorders, particularly alcohol dependence.     

Psychosis and genes with trinucleotide repeat polymorphism

Abnormal expansion of genes with trinucleotide repeat (TNR) polymorphism has been found in a number of neuropsychiatric disorders. These disorders and the major psychoses, schizophrenia and bipolar affective disorder, appear to share an interesting phenomenon: genetic anticipation. Because TNR expansion correlates with anticipation, these unstable DNA sites are considered important candidate loci for the major psychoses. We investigated genes with TNR polymorphisms, includingB1, B33, B37, and theN-cadherin gene, in unrelated Caucasian North American and Italian schizophrenics (n = 53 to 74), and matched controls. Also, unrelated Caucasian North American patients with bipolar I affective disorder were screened for the B33 andN-cadherin genes (n = 49 and 63, respectively). No unusually long alleles that would suggest abnormal expansion of the TNR were observed for any of these genes. Also, no statistically significant results were found in tests for genetic association between any of these genes and schizophrenia. For B37, a trend toward a difference in allele counts between schizophrenics and controls was observed. However, no clear evidence for a role of these TNR-containing genes in schizophrenia or bipolar affective disorders was found.     

Investigation of 17 candidate genes for personality traits confirms effects of the HTR2A gene on novelty seeking

Genes involved in serotonergic and dopaminergic neurotransmission have been hypothesized to affect different aspects of personality, but findings from genetic association studies did not provide conclusive results so far. In previous studies, however, only one or a few polymorphisms within single genes were investigated neglecting the possibility that the genetic associations might be more complex comprising several genes or gene regions. To overcome this limitation, we performed an extended genetic association study analyzing 17 serotonergic ( SLC6A4, HTR1A, HTR1B, HTR2A, HTR2C, HTR3A, HTR6, MAOA, TPH1, TPH2 ) and dopaminergic genes ( SLC6A3, DRD2, DRD3, DRD4, COMT, MAOA, TH, DBH ), which have been previously reported to be implicated with personality traits.
One hundred and ninety-five single nucleotide polymorphisms (SNPs) within these genes were genotyped with the Illumina BeadChip technology (HumanHap300, Human-1) in a sample of 366 mentally healthy Caucasians. Additionally, we tried to replicate our results in an independent sample of further 335 Caucasians. Personality traits in both samples were assessed with the German version of Cloninger's Tridimensional Personality Questionnaire.
From 30 SNPs showing associations at a nominal level of significance, two intronic SNPs, rs2770296 and rs927544, both located in the HTR2A gene, withstood correction for multiple testing. These SNPs were associated with the personality trait novelty seeking . The effect of rs927544 could be replicated for the novelty seeking subscale extravagance , and the same SNP was also associated with extravagance inthe combined samples.
Our results show that HTR2A polymorphisms modulate facets of novelty seeking behaviour in healthy adults suggesting that serotonergic neurotransmission is involved in this phenotype.     

Association of AKT1 gene variants and protein expression in both schizophrenia and bipolar disorder

The AKT1 gene has been associated with the genetic aetiology of schizophrenia. Following the overlap model of bipolar disorder and schizophrenia, we aimed to investigate AKT1 genetic variants and protein expression in both diseases. A total of 679 subjects with European ancestry were included: 384 with schizophrenia, 130 with bipolar disorder and 165 controls. Six single nucleotide polymorphisms (SNPs) were investigated for association with the diseases using single‐ and multi‐locus analyses. AKT1 and AKT2 protein levels were measured in post‐mortem brain tissues from ante‐mortem diagnosed schizophrenia (n = 30) and bipolar disorder subjects (n = 12) and matched controls. The analysis identified a significant global distortion in schizophrenia (P = 0.0026) and a weak association in bipolar disorder (P = 0.046). A sliding window procedure showed a five‐SNP haplotype (TCGAG) to be associated with schizophrenia (P = 1.22 × 10^?4) and bipolar disorder (P = 0.0041) and a four‐SNP haplotype (TCGA) with the combined sample (1.73 × 10^?5). On the basis of selected genotypes, a significant difference in protein expression emerged between subjects (P < 0.02). In conclusion, our findings, by showing the involvement of the AKT1 gene in both schizophrenia and bipolar disorder, support the role of AKT1 in the genetics of both disorders and add support to the view that there is some genetic overlap between them.     

