G2Cdb::Gene report

Gene id
Gene symbol
Homo sapiens
gamma-aminobutyric acid (GABA) A receptor, alpha 1
G00000853 (Mus musculus)

Databases (7)

ENSG00000022355 (Ensembl human gene)
2554 (Entrez Gene)
405 (G2Cdb plasticity & disease)
GABRA1 (GeneCards)
137160 (OMIM)
Marker Symbol
HGNC:4075 (HGNC)
Protein Sequence
P14867 (UniProt)

Synonyms (1)

  • EJM5

Literature (68)

Pubmed - other

  • A pilot multivariate parallel ICA study to investigate differential linkage between neural networks and genetic profiles in schizophrenia.

    Meda SA, Jagannathan K, Gelernter J, Calhoun VD, Liu J, Stevens MC and Pearlson GD

    Olin Neuropsychiatry Research Center, Institute of Living, 200 Retreat Avenue, Hartford, CT 06106, USA. smeda01@harthosp.org

    Understanding genetic influences on both healthy and disordered brain function is a major focus in psychiatric neuroimaging. We utilized task-related imaging findings from an fMRI auditory oddball task known to be robustly associated with abnormal activation in schizophrenia, to investigate genomic factors derived from multiple single nucleotide polymorphisms (SNPs) from genes previously shown to be associated with schizophrenia. Our major aim was to investigate the relationship of these genomic factors to normal/abnormal brain functionality between controls and schizophrenia patients. We studied a Caucasian-only sample of 35 healthy controls and 31 schizophrenia patients. All subjects performed an auditory oddball task, which consists of detecting an infrequent sound within a series of frequent sounds. Each subject was characterized on 24 different SNP markers spanning multiple risk genes previously associated with schizophrenia. We used a recently developed technique named parallel independent component analysis (para-ICA) to analyze this multimodal data set (Liu et al., 2008). The method aims to identify simultaneously independent components of each modality (functional imaging, genetics) and the relationships between them. We detected three fMRI components significantly correlated with two distinct gene components. The fMRI components, along with their significant genetic profile (dominant SNP) correlations were as follows: (1) Inferior frontal-anterior/posterior cingulate-thalamus-caudate with SNPs from Brain derived neurotropic factor (BDNF) and dopamine transporter (DAT) [r=-0.51; p<0.0001], (2) superior/middle temporal gyrus-cingulate-premotor with SLC6A4_PR and SLC6A4_PR_AG (serotonin transporter promoter; 5HTTLPR) [r=0.27; p=0.03], and (3) default mode-fronto-temporal gyrus with Brain derived neurotropic factor and dopamine transporter (BDNF, DAT) [r=-0.25; p=0.04]. Functional components comprised task-relevant regions (including PFC, ACC, STG and MTG) frequently identified as abnormal in schizophrenia. Further, gene-fMRI combinations 1 (Z=1.75; p=0.03), 2 (Z=1.84; p=0.03) and 3 (Z=1.67; p=0.04) listed above showed significant differences between controls and patients, based on their correlated loading coefficients. We demonstrate a framework to identify interactions between "clusters" of brain function and of genetic information. Our results reveal the effect/influence of specific interactions, (perhaps epistastatic in nature), between schizophrenia risk genes on imaging endophenotypes representing attention/working memory and goal directed related brain function, thus establishing a useful methodology to probe multivariate genotype-phenotype relationships.

    Funded by: NIBIB NIH HHS: 1 R01 EB006841, R01 EB005846, R01 EB005846-05A1, R01 EB006841, R01 EB006841-01A1, R01EB005846; NIMH NIH HHS: 2 R01 MH43775, 5 R01MH52886, R01 MH043775, R01 MH052886, R01 MH052886-08, R01 MH074797, R01 MH074797-03, R37 MH043775

    NeuroImage 2010;53;3;1007-15

  • Strong genetic evidence for a selective influence of GABAA receptors on a component of the bipolar disorder phenotype.

    Craddock N, Jones L, Jones IR, Kirov G, Green EK, Grozeva D, Moskvina V, Nikolov I, Hamshere ML, Vukcevic D, Caesar S, Gordon-Smith K, Fraser C, Russell E, Norton N, Breen G, St Clair D, Collier DA, Young AH, Ferrier IN, Farmer A, McGuffin P, Holmans PA, Wellcome Trust Case Control Consortium (WTCCC), Donnelly P, Owen MJ and O'Donovan MC

    Department of Psychological Medicine, School of Medicine, Cardiff University, Cardiff, UK. craddockn@cardiff.ac.uk

    Despite compelling evidence for a major genetic contribution to risk of bipolar mood disorder, conclusive evidence implicating specific genes or pathophysiological systems has proved elusive. In part this is likely to be related to the unknown validity of current phenotype definitions and consequent aetiological heterogeneity of samples. In the recent Wellcome Trust Case Control Consortium genome-wide association analysis of bipolar disorder (1868 cases, 2938 controls) one of the most strongly associated polymorphisms lay within the gene encoding the GABA(A) receptor beta1 subunit, GABRB1. Aiming to increase biological homogeneity, we sought the diagnostic subset that showed the strongest signal at this polymorphism and used this to test for independent evidence of association with other members of the GABA(A) receptor gene family. The index signal was significantly enriched in the 279 cases meeting Research Diagnostic Criteria for schizoaffective disorder, bipolar type (P=3.8 x 10(-6)). Independently, these cases showed strong evidence that variation in GABA(A) receptor genes influences risk for this phenotype (independent system-wide P=6.6 x 10(-5)) with association signals also at GABRA4, GABRB3, GABRA5 and GABRR3. [corrected] Our findings have the potential to inform understanding of presentation, pathogenesis and nosology of bipolar disorders. Our method of phenotype refinement may be useful in studies of other complex psychiatric and non-psychiatric disorders.

    Funded by: Medical Research Council: G0000934; Wellcome Trust: 079643

    Molecular psychiatry 2010;15;2;146-53

  • Genetic association studies of methamphetamine use disorders: A systematic review and synthesis.

    Bousman CA, Glatt SJ, Everall IP and Tsuang MT

    Department of Psychiatry, Center for Behavioral Genomics, University of California San Diego, La Jolla, 92037, USA.

    Efforts to understand the biological processes that increase susceptibility to methamphetamine (METH) use disorders (i.e., abuse, dependence, and psychosis) have uncovered several putative genotypic variants. However, to date a synthesis of this information has not been conducted. Thus, systematic searches of the current literature were undertaken for genetic-association studies of METH use disorders. Each gene's chromosomal location, function, and examined polymorphic markers were extracted. Frequencies, odds ratios and 95% confidence intervals for risk alleles, as well as sample size and power, were calculated. We uncovered 38 studies examining 39 genes, of which 18 were found to have a significant genotypic, allelic, and/or haplotypic association with METH use disorders. Three genes (COMT, DRD4, and GABRA1) were associated with METH abuse, nine (ARRB2, BDNF, CYP2D6, GLYT1, GSTM1, GSTP1, PDYN, PICK1, and SLC22A3) with METH dependence, two (AKT1 and GABRG2) with METH abuse/dependence, and four (DTNBP1, OPRM1, SNCA, and SOD2) with METH psychosis. Limitations related to phenotypic classification, statistical power, and potential publication bias in the current literature were noted. Similar to other behavioral, psychiatric, and substance use disorders, the genetic epidemiology of METH use disorders is complex and likely polygenic. National and international collaborative efforts are needed to increase the availability of large population-based samples and improve upon the power to detect genetic associations of small magnitude. Further, replication of the findings reviewed here along with further development of more rigorous methodologies and reporting protocols will aid in delineating the complex genetic epidemiology of METH use disorders.

    American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics 2009;150B;8;1025-49

  • Interaction of androsterone and progesterone with inhibitory ligand-gated ion channels: a patch clamp study.

    Ziegler E, Bodusch M, Song Y, Jahn K, Wolfes H, Steinlechner S, Dengler R, Bufler J and Krampfl K

    Department of Neurology, Medical University Hannover, 30625 Hanover, Germany. Ziegler.Elke@arcor.de

    Gamma-aminobutyric acid receptor type A (GABA(A)) receptor channels mediate fast inhibitory neurotransmission throughout the central nervous system while the expression of ionotropic glycine receptors is mainly restricted to the spinal cord and brain stem. Neuroactive steroids are well known as positive allosteric modulators of GABA(A) receptor function. Furthermore, there have been hints for an interaction of neuroactive steroids with ionotropic glycine receptors. The aim of the study was to characterize the effect of androsterone and progesterone on alpha(1) and alpha(1)beta glycine receptor and alpha(1)beta(2)gamma(2) GABA(A) receptor channels and to examine the molecular interactions between ligands and receptors. Electrophysiological recordings were performed on HEK 293 cells using the patch clamp technique in combination with an ultrafast perfusion system. A direct activation of inhibitory ionotropic receptors was observed for androsterone at GABA(A) receptor channels. A coactivation of currents elicited by nonsaturating agonist concentrations was observed with androsterone and progesterone at glycine and GABA(A) receptor channels. We could show that association of beta subunits with alpha subunits affects the sensitivity of glycine receptors to androsterone. In contrast to previous reports in which recombinant glycine receptors were inhibited by progesterone, a potentiating effect was revealed by our experiments. At concentrations of 0.1 mM and higher, there were also hints to a channel block-like mechanism. In conclusion, different molecular mechanisms of interaction between neuroactive steroids and GABA as well as glycine receptors could be identified and quantitatively described. Our data clarify the role of steroid compounds in the modulation of inhibitory receptor channel function.

    Naunyn-Schmiedeberg's archives of pharmacology 2009;380;4;277-91

  • Identification of new putative susceptibility genes for several psychiatric disorders by association analysis of regulatory and non-synonymous SNPs of 306 genes involved in neurotransmission and neurodevelopment.

    Gratacòs M, Costas J, de Cid R, Bayés M, González JR, Baca-García E, de Diego Y, Fernández-Aranda F, Fernández-Piqueras J, Guitart M, Martín-Santos R, Martorell L, Menchón JM, Roca M, Sáiz-Ruiz J, Sanjuán J, Torrens M, Urretavizcaya M, Valero J, Vilella E, Estivill X, Carracedo A and Psychiatric Genetics Network Group

    CIBER en Epidemiología y Salud Pública (CIBERESP), Instituto de Salud Carlos III, Madrid, Spain.

    A fundamental difficulty in human genetics research is the identification of the spectrum of genetic variants that contribute to the susceptibility to common/complex disorders. We tested here the hypothesis that functional genetic variants may confer susceptibility to several related common disorders. We analyzed five main psychiatric diagnostic categories (substance-abuse, anxiety, eating, psychotic, and mood disorders) and two different control groups, representing a total of 3,214 samples, for 748 promoter and non-synonymous single nucleotide polymorphisms (SNPs) at 306 genes involved in neurotransmission and/or neurodevelopment. We identified strong associations to individual disorders, such as growth hormone releasing hormone (GHRH) with anxiety disorders, prolactin regulatory element (PREB) with eating disorders, ionotropic kainate glutamate receptor 5 (GRIK5) with bipolar disorder and several SNPs associated to several disorders, that may represent individual and related disease susceptibility factors. Remarkably, a functional SNP, rs945032, located in the promoter region of the bradykinin receptor B2 gene (BDKRB2) was associated to three disorders (panic disorder, substance abuse, and bipolar disorder), and two additional BDKRB2 SNPs to obsessive-compulsive disorder and major depression, providing evidence for common variants of susceptibility to several related psychiatric disorders. The association of BDKRB2 (odd ratios between 1.65 and 3.06) to several psychiatric disorders supports the view that a common genetic variant could confer susceptibility to clinically related phenotypes, and defines a new functional hint in the pathophysiology of psychiatric diseases.

    American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics 2009;150B;6;808-16

  • GABRG1 and GABRA2 as independent predictors for alcoholism in two populations.

    Enoch MA, Hodgkinson CA, Yuan Q, Albaugh B, Virkkunen M and Goldman D

    Laboratory of Neurogenetics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892, USA. maenoch@dicbr.niaaa.nih.gov

    The chromosome 4 cluster of GABA(A) receptor genes is predominantly expressed in the brain reward circuitry and this chromosomal region has been implicated in linkage scans for alcoholism. Variation in one chromosome 4 gene, GABRA2, has been robustly associated with alcohol use disorders (AUD) although no functional locus has been identified. As HapMap data reveal moderate long-distance linkage disequilibrium across GABRA2 and the adjacent gene, GABRG1, it is possible that the functional locus is in GABRG1. We genotyped 24 SNPs across GABRG1 and GABRA2 in two population isolates: 547 Finnish Caucasian men (266 alcoholics) and 311 community-derived Plains Indian men and women (181 alcoholics). In both the Plains Indians and the Caucasians: (1) the GABRG1 haplotype block(s) did not extend to GABRA2; (2) GABRG1 haplotypes and SNPs were significantly associated with AUD; (3) there was no association between GABRA2 haplotypes and AUD; (4) there were several common (>or=0.05) haplotypes that spanned GABRG1 and GABRA2 (341 kb), three of which were present in both populations: one of these ancestral haplotypes was associated with AUD, the other two were more common in non-alcoholics; this association was determined by GABRG1; (5) in the Finns, three less common (<0.05) extended haplotypes showed an association with AUD that was determined by GABRA2. Our results suggest that there are likely to be independent, complex contributions from both GABRG1 and GABRA2 to alcoholism vulnerability.

    Funded by: Intramural NIH HHS: Z01 AA000305-04, Z99 AA999999

    Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology 2009;34;5;1245-54

  • GABA(A) receptor downregulation in brains of subjects with autism.

    Fatemi SH, Reutiman TJ, Folsom TD and Thuras PD

    Department of Psychiatry, University of Minnesota, Minneapolis, MN 55455, USA. fatem002@umn.edu

    Gamma-aminobutyric acid A (GABA(A)) receptors are ligand-gated ion channels responsible for mediation of fast inhibitory action of GABA in the brain. Preliminary reports have demonstrated altered expression of GABA receptors in the brains of subjects with autism suggesting GABA/glutamate system dysregulation. We investigated the expression of four GABA(A) receptor subunits and observed significant reductions in GABRA1, GABRA2, GABRA3, and GABRB3 in parietal cortex (Brodmann's Area 40 (BA40)), while GABRA1 and GABRB3 were significantly altered in cerebellum, and GABRA1 was significantly altered in superior frontal cortex (BA9). The presence of seizure disorder did not have a significant impact on GABA(A) receptor subunit expression in the three brain areas. Our results demonstrate that GABA(A) receptors are reduced in three brain regions that have previously been implicated in the pathogenesis of autism, suggesting widespread GABAergic dysfunction in the brains of subjects with autism.

    Funded by: NICHD NIH HHS: 5R01HD052074-01A2, R01 HD052074, R01 HD052074-02; NIMH NIH HHS: R24 MH068855

    Journal of autism and developmental disorders 2009;39;2;223-30

  • A conserved Cys-loop receptor aspartate residue in the M3-M4 cytoplasmic loop is required for GABAA receptor assembly.

    Lo WY, Botzolakis EJ, Tang X and Macdonald RL

    Program in Neuroscience, Vanderbilt University, Nashville, Tennessee 37232, USA.

    Members of the Cys-loop superfamily of ligand-gated ion channels, which mediate fast synaptic transmission in the nervous system, are assembled as heteropentamers from a large repertoire of neuronal subunits. Although several motifs in subunit N-terminal domains are known to be important for subunit assembly, increasing evidence points toward a role for C-terminal domains. Using a combination of flow cytometry, patch clamp recording, endoglycosidase H digestion, brefeldin A treatment, and analytic centrifugation, we identified a highly conserved aspartate residue at the boundary of the M3-M4 loop and the M4 domain that was required for binary and ternary gamma-aminobutyric acid type A receptor surface expression. Mutation of this residue caused mutant and partnering subunits to be retained in the endoplasmic reticulum, reflecting impaired forward trafficking. Interestingly although mutant and partnering wild type subunits could be coimmunoprecipitated, analytic centrifugation studies demonstrated decreased formation of pentameric receptors, suggesting that this residue played an important role in later steps of subunit oligomerization. We thus conclude that C-terminal motifs are also important determinants of Cys-loop receptor assembly.

    Funded by: NINDS NIH HHS: NS33300

    The Journal of biological chemistry 2008;283;44;29740-52

  • GABAA receptor promoter hypermethylation in suicide brain: implications for the involvement of epigenetic processes.

    Poulter MO, Du L, Weaver IC, Palkovits M, Faludi G, Merali Z, Szyf M and Anisman H

    Molecular Brain Research Group, Robarts Research Institute, Department of Physiology and Pharmacology University of Western Ontario, London, Ontario, Canada. mpoulter@robarts.ca

    Background: Epigenetic mechanisms may be involved in the reprogramming of gene expression in response to stressful stimuli. This investigation determined whether epigenetic phenomena might similarly be associated with suicide/depression.

    Methods: The expression of DNA methyltransferase (DNMT) mRNA was assessed in several brain regions of individuals who had committed suicide and had been diagnosed with major depression relative to that of individuals who had died suddenly as a result of factors other than suicide.

    Results: The DNMT gene transcripts' expression was altered in several brains regions of suicides, including frontopolar cortex, amygdala, and the paraventricular nucleus of the hypothalamus. Importantly, an increase of both mRNA and protein expression was found in the frontopolar cortex. In addition, although transcript abundance of various forms of DNMT was highly correlated in normal control subjects, this coordination of DNMT isoform expression was diminished in suicide brain. Further, within the frontopolar cortex, gene-specific aberrations in DNA methylation were apparent in the gamma-aminobutyric acid (GABA)(A) receptor alpha1 subunit promoter region, the transcript of which is underexpressed in suicide/major depressive disorder (MDD) brains. Indeed, three cytosine/guanosine sites were hypermethylated relative to control subjects. Finally, we found that DNMT-3B mRNA abundance was inversely correlated to alpha1 mRNA abundance.

    Conclusions: These data show that DNMT mRNA expression was altered in suicide brain, and this change in expression in the frontopolar cortex was associated with increased methylation of a gene whose mRNA expression has previously been shown to be reduced. These observations suggest that epigenetic mechanisms may be associated with altered gene expression in suicide/MDD.

    Biological psychiatry 2008;64;8;645-52

  • BDNF selectively regulates GABAA receptor transcription by activation of the JAK/STAT pathway.

    Lund IV, Hu Y, Raol YH, Benham RS, Faris R, Russek SJ and Brooks-Kayal AR

    Neuroscience Graduate Group and Department of Neurology, University of Pennsylvania, Philadelphia, PA 19104, USA.

    The gamma-aminobutyric acid (GABA) type A receptor (GABA(A)R) is the major inhibitory neurotransmitter receptor in the brain. Its multiple subunits show regional, developmental, and disease-related plasticity of expression; however, the regulatory networks controlling GABA(A)R subunit expression remain poorly understood. We report that the seizure-induced decrease in GABA(A)R alpha1 subunit expression associated with epilepsy is mediated by the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway regulated by brain-derived neurotrophic factor (BDNF). BDNF- and seizure-dependent phosphorylation of STAT3 cause the adenosine 3',5'-monophosphate (cAMP) response element-binding protein (CREB) family member ICER (inducible cAMP early repressor) to bind with phosphorylated CREB at the Gabra1:CRE site. JAK/STAT pathway inhibition prevents the seizure-induced decrease in GABA(A)R alpha1 abundance in vivo and, given that BDNF is known to increase the abundance of GABA(A)R alpha4 in a JAK/STAT-independent manner, indicates that BDNF acts through at least two distinct pathways to influence GABA(A)R-dependent synaptic inhibition.

    Funded by: NINDS NIH HHS: NS051710-01A1, NS051710-03, R01 NS051710, R01 NS051710-01A1, R01 NS051710-03

    Science signaling 2008;1;41;ra9

  • Mechanisms involved in the reduction of GABAA receptor alpha1-subunit expression caused by the epilepsy mutation A322D in the trafficking-competent receptor.

    Bradley CA, Taghibiglou C, Collingridge GL and Wang YT

    Brain Research Centre, University of British Columbia, Vancouver V6T 1Z3, Canada.

    A mutation in the alpha1-subunit (A322D) of GABA(A)Rs is responsible for juvenile myoclonic epilepsy in a large Canadian family. Previous work has identified that this mutant affects the cell expression and function of recombinant GABA(A)Rs, expressed in HEK293 cells. Here we have extended these observations by showing that the mutation promotes association with the endoplasmic reticulum chaperone calnexin and accelerates the degradation rate of the subunits approximately 2.5-fold. We also find that the mutation causes the subunit to be degraded largely by a lysosomal-dependent process. Furthermore, we find that the mutation results in receptors that are inserted into the plasma membrane but are more rapidly endocytosed by a dynamin and caveolin1-dependent mechanism. These results suggest that the mutant subunit can form functional receptors, but that these have a shorter lifetime on the plasma membrane.