Familial idiopathic basal ganglia calcification (Fahr's disease) without neurological, cognitive and psychiatric symptoms is not linked to the IBGC1 locus on chromosome 14q

Idiopathic basal ganglia calcification (IBGC) is characterised by radiological, neurological, cognitive and psychiatric abnormalities. The associations between these abnormal phenotypes and abnormal genes remain unclear despite the recent mapping to chromosome 14q of a susceptibility locus for IBGC ( IBGC1). We identified two siblings, from a large multigenerational pedigree, who had both been diagnosed with radiological IBGC, dementia, bipolar affective disorder and Parkinsonism. We assessed (1) other family members to determine whether these four phenotypes were co-segregating as symptoms of IBGC, and (2) possible IBGC linkage to the IBGC1 locus on chromosome 14q or to any known or potential dementia genes. Nine second-generation and 21 third-generation members received radiological, neurological, neuropsychological and psychiatric assessments. We genotyped all family members for microsatellite markers at the IBGC1 locus and polymorphisms of the ApoE, VLDL, alpha1-ACT, BChE-K, APP, PS1, PS2 and tau genes and tested these for linkage to IBGC, dementia and bipolar disorder. Of the ten family members with radiological intracranial calcification, all except the two index cases were normal. There was no significant association between IBGC status and severe cognitive impairment or dementia ( P=0.335) or bipolar affective disorder or Parkinsonism ( P=1.0). Linkage to the IBGC1 locus was excluded. Of the eight dementia gene markers tested, the only positive LOD score was for the ApoE epsilon4 polymorphism and dementia/severe cognitive impairment. We have identified a form of IBGC in which calcification is inherited independently of neurological, cognitive and psychiatric symptoms. This may represent a second locus for this disorder.     

Polymorphisms of Genes in Neurotransmitter Systems Were Associated with Alcohol Use Disorders in a Tibetan Population

Studies of linkage and association in various ethnic populations have revealed many predisposing genes of multiple neurotransmitter systems for alcohol use disorders (AUD). However, evidence often is contradictory regarding the contribution of most candidate genes to the susceptibility of AUD. We, therefore, performed a case-control study to investigate the possible associations of genes selected from multiple neurotransmitter systems with AUD in a homogeneous Tibetan community population in China. AUD cases (N = 281) with an alcohol use disorder identification test (AUDIT) score ≥10, as well as healthy controls (N = 277) with an AUDIT score ≤5, were recruited. All participants were genotyped for 366 single nucleotide polymorphisms (SNPs) of 34 genes selected from those involved in neurotransmitter systems. Association analyses were performed using PLINK version 1.07 software. Allelic analyses before adjustment for multiple tests showed that 15 polymorphisms within seven genes were associated with AUD (p<0.05). After adjustment for the number of SNPs genotyped within each gene, only the association of a single marker (rs10044881) in HTR4 remained statistically significant. Haplotype analysis for two SNPs in HTR4 (rs17777298 and rs10044881) showed that the haplotype AG was significantly associated with the protective effect for AUD. In conclusion, the present study discovered that the HTR4 gene may play a marked role in the pathogenesis of AUD. In addition, this Tibetan population sample marginally replicated previous evidence regarding the associations of six genes in AUD.     

Association between 1603C>T polymorphism of DBH gene and bipolar disorder in a Turkish population

Objectives

Dopamine-β-hydroxylase (DBH) is the enzyme responsible for the conversion of dopamine (DA) to norepinephrine (NE, noradrenaline) which is a key neurotransmitter in the central and peripheral nervous systems. Bipolar disorder is a major psychiatric disorder. The present study was designed to explore the associations of polymorphisms of DBH gene in Turkish patients with bipolar disorder.

Methods

− 1021C>T (rs1611115) polymorphism in promoter region, 444G>A (rs1108580) polymorphism in exon 2 and 1603C>T (rs6271; C535R) polymorphism in exon11 of DBH gene were analyzed in 106 patients with bipolar disorder and 106 healthy subjects by using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis.

Results

The results showed statistically significant associations for genotypic and allelic distribution between the 1603C>T polymorphism and bipolar disease (p = 0.0012 and p = 0.034, respectively). There was no association observed between the genotype and allelic frequencies for − 1021C>T and 444G>A polymorphisms and bipolar disorder.

Conclusions

Our data suggests that the 1603C>T polymorphism of the DBH gene is associated with susceptibility to bipolar disorder in a Turkish population.     