    Funded by: Medical Research Council: G0601813

    The Journal of biological chemistry 2008;283;32;22043-50

  • The anesthetic-like effects of diverse compounds on wild-type and mutant gamma-aminobutyric acid type A and glycine receptors.

    Yang L and Sonner JM

    Department of Anesthesia and Perioperative Care, Room S-455i, University of California, San Francisco, CA 94143-0464, USA.

    Introduction: No theory of inhaled anesthetic action requires volatility of the anesthetic to accomplish the biophysical interaction of anesthetic with biological target. The identification of mutations that attenuate the effect of inhaled anesthetics on various receptors raises the possibility that nonvolatile compounds with anesthetic effects can be identified with the aid of these receptors. In previous studies, we identified compounds that were either charged or had an exceptionally low vapor pressure and which modulated anesthetic-sensitive receptors in a manner similar to inhaled anesthetics. We tested whether these, and another charged compound, shared a common mechanism with volatile anesthetics, by comparing their effect on wild-type gamma-aminobutyric acid type A (GABA(A)) or glycine receptors and mutant receptors that were engineered to be relatively resistant to inhaled anesthetics.

    Methods: The effect of beta-hydroxybutyric acid, ammonium chloride, diethylhexyl phthalate, and GABA were tested on homomeric alpha1 and mutant alpha1 (S267I) glycine receptors. The effect of sodium dodecyl sulfate and glycine were tested on alpha1 b2 gamma2s and mutant alpha1(S270I) beta2 gamma2s GABA(A) receptors. Receptors were expressed in Xenopus laevis oocytes and studied using two-electrode voltage clamping. For both GABA(A) and glycine receptors, isoflurane and ethanol were used as positive controls and propofol as a negative control (i.e., unaffected by the mutation).

    Results: Beta-hydroxybutyric acid, ammonium chloride, diethylhexyl phthalate, and GABA all enhanced glycine receptor function. This effect was reduced by the S267I mutations. Sodium dodecyl sulfate and glycine enhanced GABA(A) receptor function, and the S270I mutation attenuated this effect.

    Conclusion: These findings support the hypothesis that the compounds studied modulate GABA(A) or glycine receptors by a mechanism similar to that of isoflurane and ethanol. Comparing the effect of drugs on anesthetic-sensitive wild-type receptors with relatively less sensitive mutant receptors may help identify compounds with anesthetic effects.

    Funded by: NIGMS NIH HHS: R01 GM069379

    Anesthesia and analgesia 2008;106;3;838-45, table of contents

  • Molecular modeling and mutagenesis reveals a tetradentate binding site for Zn2+ in GABA(A) alphabeta receptors and provides a structural basis for the modulating effect of the gamma subunit.

    Trudell JR, Yue ME, Bertaccini EJ, Jenkins A and Harrison NL

    Department of Anesthesia, Stanford University School of Medicine, Stanford, California 94305-5117, USA. trudell@stanford.edu

    Gamma-aminobutyric acid type A receptors (GABA(A)-R) containing alpha1beta2gamma2 subunits are weakly inhibited by Zn2+, whereas receptors containing only the alpha1beta2 subunits are strongly inhibited. We built homology models of the ion pores of alpha1beta2 and alpha1beta2gamma2 GABA(A)-R using coordinates of the nicotinic acetylcholine receptor as a template. Threading the GABA(A)-R beta2 sequence onto this template placed the 17' histidine and the 20' glutamate residues at adjacent locations in the mouth of the pore, such that a nearly ideal tetradentate site for Zn2+ was formed from two histidine and two glutamate residues between adjacent beta subunits in the alpha1beta2 GABA(A)-R. Following optimization with CHARMM, the distance between the alpha-carbons of the adjacent histidine residues was approximately 9.2 A, close to the ideal distance for a Zn2+ binding site. Loss of inhibition by Zn2+ in alpha1beta2gamma2 GABA(A)-R can be explained by the geometry of these residues in the arrangement alpha1beta2gamma2alpha1beta2, in which the nearest C-alpha-C-alpha distance between the histidine residues is 15.5 A, too far apart for an energetically optimal Zn2+ binding site. We then mutated the gamma subunit at the 17' and/or 20' positions. Zn2+ inhibition was not restored in alpha1beta2gamma2 (I282H) receptors. A novel finding is that the modeling shows the native 20' lysine in gamma2 can compete with Zn2+ for binding to the inserted 17' histidine. Sensitivity to Zn2+ was restored in the double mutant receptor, alpha1beta2gamma2 (I282H; K285E), in which the competition with lysine was removed and a more favorable Zn2+ binding site was formed.

    Funded by: NIAAA NIH HHS: AA 13646, AA 16393, AA13378; NIGMS NIH HHS: GM073959

    Journal of chemical information and modeling 2008;48;2;344-9

  • Juvenile myoclonic epilepsy with generalised and focal electroencephalographic abnormalities: a case report with a molecular genetic study.

    Bartocci A, Elia M, Calì F, Tiacci C, Cantisani AT and Perticoni G

    S.C. di Neurofisiopatologia-Azienda Ospedaliera R. Silvestrini, S. Andrea della Fratte, I-06156, Perugia, Italy. arnybart@infinito.it

    This is the case of a 16-year-old girl with juvenile myoclonic epilepsy (JME) and maternal family history positive for epilepsy and febrile seizures, presenting ictal and interictal generalised, as well as focal paroxysmal abnormalities over the right central-temporal regions activated during sleep. The brain magnetic resonance image was normal and the seizures responded to therapy with valproate and lamotrigine. A molecular genetic analysis led to the identification of a polymorphism (A-->G) in position 10 in the intron 3 (rs949626) of the EFHC1 gene; and a polymorphism (T-->C) of the exon of the GABRA1 gene, without aminoacidic exchange. In the literature this is the first case of JME with electroencephalograph focal epileptiform abnormalities, but without EFHC1 and GABRA1 gene mutations.

    Neurological sciences : official journal of the Italian Neurological Society and of the Italian Society of Clinical Neurophysiology 2007;28;5;276-8

  • Screening of GABA(A)-receptor gene mutations in primary dystonia.

    Shang H, Lang D, Burgunder JM and Kaelin-Lang A

    Department of Neurology, West China Hospital, SiChuan University, Chengdu, China.

    Several lines of evidence suggest that GABA-ergic neurotransmission plays a role in the pathogenesis of primary dystonia in humans. In this study, we tested the hypothesis that mutations in the GABRA1, GABRB3, and GABRG2 genes encoding the alpha1, beta3, and gamma subunits of the GABA(A) receptor are involved in familial primary dystonia. All exons and exon-intron boundaries of the above genes were amplified by PCR from genomic DNA in 28 patients who had primary dystonia and a positive family history but had no mutation in any other genes known to be involved in primary dystonia. The PCR products were analyzed by single strand conformation polymorphism followed by sequencing of variant conformers compared with normal controls (n = 54). We found no mutations in these genes. We did, however, find a new polymorphism, 559 + 80G>A in intron 5 of GABRA1, and we also confirmed several that were previously reported, including 315C>T in exon 3 and 588C>T in exon 5 of GABRG2, but there were no significant differences between controls and patients in the allele and genotype frequencies of these polymorphisms. In conclusion, mutations of GABRA1, GABRB3, and GABRG2 appear not to play a major role in the development of familial primary dystonia.

    European journal of neurology 2007;14;10;1179-81

  • The GABAA receptor alpha1 subunit epilepsy mutation A322D inhibits transmembrane helix formation and causes proteasomal degradation.

    Gallagher MJ, Ding L, Maheshwari A and Macdonald RL

    Department of Neurology, Vanderbilt University, Nashville, TN 37232-8552, USA. martin.gallagher@vanderbilt.edu

    A form of autosomal dominant juvenile myoclonic epilepsy is caused by a nonconservative missense mutation, A322D, in the GABAA receptor alpha1 subunit M3 transmembrane helix. We reported previously that the A322D mutation reduced total and surface alpha1(A322D) subunit protein and that residual alpha1(A322D) subunit resided in the endoplasmic reticulum. Here, we demonstrate that the reduction in alpha1(A322D) expression results from rapid endoplasmic reticulum-associated degradation of the alpha1(A322D) subunit through the ubiquitin-proteasome system. We provide direct evidence that the alpha1(A322D) subunit misfolds and show that in at least 33% of alpha1(A322D) subunits, M3 failed to insert into the lipid bilayer. We constructed a series of mutations in the M3 domain and empirically determined the apparent free energy cost (DeltaGapp) of membrane insertion failure, and we show that the DeltaGapp correlated directly with the recently elucidated transmembrane sequence code (DeltaGLep). These data provide a biochemical mechanism for the pathogenesis of this epilepsy mutation and demonstrate that DeltaGLep predicts the efficiency of lipid partitioning of a naturally occurring protein's transmembrane domain expressed in vivo. Finally, we calculated the DeltaDeltaGLep for 277 known transmembrane missense mutations associated with Charcot-Marie-Tooth disease, diabetes insipidus, retinitis pigmentosa, cystic fibrosis, and severe myoclonic epilepsy of infancy and showed that the majority of these mutations also are likely to destabilize transmembrane domain membrane insertion, but that only a minority of the mutations would be predicted to be as destabilizing as the A322D mutation.

    Funded by: NINDS NIH HHS: K08 NS044257, NS33300, NS39479, NS44257, R01 NS033300, R01 NS039479

    Proceedings of the National Academy of Sciences of the United States of America 2007;104;32;12999-3004

  • Association analysis of GABA receptor subunit genes on 5q33 with heroin dependence in a Chinese male population.

    Loh EW, Tang NL, Lee DT, Liu SI and Stadlin A

    Division of Psychiatry and Drug Abuse Research, National Health Research Institutes at Taipei City Psychiatric Center, Sinyi District, Taipei City, Taiwan.

    GABAA receptor subunit genes clustered on 5q33 play a role in the development of alcoholism and methamphetamine use disorder without psychosis. The present study explored the possible contribution of the same subunit genes to the development of heroin dependence. Single nucleotide polymorphisms (SNPs) of the GABAA receptor subunits GABRB2, GABRA6, GABRA1, and GABRG2 were examined in 178 male Han Chinese heroin-dependent and 170 male control subjects. A significant difference in allele frequency for the SNP rs211014 in the GABAAgamma2 receptor subunit gene between cases and controls was identified (P = 0.015). A possible mechanism for the involvement of the GABA receptor subunit genes on 5q33 in the development of heroin dependence is discussed.

    American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics 2007;144B;4;439-43

  • Dysfunction of GABAA receptor glycolysis-dependent modulation in human partial epilepsy.