Functional polymorphisms of HSPA5: possible association with bipolar disorder

Altered endoplasmic reticulum stress (ER) response signaling is suggested in bipolar disorder. Previously, we preliminarily reported the genetic association of HSPA5 (GRP78/BiP) with bipolar disorder. Here, we extended our analysis by increasing the number of Japanese case-control samples and NIMH Genetics Initiative bipolar trio samples (NIMH trios), and also analyzed schizophrenia samples. In Japanese, nominally significant association of one haplotype was observed in extended samples of bipolar disorder but not in schizophrenia. In NIMH trios, no association was found in total samples. However, an exploratory analysis suggested that the other haplotype was significantly over-transmitted to probands only from the paternal side. The associated haplotype in Japanese or NIMH pedigrees shared three common polymorphisms in the promotor, which was found to alter promotor activity. These findings suggested promotor polymorphisms of HSPA5 may affect the interindividual variability of ER stress response and may confer a genetic risk factor for bipolar disorder.     

The interaction between TPH2 promoter haplotypes and clinical-demographic risk factors in suicide victims with major psychoses

Tryptophan hydroxylase isoform 2 (TPH2) is a rate-limiting enzyme in the biosynthesis of serotonin (5-HT) and is predominantly localized in the brain. Previous studies have suggested that there is an association between serotonergic dysfunction in the brain and suicidality. This study was designed to examine whether the -473T > A and -8396G > C polymorphisms of the TPH2 gene may be associated with completed suicide in subjects with major psychoses from the Stanley Foundation Brain Bank sample. TPH2 genotypes were determined in 69 subjects with a diagnosis of schizophrenia or bipolar disorder, among which 22 died by suicide. Genomic DNA was amplified by polymerase chain reaction and typed by automated methods. Both markers were found to be in Hardy-Weinberg equilibrium and in strong linkage disequilibrium. No association with history of suicide was found for either polymorphism. Haplotype analysis with EHAP showed no association between completed suicide and haplotype distribution (chi2 = 1.877; 3 df; P = 0.598). Nor was there any association between suicide and these genetic markers even when clinical-demographic factors were considered as covariates in the haplotype analysis. These findings suggest that these 5' marker haplotypes in the TPH2 gene do not influence suicidal behaviour.     

Receptor-Mediated Activation of G Proteins Is Increased in Postmortem Brains of Bipolar Affective Disorder Subjects

Abstract: Guanine nucleotide binding proteins (G proteins) have been implicated in the pathophysiology of bipolar affective disorder. In the present investigation receptor-mediated G protein activation and changes in G protein trimeric state were examined in frontal cortical membranes obtained from postmortem brains of bipolar affective disorder subjects and from age-, sex-, and postmortem interval-matched controls. Stimulation of cortical membranes with serotonin, isoproterenol, or carbachol increased guanosine 5′-O-(3-[³⁵S]thiophosphate) ([³⁵S]GTPγS) binding to specific G_α proteins in a receptor-selective manner. The abilities of these receptor agonists to stimulate the binding of [³⁵S]GTPγS to the G_α proteins was enhanced in membranes from bipolar brains. Immunoblot analyses showed increases in the levels of membrane 45- and 52-kDa G_αs proteins but no changes in the amounts of G_αi, G_αo, G_αz, G_αq/11, or G_β proteins in membrane or cytosol fractions of bipolar brain homogenates. Pertussis toxin (PTX)-activated ADP-ribosylations of G_αi and G_αo were enhanced by ～80% in membranes from bipolar compared with control brains, suggesting an increase in the levels of the trimeric state of these G proteins in bipolar disorder. Serotonin-induced, magnesium-dependent reduction in PTX-mediated ADP-ribosylation of G_αi/G_αo in cortical membranes from bipolar brains was greater than that observed in controls, providing further evidence for enhanced receptor-G protein coupling in bipolar brain membranes. In addition, the amounts of G_β proteins that coimmunoprecipitated with the G_α proteins were also elevated in bipolar brains. The data show that in bipolar brain membrane there is enhanced receptor-G protein coupling and an increase in the trimeric state of the G proteins. These changes may contribute to produce exaggerated transmembrane signaling and to the alterations in affect that characterize bipolar affective disorder.     