    Laschet JJ, Kurcewicz I, Minier F, Trottier S, Khallou-Laschet J, Louvel J, Gigout S, Turak B, Biraben A, Scarabin JM, Devaux B, Chauvel P and Pumain R

    Institut National de la Santé et de la Recherche Médicale, Unité 573, F-75014 Paris, France. jacques.laschet@broca.inserm.fr

    A reduction in GABAergic neurotransmission has been put forward as a pathophysiological mechanism for human epilepsy. However, in slices of human epileptogenic neocortex, GABAergic inhibition can be clearly demonstrated. In this article we present data showing an increase in the functional lability of GABAergic inhibition in epileptogenic tissue compared with nonepileptogenic human tissue. We have previously shown that the glycolytic enzyme GAPDH is the kinase involved in the glycolysis-dependent endogenous phosphorylation of the alpha1-subunit of GABA(A) receptor, a mechanism necessary for maintaining GABA(A) function. In human epileptogenic cortex obtained during curative surgery of patients with partial seizures, we demonstrate an intrinsic deficiency of GABA(A) receptor endogenous phosphorylation resulting in an increased lability of GABAergic currents in neurons isolated from this tissue when compared with neurons from nonepileptogenic human tissue. This feature was not related to a reduction in the number of GABA(A) receptor alpha1-subunits in the epileptogenic tissue as measured by [(3)H]flunitrazepam photoaffinity labeling. Maintaining the receptor in a phosphorylated state either by favoring the endogenous phosphorylation or by inhibiting a membrane-associated phosphatase is needed to sustain GABA(A) receptor responses in epileptogenic cortex. The increased functional lability induced by the deficiency in phosphorylation can account for transient GABAergic disinhibition favoring seizure initiation and propagation. These findings imply new therapeutic approaches and suggest a functional link to the regional cerebral glucose hypometabolism observed in patients with partial epilepsy, because the dysfunctional GABAergic mechanism depends on the locally produced glycolytic ATP.

    Proceedings of the National Academy of Sciences of the United States of America 2007;104;9;3472-7

  • GABA(A)-receptor mRNA expression in the prefrontal and temporal cortex of ALS patients.

    Petri S, Kollewe K, Grothe C, Hori A, Dengler R, Bufler J and Krampfl K

    Department of Neurology, Hannover Medical School, Carl-Neuberg-Str. 1, 30623 Hannover, Germany. petri.susanne@MH-Hannover.de

    There is evidence that excitotoxic cell death is involved in the pathogenesis of amyotrophic lateral sclerosis (ALS). Electrophysiological and histological studies support the pathophysiological concept of an impaired inhibitory, namely GABAergic, control of the motoneurons in the cerebral cortex of ALS patients. Recently, pathological, neuropsychological and functional imaging data have challenged the view that ALS is a disorder restricted to the motor system. The aim of our study was to investigate the expression of the most abundant GABA(A)-receptor subunit mRNAs and the GABA synthesizing enzyme glutamic acid decarboxylase (GAD) in the prefrontal, temporal, occipital and cerebellar cortex of ALS patients compared to tissue of control persons. We performed in situ hybridization histochemistry (ISH) on human post-mortem cortex sections of ALS patients (n=5) and age-matched controls with no history of neurological disease (n=5). In the prefrontal and temporal cortex of ALS patients, we detected significantly reduced mRNA expression of the alpha1-subunit, while the GABA synthesizing enzyme glutamic acid decarboxylase (GAD) was significantly upregulated in these regions. In the occipital and cerebellar cortex, we did not see disease-specific differences of the mRNA expression of the investigated subunits.

    Journal of the neurological sciences 2006;250;1-2;124-32

  • Mutations in the GABRA1 and EFHC1 genes are rare in familial juvenile myoclonic epilepsy.

    Ma S, Blair MA, Abou-Khalil B, Lagrange AH, Gurnett CA and Hedera P

    Department of Neurology, Vanderbilt University, 465 21st Avenue South, 6140 MRB III, Nashville, TN 37232-8552, USA.

    Juvenile myoclonic epilepsy (JME), accounting for approximately 25% of idiopathic generalized epilepsies, is genetically heterogeneous. Mutations in the alpha-1 subunit of the GABAA receptor (GABRA1) and EFHC1 genes have been reported in a few families with autosomal dominant (AD) JME. We have investigated the contribution of these two genes to familial JME in our cohort of 54 JME Caucasian families. Syndromic classification of JME was based on previously published criteria. We considered kindreds with at least one affected first-degree relative and the evidence of a vertical transmission as definite AD JME, and families with at least one affected second-degree relative as probable AD JME. We included 33 families meeting criteria for definitive AD JME and 21 that were classified as probable AD JME. None of these families were considered informative enough to analyze candidate loci for JME using linkage analysis. We have systematically screened coding exons of these two genes using temperature gradient capillary electrophoresis. Every heteroduplex with an abnormal mobility was sequenced. No disease-causing mutations in the GABRA1 gene were identified. Analysis of EFHC1 gene found one putative disease-causing mutation R221H that was previously reported as a tandem mutation. Several synonymous and non-synonymous coding polymorphisms were identified but the allelic frequency did not differ between controls and affected individuals. Our data suggests that the majority of familial AD JME is not caused by mutations in the GABRA1 and EFHC1 genes.

    Funded by: NCRR NIH HHS: RR00095; NINDS NIH HHS: K08NS42743, K12NS01690

    Epilepsy research 2006;71;2-3;129-34

  • Association between GABRA1 and drinking behaviors in the collaborative study on the genetics of alcoholism sample.

    Dick DM, Plunkett J, Wetherill LF, Xuei X, Goate A, Hesselbrock V, Schuckit M, Crowe R, Edenberg HJ and Foroud T

    Department of Psychiatry, Washington University School of Medicine, St. Louis, Missouri 63110, USA. dickd@wustl.edu

    Background: A wealth of literature supports the role of gamma-aminobutyric acid (GABA) in neurobiological pathways contributing to alcohol dependence and related phenotypes. Animal studies have consistently tied rodent homologs of the GABAA receptor genes on human chromosome 5q to alcohol-related behaviors; however, human studies have produced mixed results. Family-based association analyses previously conducted in the Collaborative Study on the Genetics of Alcoholism (COGA) sample yielded no evidence of association with Diagnostic and Statistical Manual of Mental Disorder-fourth edition (DSM-IV) alcohol dependence and these genes. As a follow-up to that study, we examined several alcohol-related behaviors in the COGA sample as follows: (1) a broader definition of alcohol dependence, including DSM-III-R symptoms and Feighner criteria (referred to as COGA alcohol dependence); (2) withdrawal; (3) history of alcohol-induced blackouts; (4) level of response to alcohol; (5) age of onset of regular drinking; and (6) age at first drunkenness.

    Methods: Family-based association tests were conducted, using multiple single-nucleotide polymorphisms (SNPs) in each of the 4 GABAA receptor genes on chromosome 5q.

    Results: In GABRA1, we found evidence of association with several of the drinking behavior phenotypes, including COGA alcohol dependence, history of blackouts, age at first drunkenness, and level of response to alcohol. We did not find consistent evidence of association with the remaining genes and any of the phenotypes.

    Conclusions: We found evidence for association between GABRA1 and COGA alcohol dependence, history of blackouts, age at first drunkenness, and level of response to alcohol. These analyses suggest that efforts to characterize genetic contributions to alcohol dependence may benefit by examining alcohol-related behaviors in addition to clinical alcohol dependence diagnoses.

    Alcoholism, clinical and experimental research 2006;30;7;1101-10

  • Investigation of autism and GABA receptor subunit genes in multiple ethnic groups.

    Collins AL, Ma D, Whitehead PL, Martin ER, Wright HH, Abramson RK, Hussman JP, Haines JL, Cuccaro ML, Gilbert JR and Pericak-Vance MA

    Center for Human Genetics, Duke University Medical Center, Durham, NC, USA.

    Autism is a neurodevelopmental disorder of complex genetics, characterized by impairment in social interaction and communication, as well as repetitive behavior. Multiple lines of evidence, including alterations in levels of GABA and GABA receptors in autistic patients, indicate that the GABAergic system, which is responsible for synaptic inhibition in the adult brain, may be involved in autism. Previous studies in our lab indicated association of noncoding single nucleotide polymorphisms (SNPs) within a GABA receptor subunit gene on chromosome 4, GABRA4, and interaction between SNPs in GABRA4 and GABRB1 (also on chromosome 4), within Caucasian autism patients. Studies of genetic variation in African-American autism families are rare. Analysis of 557 Caucasian and an independent population of 54 African-American families with 35 SNPs within GABRB1 and GABRA4 strengthened the evidence for involvement of GABRA4 in autism risk in Caucasians (rs17599165, p=0.0015; rs1912960, p=0.0073; and rs17599416, p=0.0040) and gave evidence of significant association in African-Americans (rs2280073, p=0.0287 and rs16859788, p=0.0253). The GABRA4 and GABRB1 interaction was also confirmed in the Caucasian dataset (most significant pair, rs1912960 and rs2351299; p=0.004). Analysis of the subset of families with a positive history of seizure activity in at least one autism patient revealed no association to GABRA4; however, three SNPs within GABRB1 showed significant allelic association; rs2351299 (p=0.0163), rs4482737 (p=0.0339), and rs3832300 (p=0.0253). These results confirmed our earlier findings, indicating GABRA4 and GABRB1 as genes contributing to autism susceptibility, extending the effect to multiple ethnic groups and suggesting seizures as a stratifying phenotype.

    Funded by: NCRR NIH HHS: M01 RR-00095, M01 RR000095; NIA NIH HHS: R01 AG020135, R01 AG20135; NINDS NIH HHS: NS26630, P01 NS026630, R01 NS036768, R01 NS042165, R01 NS36768, R01 NS42165

    Neurogenetics 2006;7;3;167-74

  • An asymmetric contribution to gamma-aminobutyric type A receptor function of a conserved lysine within TM2-3 of alpha1, beta2, and gamma2 subunits.

    Hales TG, Deeb TZ, Tang H, Bollan KA, King DP, Johnson SJ and Connolly CN

    Department of Pharmacology & Physiology.

    Mutations that impair the expression and/or function of gamma-aminobutyric acid type A (GABAA) receptors can lead to epilepsy. The familial epilepsy gamma2(K289M) mutation affects a basic residue conserved in the TM2-3 linker of most GABAA subunits. We investigated the effect on expression and function of the Lys --> Met mutation in mouse alpha1(K278M), beta2(K274M), and gamma2(K289M) subunits. Compared with cells expressing wild-type and alpha1beta2gamma2(K289M) receptors, cells expressing alpha1(K278M)beta2gamma2 and alpha1beta2(K274M)gamma2 receptors exhibited reduced agonist-evoked current density and reduced GABA potency, with no change in single channel conductance. The low current density of alpha1beta2(K274M)gamma2 receptors coincided with reduced surface expression. By contrast the surface expression of alpha1(K278M)beta2gamma2 receptors was similar to wild-type and alpha1beta2gamma2(K289M) receptors suggesting that the alpha1(K278M) impairs function. In keeping with this interpretation GABA-activated channels mediated by alpha1(K278M)beta2gamma2 receptors had brief open times. To a lesser extent gamma2(K289M) also reduced mean open time, whereas beta2(K274M) had no effect. We used propofol as an alternative GABAA receptor agonist to test whether the functional deficits of mutant subunits were specific to GABA activation. Propofol was less potent as an activator of alpha1(K278M)beta2gamma2 receptors. By contrast, neither beta2(K274M) nor gamma2(K289M) affected the potency of propofol. The beta2(K274M) construct was unique in that it reduced the efficacy of propofol activation relative to GABA. These data suggest that the alpha1 subunit Lys-278 residue plays a pivotal role in channel gating that is not dependent on occupancy of the GABA binding site. Moreover, the conserved TM2-3 loop lysine has an asymmetric function in different GABAA subunits.

    Funded by: NIGMS NIH HHS: GM058037

    The Journal of biological chemistry 2006;281;25;17034-43

  • Genetic analysis of the GABRA1 gene in patients with essential tremor.

    Deng H, Xie WJ, Le WD, Huang MS and Jankovic J

    Department of Neurology, Baylor College of Medicine, 6501 Fannin Street, Houston, TX 77030, USA.

    The gamma-aminobutyric acid A (GABA-A) receptor mediates inhibitory neurotransmission in the brain and as such may be involved in certain neurological movement disorders, such as tremor. GABA-A receptor alpha 1 (Gabra)(-/-) mice have been reported to exhibit postural and kinetic, alcohol-responsive, tremor that is characteristic of essential tremor (ET), the most common form of tremor. To determine whether ET is associated with the GABRA1 gene mutation, we screened 76 patients with familial ET and found a novel nucleotide variant: IVS8+24 G>T (nt 6119289) in a male patient, and a known 156T>C polymorphism (nt 6090903) in exon 4 in 41% patients, which results in a silent mutation (G52G). No significant association between 156T>C variant and disease risk was found (adjusted OR=0.95, 95% CI=0.57-1.61; p=0.858) by further analysis of 121 familial ET patients and 114 normal controls, except a novel 96A>G (Q32Q; nt 6090743) variant, found in a normal control. Since the 156T>C variant appears to be not pathogenically relevant, our results suggest that missense, nonsense or splice site mutation in the coding region of the GABRA1 gene is not a major genetic cause of ET in Caucasian subjects.

    Funded by: NINDS NIH HHS: NS 043567, NS 40370

    Neuroscience letters 2006;401;1-2;16-9

  • Allelic variants of the gamma-aminobutyric acid-A receptor alpha1-subunit gene (GABRA1) are not associated with idiopathic gonadotropin-dependent precocious puberty in girls with and without electroencephalographic abnormalities.

    Brito VN, Mendonca BB, Guilhoto LM, Freitas KC, Arnhold IJ and Latronico AC

    Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo Disciplina de Endocrinologia e Metabologia, Avenue Dr. Eneas de Carvalho Aguiar, 155-2 andar Bloco 6, 05403900 Sao Paulo, Brazil. vinbrito@uol.com.br

    Context: gamma-Aminobutyric acid (GABA) is a dominant inhibitory neurotransmitter involved in the modulation of brain electric activity and puberty onset in primates. GABA inhibitory effects on GnRH neurons are mainly mediated by GABA-A receptor alpha1-subunit.

    Objective: The objective of this study was to investigate functional mutations or polymorphisms of the GABA-A receptor alpha1-subunit gene (GABRA1) in girls with idiopathic gonadotropin-dependent precocious puberty (GDPP) with and without electroencephalographic (EEG) abnormalities.

    Design: The entire coding region of GABRA1 was sequenced in all patients. Two known GABRA1 polymorphisms were investigated by GeneScan software analysis or enzymatic restriction. Seventy-three normal women were used as controls for genetic study. EEG tracings were recorded in 23 girls with GDPP and 17 girls with adequate pubertal development.

    Setting: The study was performed at a university hospital.

    Patients: Thirty-one girls from 28 unrelated families with idiopathic GDPP were studied.

    Results: Automatic sequencing revealed no functional mutations in girls with GDPP. Seven different GABRA1 polymorphisms, including two exonic (156T>C and 1323G>A) and five intronic [IVS2-712(GT)n, IVS3+12A>T, IVS8+45T>G, IVS9+76A>G, and IVS10+15G>A], were found in GDPP girls and controls. Abnormal EEG tracings were found in 26% of 23 girls with GDPP, two of them with epilepsy. The genotype and allele frequencies of the GABRA1 polymorphisms were not statistically different between unrelated GDPP girls and controls or between GDPP girls with or without EEG abnormalities.

    Conclusions: GABRA1 functional mutations or polymorphisms are not associated with the intrinsic mechanism of GDPP in girls with and without EEG abnormalities.

    The Journal of clinical endocrinology and metabolism 2006;91;6;2432-6

  • Association between alcoholism and the genetic polymorphisms of the GABAA receptor genes on chromosome 5q33-34 in Korean population.

    Park CS, Park SY, Lee CS, Sohn JW, Hahn GH and Kim BJ

    Department of Psychiatry, College of Medicine, Gyeongsang National University, Jinju, Korea.

    Family, twin, and adoption studies have demonstrated that genes play an important role in the development of alcoholism. We investigated the association between alcoholism and the genetic polymorphisms of the GABAA receptor genes on chromosome 5q33-34 in Korean population. The genotype of the GABAA receptor gene polymorphisms were determined by performing polymerase chain reaction genotyping for 172 normal controls and 162 male alcoholics who are hospitalized in alcoholism treatment institute. We found a significant association between the genetic polymorphisms of the GABAA alpha1 and GABAA alpha6 receptor gene and alcoholism. The GG genotype of the GABAA alpha1 receptor gene was associated with the onset age of alcoholism and alcohol withdrawal symptoms, and a high score on the Korean version of the ADS. However, there was no association between the genetic polymorphisms of the GABAA beta2 and gamma2 receptor gene and alcoholisms. Our finding suggest that genetic polymorphisms of the GABAA alpha1 and GABAA alpha6 receptor gene may be associated with the development of alcoholism and that the GG genotype of the GABAA alpha1 receptor gene play an important role in the development of the early onset and the severe type of alcoholism.

    Journal of Korean medical science 2006;21;3;533-8

  • The alpha1 subunit of GABAA receptor is repressed by c-myc and is pro-apoptotic.

    Vaknin UA and Hann SR

    Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2175, USA.

    The c-myc oncoprotein plays a critical role in the regulation of cellular proliferation and apoptosis. To mediate these biological functions, a variety of target genes are activated or repressed by c-myc, but few genes have yet been identified that directly mediate c-myc's role in proliferation or apoptosis. During a screen for genes that are repressed by c-myc, we identified the alpha1 subunit of gamma aminobutyric acid receptor (GABAAR-alpha1) as a novel target of c-myc. GABAAR is the major inhibitory neurotransmitter receptor in the mammalian central nervous system and is involved in developmental events in the brain, such as neurite outgrowth, neuronal survival, neuronal migration, and proliferation. We show here that GABAAR-alpha1 expression is rapidly and directly repressed by c-myc. GABAAR-alpha1 expression is elevated in c-myc null cells and upregulation of GABAAR-alpha1 correlates with downregulation of c-myc protein expression during neuronal cell differentiation. We also show that overexpression of GABAAR-alpha1 causes apoptosis, which is blocked by the coexpression of Bcl-2 or Bcl-XL. Induction of apoptosis is specific for the alpha1 subunit, since neither the beta1 or beta2 subunits of GABAAR induced apoptosis. Derepression of GABAAR-alpha1 expression upon downregulation of c-myc represents a unique apoptotic mechanism and a distinct function for the alpha1 subunit, independent of its role as a component of the GABAAR in the plasma membrane. In addition, the regulation of GABAAR-alpha1 expression by c-myc provides a potential direct role for the Myc proteins in neurological processes and neurodegenerative disorders.

    Funded by: NCI NIH HHS: R01 CA47399

    Journal of cellular biochemistry 2006;97;5;1094-103

  • Identification of amino acid residues important for assembly of GABA receptor alpha1 and gamma2 subunits.

    Sarto-Jackson I, Ramerstorfer J, Ernst M and Sieghart W

    Division of Biochemistry and Molecular Biology, Centre for Brain Research, Medical University of Vienna, Austria.

    Comparative models of GABA(A) receptors composed of alpha1 beta3 gamma2 subunits were generated using the acetylcholine-binding protein (AChBP) as a template and were used for predicting putative engineered cross-link sites between the alpha1 and the gamma2 subunit. The respective amino acid residues were substituted by cysteines and disulfide bond formation between subunits was investigated on co-transfection into human embryonic kidney (HEK) cells. Although disulfide bond formation between subunits could not be observed, results indicated that mutations studied influenced assembly of GABA(A) receptors. Whereas residue alpha1A108 was important for the formation of assembly intermediates with beta3 and gamma2 subunits consistent with its proposed location at the alpha1(+) side of GABA(A) receptors, residues gamma2T125 and gamma2P127 were important for assembly with beta3 subunits. Mutation of each of these residues also caused an impaired expression of receptors at the cell surface. In contrast, mutated residues alpha1F99C, alpha1S106C or gamma2T126C only impaired the formation of receptors at the cell surface when co-expressed with subunits in which their predicted interaction partner was also mutated. These data are consistent with the prediction that the mutated residue pairs are located close to each other.

    Journal of neurochemistry 2006;96;4;983-95

  • Diversification of transcriptional modulation: large-scale identification and characterization of putative alternative promoters of human genes.

    Kimura K, Wakamatsu A, Suzuki Y, Ota T, Nishikawa T, Yamashita R, Yamamoto J, Sekine M, Tsuritani K, Wakaguri H, Ishii S, Sugiyama T, Saito K, Isono Y, Irie R, Kushida N, Yoneyama T, Otsuka R, Kanda K, Yokoi T, Kondo H, Wagatsuma M, Murakawa K, Ishida S, Ishibashi T, Takahashi-Fujii A, Tanase T, Nagai K, Kikuchi H, Nakai K, Isogai T and Sugano S

    Life Science Research Laboratory, Central Research Laboratory, Hitachi, Ltd., Kokubunji, Tokyo, 185-8601, Japan.