Association of dopamine‐ and serotonin‐related genes with canine aggression

Human‐directed canine aggression was studied using 50 aggressive and 81 non‐aggressive dogs. We examined 62 single nucleotide polymorphisms (SNPs) occurring in or in the close vicinity of 16 neurotransmitter‐related genes. Allelic associations with aggression were identified for DRD1, HTR1D, HTR2C and SLC6A1. Risk or protective haplotypes for aggressive behaviour based on 2–5 SNPs were identified. The frequency of aggressive dogs varied significantly between the haplotypes within loci and the odds ratios of aggression in dogs with risk haplotypes compared with protective haplotypes varied from 4.4 (HTR2C) to 9.0 (SLC6A1). A risk haplotype across the neurotransmitter receptor gene HTR1D harboured a non‐synonymous SNP with a potential effect on protein function. We identified no haplotypes in complete association with the recorded phenotypes, supporting a complex inheritance of aggression.     

Affective Neural Responses Modulated by Serotonin Transporter Genotype in Clinical Anxiety and Depression

Serotonin transporter gene variants are known to interact with stressful life experiences to increase chances of developing affective symptoms, and these same variants have been shown to influence amygdala reactivity to affective stimuli in non-psychiatric populations. The impact of these gene variants on affective neurocircuitry in anxiety and mood disorders has been studied less extensively. Utilizing a triallelic assay (5-HTTLPR and rs25531) to assess genetic variation linked with altered serotonin signaling, this fMRI study investigated genetic influences on amygdala and anterior insula activity in 50 generalized anxiety disorder patients, 26 of whom also met DSM-IV criteria for social anxiety disorder and/or major depressive disorder, and 39 healthy comparison subjects. A Group x Genotype interaction was observed for both the amygdala and anterior insula in a paradigm designed to elicit responses in these brain areas during the anticipation of and response to aversive pictures. Patients who are S/L_G carriers showed less activity than their L_A/L_A counterparts in both regions and less activity than S/L_G healthy comparison subjects in the amygdala. Moreover, patients with greater insula responses reported higher levels of intolerance of uncertainty, an association that was particularly pronounced for patients with two L_A alleles. A genotype effect was not established in healthy controls. These findings link the serotonin transporter gene to affective circuitry findings in anxiety and depression psychopathology and further suggest that its impact on patients may be different from effects typically observed in healthy populations.     

Further confirmation of the association between anxiety and CTNND2: replication in humans

The rat genome sequencing and mapping consortium found evidence for an association between the catenin‐δ2 gene (CTNND2) and anxious behaviour. We replicated these results in humans by carrying out a genetic association test in patients with panic disorder, social phobia, generalized anxiety disorder and/or agoraphobia (N = 1714) and controls (N = 4125). We further explored the association between CTNND2 and other psychiatric disorders based on publicly available genome‐wide association results. A gene‐based test showed that single nucleotide polymorphisms (SNPs) in CTNND2 have a significantly increased signal (P < 1e^?5) and decreased P‐values. Single nucleotide polymorphism rs1012176 showed the strongest association with any anxiety disorder (odds ratio: 0.8128, SE = 0.063, P = 0.00099), but this effect was not significant after correction for multiple testing. In available genome‐wide association results from the Psychiatric Genomics Consortium we found that SNPs in CTNND2 collectively showed an increased signal for schizophrenia (P < 1e^?5) and major depressive disorder (P < 1e^?5), but not for bipolar disorder. These signals remained significant after correction for potential confounders. The association between CTNND2 and anxiety was not strong enough to be picked up in the current generation of human genome‐wide analyses, indicating the usefulness of and need for animal genetic studies to identify candidate genes for further study in human samples .     

Refined localization of a gene for pediatric gastroesophageal reflux makes HTR2A an unlikely candidate gene

Our objective in this study was to determine whether mutations in the gene for the 5-hydroxytryptamine receptor 2A (HTR2A) cause the autosomal dominant form of severe pediatric gastroesophageal reflux (GER), which we had previously mapped to a 21-cM region at chromosome 13q14. Direct sequencing of the HTR2A gene was carried out on DNA from affected and unaffected members of families with severe pediatric GER displaying genetic linkage to the HTR2A locus. In addition, we performed high-resolution linkage mapping within the GER gene region using additional polymorphic markers closely linked to HTR2A. Several previously reported polymorphisms in the HTR2A gene were identified in three families affected with GER. In addition, we identified a novel polymorphism at nucleotide -1273 in the HTR2A promoter. No mutant allele cosegregated exclusively with the GER phenotype in any family. Linkage analysis using additional polymorphic markers narrowed the region of the GER gene to a 9 cM interval between markers D13S263 and CAGR1, formally excluding HTR2A as a candidate gene. In conclusion, sequence analysis of HTR2A and linkage analysis argue against mutations in HTR2A being a cause of severe pediatric GER.     