    By analyzing 1,780,295 5'-end sequences of human full-length cDNAs derived from 164 kinds of oligo-cap cDNA libraries, we identified 269,774 independent positions of transcriptional start sites (TSSs) for 14,628 human RefSeq genes. These TSSs were clustered into 30,964 clusters that were separated from each other by more than 500 bp and thus are very likely to constitute mutually distinct alternative promoters. To our surprise, at least 7674 (52%) human RefSeq genes were subject to regulation by putative alternative promoters (PAPs). On average, there were 3.1 PAPs per gene, with the composition of one CpG-island-containing promoter per 2.6 CpG-less promoters. In 17% of the PAP-containing loci, tissue-specific use of the PAPs was observed. The richest tissue sources of the tissue-specific PAPs were testis and brain. It was also intriguing that the PAP-containing promoters were enriched in the genes encoding signal transduction-related proteins and were rarer in the genes encoding extracellular proteins, possibly reflecting the varied functional requirement for and the restricted expression of those categories of genes, respectively. The patterns of the first exons were highly diverse as well. On average, there were 7.7 different splicing types of first exons per locus partly produced by the PAPs, suggesting that a wide variety of transcripts can be achieved by this mechanism. Our findings suggest that use of alternate promoters and consequent alternative use of first exons should play a pivotal role in generating the complexity required for the highly elaborated molecular systems in humans.

    Genome research 2006;16;1;55-65

  • Genetic investigation of chromosome 5q GABAA receptor subunit genes in schizophrenia.

    Petryshen TL, Middleton FA, Tahl AR, Rockwell GN, Purcell S, Aldinger KA, Kirby A, Morley CP, McGann L, Gentile KL, Waggoner SG, Medeiros HM, Carvalho C, Macedo A, Albus M, Maier W, Trixler M, Eichhammer P, Schwab SG, Wildenauer DB, Azevedo MH, Pato MT, Pato CN, Daly MJ and Sklar P

    Program in Medical and Population Genetics, Broad Institute of Harvard and Massachusetts Institute of Technology, Cambridge, MA, USA.

    We previously performed a genome-wide linkage scan in Portuguese schizophrenia families that identified a risk locus on chromosome 5q31-q35. This finding was supported by meta-analysis of 20 other schizophrenia genome-wide scans that identified 5q23.2-q34 as the second most compelling susceptibility locus in the genome. In the present report, we took a two-stage candidate gene association approach to investigate a group of gamma-aminobutyric acid (GABA) A receptor subunit genes (GABRA1, GABRA6, GABRB2, GABRG2, and GABRP) within our linkage peak. These genes are plausible candidates based on prior evidence for GABA system involvement in schizophrenia. In the first stage, associations were detected in a Portuguese patient sample with single nucleotide polymorphisms (SNPs) and haplotypes in GABRA1 (P=0.00062-0.048), GABRP (P=0.0024-0.042), and GABRA6 (P=0.0065-0.0088). The GABRA1 and GABRP findings were replicated in the second stage in an independent German family-based sample (P=0.0015-0.043). Supportive evidence for association was also obtained for a previously reported GABRB2 risk haplotype. Exploratory analyses of the effects of associated GABRA1 haplotypes on transcript levels found altered expression of GABRA6 and coexpressed genes of GABRA1 and GABRB2. Comparison of transcript levels in schizophrenia patients and unaffected siblings found lower patient expression of GABRA6 and coexpressed genes of GABRA1. Interestingly, the GABRA1 coexpressed genes include synaptic and vesicle-associated genes previously found altered in schizophrenia prefrontal cortex. Taken together, these results support the involvement of the chromosome 5q GABAA receptor gene cluster in schizophrenia, and suggest that schizophrenia-associated haplotypes may alter expression of GABA-related genes.

    Funded by: NIMH NIH HHS: MH058693, MH52618

    Molecular psychiatry 2005;10;12;1074-88, 1057

  • Endoplasmic reticulum retention and associated degradation of a GABAA receptor epilepsy mutation that inserts an aspartate in the M3 transmembrane segment of the alpha1 subunit.

    Gallagher MJ, Shen W, Song L and Macdonald RL

    Department of Neurology, Vanderbilt University, Nashville, Tennessee 37232, USA.

    A GABA(A) receptor alpha1 subunit epilepsy mutation (alpha1(A322D)) introduces a negatively charged aspartate residue into the hydrophobic M3 transmembrane domain of the alpha1 subunit. We reported previously that heterologous expression of alpha1(A322D)beta2gamma2 receptors in mammalian cells resulted in reduced total and surface alpha1 subunit protein. Here we demonstrate the mechanism of this reduction. Total alpha1(A322D) subunit protein was reduced relative to wild type protein by a similar amount when expressed alone (86 +/- 6%) or when coexpressed with beta2 and gamma2S subunits (78 +/- 6%), indicating an expression reduction prior to subunit oligomerization. In alpha1beta2gamma2S receptors, endoglycosidase H deglycosylated only 26 +/- 5% of alpha1 subunits, consistent with substantial protein maturation, but in alpha1(A322D)beta2gamma2S receptors, endoglycosidase H deglycosylated 91 +/- 4% of alpha1(A322D) subunits, consistent with failure of protein maturation. To determine the cellular localization of wild type and mutant subunits, the alpha1 subunit was tagged with yellow (alpha1-YFP) or cyan (alpha1-CFP) fluorescent protein. Confocal microscopic imaging demonstrated that 36 +/- 4% of alpha1-YFPbeta2gamma2 but only 5 +/- 1% alpha1(A322D)-YFPbeta2gamma2 colocalized with the plasma membrane, whereas the majority of the remaining receptors colocalized with the endoplasmic reticulum (55 +/- 4% alpha1-YFPbeta2gamma2S, 86 +/- 3% alpha1(A322D)-YFP). Heterozygous expression of alpha1-CFPbeta2gamma2S and alpha1(A322D)-YFPbeta2gamma2S or alpha1-YFPbeta2gamma2S and alpha1(A322D)-CFPbeta2gamma2S receptors showed that membrane GABA(A) receptors contained primarily wild type alpha1 subunits. These data demonstrate that the A322D mutation reduces alpha1 subunit expression after translation, but before assembly, resulting in endoplasmic reticulum-associated degradation and membrane alpha1 subunits that are almost exclusively wild type subunits.

    Funded by: NINDS NIH HHS: K08 NS044257, K08NS44257-01, NS33300, NS39479

    The Journal of biological chemistry 2005;280;45;37995-8004

  • Mutation screen of GABRA1, GABRB2 and GABRG2 genes in Japanese patients with absence seizures.

    Ito M, Ohmori I, Nakahori T, Ouchida M and Ohtsuka Y

    Department of Child Neurology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Science. 2-5-1 Shikata-cho, Okayama 700-8558, Japan.

    Absence seizures are classified into typical and atypical absences according to clinical and EEG characteristics. Although missense mutations in the GABA(A) receptor gamma2 subunits (GABRG2) gene have recently been detected in two families with typical absence seizures, no study has been carried out to clarify the relationship between atypical absence and GABA(A) receptors. We performed mutation analysis of all the coding exons of GABA(A) receptor alpha1, beta2 and gamma2 subunit (GABRA1, GABRB2 and GABRG2) genes by direct sequencing to clarify whether there was common molecular biological mechanism underlying both typical and atypical absences. We recruited 52 unrelated Japanese patients, thirty-eight with typical absences and 14 with atypical absences. They consisted of 38 with childhood absence epilepsy, three with Lennox-Gastaut syndrome, two with epilepsy with myoclonic-astatic seizures and nine with epilepsy with continuous spike-waves during slow wave sleep. All of the subjects were idiopathic or cryptogenic cases without any organic brain lesions or underlying diseases. We detected five polymorphisms (T156C in GABRA1, C1194T in GABRB2, and C315T, T588C and C1230T in GABRG2), and they are silent mutations. In conclusion, mutations in GABRA1, GABRB2 and GABRG2 do not seem to be a major genetic cause of epilepsy with typical and atypical absences in Japanese subjects.

    Neuroscience letters 2005;383;3;220-4

  • Molecular analysis of the A322D mutation in the GABA receptor alpha-subunit causing juvenile myoclonic epilepsy.

    Krampfl K, Maljevic S, Cossette P, Ziegler E, Rouleau GA, Lerche H and Bufler J

    Neurologische Klinik, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625 Hannover, Germany. krampfl.klaus@mh-hannover.de

    Juvenile myoclonic epilepsy (JME) belongs to the most common forms of hereditary epilepsy, the idiopathic generalized epilepsies. Although the mode of inheritance is usually complex, mutations in single genes have been shown to cause the disease in some families with autosomal dominant inheritance. The first mutation in a multigeneration JME family has been recently found in the alpha1-subunit of the GABAA receptor (GABRA1), predicting the single amino acid substitution A322D. We further characterized the functional consequences of this mutation by coexpressing alpha1-, beta2- and gamma2-subunits in human embryonic kidney (HEK293) cells. By using an ultrafast application system, mutant receptors have shown reduced macroscopic current amplitudes at saturating GABA concentrations and a highly reduced affinity to GABA compared to the wild-type (WT). Dose-response curves for current amplitudes, activation kinetics, and GABA-dependent desensitization parameters showed a parallel shift towards 30- to 40-fold higher GABA concentrations. Both deactivation and resensitization kinetics were considerably accelerated in mutant channels. In addition, mutant receptors labelled with enhanced green fluorescent protein (EGFP) were not integrated in the cell membrane, in contrast to WT receptors. Therefore, the A322D mutation leads to a severe loss-of-function of the human GABAA receptor by several mechanisms, including reduced surface expression, reduced GABA-sensitivity, and accelerated deactivation. These molecular defects could decrease and shorten the resulting inhibitory postsynaptic currents (IPSCs) in vivo, which can induce a hyperexcitability of the postsynaptic membrane and explain the occurrence of epileptic seizures.

    The European journal of neuroscience 2005;22;1;10-20

  • The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC).

    Gerhard DS, Wagner L, Feingold EA, Shenmen CM, Grouse LH, Schuler G, Klein SL, Old S, Rasooly R, Good P, Guyer M, Peck AM, Derge JG, Lipman D, Collins FS, Jang W, Sherry S, Feolo M, Misquitta L, Lee E, Rotmistrovsky K, Greenhut SF, Schaefer CF, Buetow K, Bonner TI, Haussler D, Kent J, Kiekhaus M, Furey T, Brent M, Prange C, Schreiber K, Shapiro N, Bhat NK, Hopkins RF, Hsie F, Driscoll T, Soares MB, Casavant TL, Scheetz TE, Brown-stein MJ, Usdin TB, Toshiyuki S, Carninci P, Piao Y, Dudekula DB, Ko MS, Kawakami K, Suzuki Y, Sugano S, Gruber CE, Smith MR, Simmons B, Moore T, Waterman R, Johnson SL, Ruan Y, Wei CL, Mathavan S, Gunaratne PH, Wu J, Garcia AM, Hulyk SW, Fuh E, Yuan Y, Sneed A, Kowis C, Hodgson A, Muzny DM, McPherson J, Gibbs RA, Fahey J, Helton E, Ketteman M, Madan A, Rodrigues S, Sanchez A, Whiting M, Madari A, Young AC, Wetherby KD, Granite SJ, Kwong PN, Brinkley CP, Pearson RL, Bouffard GG, Blakesly RW, Green ED, Dickson MC, Rodriguez AC, Grimwood J, Schmutz J, Myers RM, Butterfield YS, Griffith M, Griffith OL, Krzywinski MI, Liao N, Morin R, Morrin R, Palmquist D, Petrescu AS, Skalska U, Smailus DE, Stott JM, Schnerch A, Schein JE, Jones SJ, Holt RA, Baross A, Marra MA, Clifton S, Makowski KA, Bosak S, Malek J and MGC Project Team

    The National Institutes of Health's Mammalian Gene Collection (MGC) project was designed to generate and sequence a publicly accessible cDNA resource containing a complete open reading frame (ORF) for every human and mouse gene. The project initially used a random strategy to select clones from a large number of cDNA libraries from diverse tissues. Candidate clones were chosen based on 5'-EST sequences, and then fully sequenced to high accuracy and analyzed by algorithms developed for this project. Currently, more than 11,000 human and 10,000 mouse genes are represented in MGC by at least one clone with a full ORF. The random selection approach is now reaching a saturation point, and a transition to protocols targeted at the missing transcripts is now required to complete the mouse and human collections. Comparison of the sequence of the MGC clones to reference genome sequences reveals that most cDNA clones are of very high sequence quality, although it is likely that some cDNAs may carry missense variants as a consequence of experimental artifact, such as PCR, cloning, or reverse transcriptase errors. Recently, a rat cDNA component was added to the project, and ongoing frog (Xenopus) and zebrafish (Danio) cDNA projects were expanded to take advantage of the high-throughput MGC pipeline.

    Funded by: PHS HHS: N01-C0-12400

    Genome research 2004;14;10B;2121-7

  • Association studies of neurotransmitter gene polymorphisms in alcoholic Caucasians.

    Foley PF, Loh EW, Innes DJ, Williams SM, Tannenberg AE, Harper CG and Dodd PR

    Department of Biochemistry, School of Molecular and Microbial Sciences, University of Queensland, St. Lucia, Brisbane, Australia. p.foley@mailbox.uq.edu.au

    Ethanol enhances mesolimbic/cortical dopamine activity in reward and reinforcement circuits. We investigated the hypothesis that risk for alcoholism may be mediated by genes for neurotransmitters associated with the dopamine reward system as well as genes for enzymes involved in ethanol metabolism. DNA was extracted from brain tissue collected at autopsy from pathologically characterized alcoholics and controls. PCR-based assays showed that alcoholism was associated with polymorphisms of the dopamine D2 receptor (DRD2) TaqI B (P = .029) and the GABAA-beta2 subunit C1412T (P = .012) genes, but not with the glutamate receptor subunit gene NMDAR2B (366C/G), the serotonin transporter gene (5HTTL-PR), the dopamine transporter gene DAT1(SLC6A3), the dopamine D2 receptor gene DRD2 TaqI A, or the GABAA alpha1(A15G), alpha6(T1519C), and gamma2(G3145A) subunit genes. The glial glutamate transporter gene EAAT2 polymorphism G603A was associated with alcoholic cirrhosis (P = .048). The genotype for the most active alcohol dehydrogenase enzyme ADH1C was associated with a lower risk of alcoholism (P = .026) and was less prevalent in alcoholics with DRD2TaqIA2/A2 (P = .047), GABAA-beta2 1412C/C (P = .01), or EAAT2 603G/A (P = .022) genotypes. Combined DRD2TaqI A or B with GABAA-beta2 or EAAT2 G603A genotypes may have a concerted influence in the predisposition to alcoholism.

    Annals of the New York Academy of Sciences 2004;1025;39-46

  • GABAA receptor alpha1 and alpha6 subunits mediate cell surface anchoring in cultured cells.

    Peran M, Hooper H, Rayner SL, Stephenson FA and Salas R

    Departamento de Bioquímica, Facultad de Medicina, Universidad de Málaga, Campus de Teatinos, Málaga 29080, Spain.

    The clustering and immobility of gamma-aminobutyric acid type A receptors (GABAARs) at discrete and functionally significant domains on the nerve cell surface is an important determinant in the integration of synaptic inputs. To investigate the role that different GABAAR alpha subunit isoforms play in determining receptor mobility, alphaxbeta3gamma2s subunits (where x = subunit isoforms 1-6) were co-transfected into COS 7 and human embryonic kidney (HEK) 293 cells and the surface mobility of these recombinant complexes was measured by fluorescence photobleach recovery (FPR). In addition, the lateral mobility of endogenous GABAARs in cerebellar granule (CG) cells was measured. We show that the alpha1 and alpha6 subunits immobilize recombinant GABAAR in transfected cells. This is consistent with the immobility of native receptors in CG cells, which express alpha1 and alpha6.

    Neuroscience letters 2004;364;2;67-70

  • The juvenile myoclonic epilepsy GABA(A) receptor alpha1 subunit mutation A322D produces asymmetrical, subunit position-dependent reduction of heterozygous receptor currents and alpha1 subunit protein expression.

    Gallagher MJ, Song L, Arain F and Macdonald RL

    Department of Neurology, Vanderbilt University, Nashville, Tennessee 37212, USA.

    Individuals with autosomal dominant juvenile myoclonic epilepsy are heterozygous for a GABA(A) receptor alpha1 subunit mutation (alpha1A322D). GABA(A) receptor alphabetagamma subunits are arranged around the pore in a beta-alpha-beta-alpha-gamma sequence (counterclockwise from the synaptic cleft). Therefore, each alpha1 subunit has different adjacent subunits, and heterozygous expression of alpha1(A322D), beta, and gamma subunits could produce receptors with four different subunit arrangements: beta-alpha1-beta-alpha1-gamma (wild type); beta-alpha1(A322D)-beta-alpha1-gamma (Het(betaalphabeta)); beta-alpha1-beta-alpha1(A322D)-gamma (Het(betaalphagamma));beta-alpha1(A322D)-beta-alpha1(A322D)-gamma (homozygous). Expression of a 1:1 mixture of wild-type andalpha1(A322D) subunits with beta2S and gamma2S subunits (heterozygous transfection) produced smaller currents than wild type and much larger currents than homozygous mutant transfections. Western blot and biotinylation assays demonstrated that the amount of total and surface alpha1 subunit from heterozygous transfections was also intermediate between those of wild-type and homozygous mutant transfections. alpha1(A322D) mutations were then made in covalently tethered triplet (gamma2S-beta2S-alpha1) and tandem (beta2S-alpha1) concatamers to target selectively alpha1(A322D) to each of the asymmetric alpha1 subunits. Coexpression of mutant and wild-type concatamers resulted in expression of either Het(betaalphabeta) or Het(betaalphagamma) receptors. Het(betaalphabeta) currents were smaller than wild type and much larger than Het(betaalphagamma) and homozygous currents. Furthermore, Het(betaalphabeta) transfections contained less beta-alpha concatamer than wild type but more than both Het(betaalphagamma) and homozygous mutant transfections. Thus, whole-cell currents and protein expression of heterozygous alpha1(A322D)beta2Sgamma2S receptors depended on the position of the mutant alpha1 subunit, and GABA(A) receptor currents in heterozygous individuals likely result primarily from wild-type and Het(betaalphabeta) receptors with little contribution from Het(betaalphagamma) and homozygous receptors.

    Funded by: NINDS NIH HHS: K08 NS044257, K08 NS44257-01, NS33300, NS39479

    The Journal of neuroscience : the official journal of the Society for Neuroscience 2004;24;24;5570-8

  • Gating allosterism at a single class of etomidate sites on alpha1beta2gamma2L GABA A receptors accounts for both direct activation and agonist modulation.

    Rüsch D, Zhong H and Forman SA

    Department of Anesthesia and Critical Care, Massachusetts General Hospital, Boston, MA 02114, USA.

    At clinical concentrations, the potent intravenous general anesthetic etomidate enhances gamma-aminobutyric acid, type A (GABA(A)) receptor activity elicited with low gamma-aminobutyric acid (GABA) concentrations, whereas much higher etomidate concentrations activate receptors in the absence of GABA. Therefore, GABA(A) receptors may possess two types of etomidate sites: high affinity GABA-modulating sites and low affinity channel-activating sites. However, GABA modulation and direct activation share stereoselectivity for the (R)(+)-etomidate isomer and display parallel dependence on GABA(A) beta subunit isoforms, suggesting that these two actions may be mediated by a single class of etomidate site(s) that exert one or more molecular effects. In this study, we assessed GABA modulation by etomidate using leftward shifts of electrophysiological GABA concentration responses in cells expressing human alpha1beta2gamma2L receptors. Etomidate at up to 100 microm reduced GABA EC(50) values by over 100-fold but without apparent saturation, indicating the absence of high affinity etomidate sites. In experiments using a partial agonist, P4S, etomidate both reduced EC(50) and increased maximal efficacy, demonstrating that etomidate shifts the GABA(A) receptor gating equilibrium toward open states. Results were quantitatively analyzed using equilibrium receptor gating models, wherein a postulated class of equivalent etomidate sites both directly activates receptors and enhances agonist gating. A Monod-Wyman-Changeux co-agonist mechanism with two equivalent etomidate sites that allosterically enhance GABA(A) receptor gating independently of agonist binding most simply accounts for direct activation and agonist modulation. This model also correctly predicts the actions of etomidate on GABA(A) receptors containing a point mutation that increases constitutive gating activity.

    Funded by: NIGMS NIH HHS: P01 GM 58448, R01 GM 66724

    The Journal of biological chemistry 2004;279;20;20982-92

  • Modelling extracellular domains of GABA-A receptors: subtypes 1, 2, 3, and 5.