Promoter and functional polymorphisms of HTR2C and suicide victims

In Europe, the countries with the highest suicide rates form a so-called J-curve, which starts in Finland and extends down to Slovenia—a country with one of the world's highest suicide rates. So far, the strongest association between suicide and genes has been shown for the serotonergic system. A functional polymorphism 68G>C (Cys23Ser) and a promoter polymorphism–995G>A of serotonin receptor 2C (HTR2C) have already been investigated, but no associations with suicide were determined. In the present study 334 suicide victims and 211 controls of Slovenian origin were genotyped for the above-mentioned polymorphisms using standard methods. In the case of the polymorphism–995G>A no association with suicide was found. However, a significant association was observed between female suicide victims and polymorphism 68G>C. The significance remained when we combined alleles of female and male populations. An excess of GG genotype and allele G was observed. However, no statistically important differences were present when only males were analyzed. Haplotype analysis on female population showed marginal association of haplotype G-C with suicide. The present study speaks for the plausible implication of the HTR2C in suicide susceptibility.     

Analysis of serotonin receptor 2A gene (HTR2A): Association study with autism spectrum disorder in the Indian population and investigation of the gene expression in peripheral blood leukocytes

Several studies suggest involvement of serotoninergic system in the pathophysiology of Autism Spectrum Disorder (ASD). The 5-HT receptor binding studies using ³H-lysergic acid diethylamide (³H-LSD) and linkage analysis provided evidences to consider HTR2A as a potential candidate gene for ASD. The three SNPs, −1438A/G (rs6311), 102T/C (rs6313) and 1354C/T (rs6314) of HTR2A have been well studied in the etiology of various neuropsychiatric disorders. But studies on association of this gene with ASD are limited to two reports from American and Korean populations. Additionally there are reports, which demonstrated paternal imprinting of HTR2A with expression from only one allele. So far no reports are available on HTR2A and its association with any neuropsychiatric disorders from Indian population. Therefore, the present study investigates association of the above mentioned three markers of HTR2A with ASD in Indian population using population and family-based approaches. The study also deals with allelic expression pattern of HTR2A in Peripheral Blood Leukocytes (PBLs) to understand the parental imprinting status. The genotyping analyses were carried out for probands, parents and controls. The subsequent association analyses did not show association of these markers with ASD. So, HTR2A is unlikely to be a genetic marker for ASD in Indian population. The expression analyses showed absence of monoallelic expression, suggesting lack of parental imprinting of HTR2A gene. However, we noticed methylation of the CpG sites at −1438A/G and 102T/C loci of HTR2A gene. Further bioinformatics analysis revealed absence of CpG islands in the promoter of the gene supporting biallelic expression pattern of HTR2A in PBLs.     

Analysis of serotonin receptor 2A gene (HTR2A): Association study with autism spectrum disorder in the Indian population and investigation of the gene expression in peripheral blood leukocytes

Several studies suggest involvement of serotoninergic system in the pathophysiology of Autism Spectrum Disorder (ASD). The 5-HT receptor binding studies using ³H-lysergic acid diethylamide (³H-LSD) and linkage analysis provided evidences to consider HTR2A as a potential candidate gene for ASD. The three SNPs, −1438A/G (rs6311), 102T/C (rs6313) and 1354C/T (rs6314) of HTR2A have been well studied in the etiology of various neuropsychiatric disorders. But studies on association of this gene with ASD are limited to two reports from American and Korean populations. Additionally there are reports, which demonstrated paternal imprinting of HTR2A with expression from only one allele. So far no reports are available on HTR2A and its association with any neuropsychiatric disorders from Indian population. Therefore, the present study investigates association of the above mentioned three markers of HTR2A with ASD in Indian population using population and family-based approaches. The study also deals with allelic expression pattern of HTR2A in Peripheral Blood Leukocytes (PBLs) to understand the parental imprinting status. The genotyping analyses were carried out for probands, parents and controls. The subsequent association analyses did not show association of these markers with ASD. So, HTR2A is unlikely to be a genetic marker for ASD in Indian population. The expression analyses showed absence of monoallelic expression, suggesting lack of parental imprinting of HTR2A gene. However, we noticed methylation of the CpG sites at −1438A/G and 102T/C loci of HTR2A gene. Further bioinformatics analysis revealed absence of CpG islands in the promoter of the gene supporting biallelic expression pattern of HTR2A in PBLs.