    Chou KC

    Tianjin Institute of Bioinformatics and Drug Discovery, Tianjin 300074, China. kchou@san.rr.com

    GABA is the main inhibitory neurotransmitter in the mammalian central nervous system. When GABA binds to the ubiquitous GABA-A receptors on neurons, chloride channels are activated leading to a rapid increase in chloride conductance that depresses excitatory depolarization. The GABA-A receptors are targets for many clinically important drugs, such as the benzodiazepines, general anaesthetics, and barbiturates. All of these drugs enhance the chloride current activated by GABA. Of the GABA-A receptor family, the subtype 2 is critical for the treatment of anxiety spectrum disorders. To avoid unwanted side effects, it is necessary to find highly selective drugs that interact only with subtype 2 but not with the related receptors such as subtypes 1, 3, and 5. To realize such a goal, it is important to have not only the 3D (dimensional) structure of subtype 2 but also the 3D structures of subtypes 1, 3, and 5. In this study, the 3D structures of all the four subtypes of GABA-A receptors have been derived. The computer-modeled heteropentameric structures bear the following features: (1) each of the five subunits in the pentamer has an intrachain disulfide bond, a hallmark of ligand-gated pentameric channels; (2) those residues which are sensitive to the binding of the benzodiazepine site ligands are grouped around the alpha1,2,3,5/gamma2 interfaces; and (3) those residues which are sensitive to the binding of GABA molecules are grouped around the alpha1,2,3,5/beta2 interfaces. All these findings are fully consistent with experimental observations. Meanwhile, for those sensitive or key residues, a close look at their subtle difference among the four subtypes has been provided through a highlighted superposition picture. In addition to providing the atomic coordinates, the predicted structures have further clarified some ambiguities that could not been uniquely determined by the existing experimental data, such as the directionality of the subunit arrangement in the heteropentamers. The 3D models may provide a reasonable structural frame or footing for designing highly selective drugs. The present models might be also useful in understanding the basic mechanism of operation of the GABA-A receptors, stimulating novel strategies for developing more specific drugs and better treatments.

    Biochemical and biophysical research communications 2004;316;3;636-42

  • A mutation in the GABAA receptor alpha 1 subunit linked to human epilepsy affects channel gating properties.

    Fisher JL

    Department of Pharmacology, Physiology and Neuroscience, University of South Carolina School of Medicine, Blg 1 Rm D23, Columbia, SC 29209, USA. jfisher@med.sc.edu

    A genetic component is associated with the development of many forms of epilepsy. Recently, mutations in the GABAA receptor have been linked to several inherited epilepsies. One of these mutations is a non-conservative change of alanine to aspartate in the third transmembrane domain of the alpha1 subunit. To determine the functional consequences of this alteration, mutated alpha subunits were transiently transfected along with wild-type beta3 and gamma2L subunits into HEK-293T cells. The mutated alpha1(A294D) subunit reduced GABA sensitivity of the receptor, increased the deactivation rate and slowed desensitization. The mutation caused a reduction in channel open time but no change in single channel conductance. Studies with additional mutants, altering the charge and/or size of the side-chain, indicated that both size and hydrophobicity of the residue at this location influence channel gating. The effects on GABA sensitivity, deactivation rate and channel open time are consistent with a reduced efficacy of channel gating, and would be expected to decrease GABAergic neurotransmission. The alpha1 subtype is the most widely expressed of the alpha subunits, with expression increasing throughout development. Therefore, production of the mutated subunit could cause global hyperexcitability throughout the brain, leading to generalized seizures with juvenile onset.

    Neuropharmacology 2004;46;5;629-37

  • Comparative cellular distribution of GABAA and GABAB receptors in the human basal ganglia: immunohistochemical colocalization of the alpha 1 subunit of the GABAA receptor, and the GABABR1 and GABABR2 receptor subunits.

    Waldvogel HJ, Billinton A, White JH, Emson PC and Faull RL

    Department of Anatomy with Radiology, Faculty of Medical and Health Sciences, University of Auckland, Private Bag 92019, Auckland, New Zealand. h.waldvogel@auckland.ac.nz

    The GABA(B) receptor is a G-protein linked metabotropic receptor that is comprised of two major subunits, GABA(B)R1 and GABA(B)R2. In this study, the cellular distribution of the GABA(B)R1 and GABA(B)R2 subunits was investigated in the normal human basal ganglia using single and double immunohistochemical labeling techniques on fixed human brain tissue. The results showed that the GABA(B) receptor subunits GABA(B)R1 and GABA(B)R2 were both found on the same neurons and followed the same distribution patterns. In the striatum, these subunits were found on the five major types of interneurons based on morphology and neurochemical labeling (types 1, 2, 3, 5, 6) and showed weak labeling on the projection neurons (type 4). In the globus pallidus, intense GABA(B)R1 and GABA(B)R2 subunit labeling was found in large pallidal neurons, and in the substantia nigra, both pars compacta and pars reticulata neurons were labeled for both receptor subunits. Studies investigating the colocalization of the GABA(A) alpha(1) subunit and GABA(B) receptor subunits showed that the GABA(A) receptor alpha(1) subunit and the GABA(B)R1 subunit were found together on GABAergic striatal interneurons (type 1 parvalbumin, type 2 calretinin, and type 3 GAD neurons) and on neurons in the globus pallidus and substantia nigra pars reticulata. GABA(B)R1 and GABA(B)R2 were found on substantia nigra pars compacta neurons but the GABA(A) receptor alpha(1) subunit was absent from these neurons. The results of this study provide the morphological basis for GABAergic transmission within the human basal ganglia and provides evidence that GABA acts through both GABA(A) and GABA(B) receptors. That is, GABA acts through GABA(B) receptors, which are located on most of the cell types of the striatum, globus pallidus, and substantia nigra. GABA also acts through GABA(A) receptors containing the alpha(1) subunit on specific striatal GABAergic interneurons and on output neurons of the globus pallidus and substantia nigra pars reticulata.

    The Journal of comparative neurology 2004;470;4;339-56

  • Possible association between a haplotype of the GABA-A receptor alpha 1 subunit gene (GABRA1) and mood disorders.

    Horiuchi Y, Nakayama J, Ishiguro H, Ohtsuki T, Detera-Wadleigh SD, Toyota T, Yamada K, Nankai M, Shibuya H, Yoshikawa T and Arinami T

    Department of Medical Genetics, Institute of Basic Medical Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.

    Background: The gamma-aminobutyric acid (GABA) neurotransmitter system has been implicated in the pathogenesis of mood disorders. The GABRA1 gene encodes one of the subunits of GABA-A receptor and is located on human chromosome 5q34-q35, which is a region reportedly linked to mood disorders. We examined the GABRA1 gene as a candidate for mood disorders.

    Methods: We performed mutation screening of GABRA1 in 24 Japanese bipolar patients and evaluated associations in Japanese case-control subjects consisting of 125 patients with bipolar disorder, 147 patients with depressive disorders, and 191 healthy control subjects. Associations were confirmed in the National Institute of Mental Health (NIMH) Initiative Bipolar Pedigrees, which consists of 88 multiplex pedigrees with 480 informative persons.

    Results: We identified 13 polymorphisms in the GABRA1 gene. Nonsynonymous mutations were not found. Association of a specific haplotype with affective disorders was suggested in the Japanese case-control population (corrected p=.0008). This haplotype association was confirmed in the NIMH pedigrees (p=.007).

    Conclusions: These results indicate that the GABRA1 gene may play a role in the etiology of bipolar disorders.

    Funded by: NIMH NIH HHS: U01 MH46274, U01 MH46280, U01 MH46282

    Biological psychiatry 2004;55;1;40-5

  • Recombinant alpha 1 beta 2 gamma 2 GABA(A) receptors expressed in HEK293 and in QT6 cells show different kinetics.

    Mercik K, Pytel M and Mozrzymas JW

    Department of Biophysics, Wroclaw Medical University, ul. Chalz.cansls;ubińskiego 10, 50-368, Wroclaw, Poland.

    Neuronal gamma-aminobutyric acid (GABA)(A) receptors are extremely heterogeneous and therefore GABAergic currents represent responses of unknown mixture of receptor subtypes. Expression of recombinant receptors in foreign cells allows to investigate a defined receptor subtype but its properties can be altered due to, e.g., differences in the endogenous modulators. In the present study the alpha1beta2gamma2 receptors were expressed in HEK293 and QT-6 cells and current responses to ultrafast GABA applications were recorded. Rise time and rapid deactivation component were faster in responses recorded from QT-6 cells. Moreover, in QT-6 cells desensitization was faster and more profound. Recovery in the paired pulse experiments was faster in HEK293 cells. In conclusion, we provide evidence that recombinant receptors may show functional differences when expressed in different cells.

    Neuroscience letters 2003;352;3;195-8

  • Absence of GABRA1 Ala322Asp mutation in juvenile myoclonic epilepsy families from India.

    Kapoor A, Vijai J, Ravishankar HM, Satishchandra P, Radhakrishnan K and Anand A

    Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Jakkur, Bangalore 560 064, India. anand@jncasr.ac.in

    An Ala322Asp mutation in the GABRA1 gene was recently reported to be responsible for causing the autosomal dominant (AD) form of juvenile myoclonic epilepsy (JME) in a French-Canadian family. To study if JME families from India exhibiting the AD mode of inheritance carry the Ala322Asp mutation, we examined 35 unrelated JME-affected individuals from such families for the Ala322Asp mutation in GABRA1. Ala322Asp mutation was not observed in any of these JME-affected individuals, suggesting that this mutation is unlikely to be a predominant mutation involved in causation of epilepsy. To evaluate the possibility of other mutation(s) in and around GABRA1 that may predispose to JME, we compared the allele frequencies at two marker loci, D5S2118 and D5S422, flanking GABRA1, in probands and 100 matched population controls. One of the allele frequencies at D5S422 shows a significant difference between the cases and controls (chi-square = 11.44, d.f. = 1, P = 0.0007), suggesting genetic association between JME and genes located in the proximity of the DNA marker.

    Journal of genetics 2003;82;1-2;17-21

  • Interaction of calcineurin and type-A GABA receptor gamma 2 subunits produces long-term depression at CA1 inhibitory synapses.

    Wang J, Liu S, Haditsch U, Tu W, Cochrane K, Ahmadian G, Tran L, Paw J, Wang Y, Mansuy I, Salter MM and Lu YM

    Neuroscience Research Group, Department of Physiology and Biophysics, Faculty of Medicine, University of Calgary, Calgary, Canada, T2N 4N1.

    Long-term depression (LTD) is an activity-dependent weakening of synaptic efficacy at individual inhibitory synapses, a possible cellular model of learning and memory. Here, we show that the induction of LTD of inhibitory transmission recruits activated calcineurin (CaN) to dephosphorylate type-A GABA receptor (GABA(A)Rs) via the direct binding of CaN catalytic domain to the second intracellular domain of the GABA(A)R-gamma(2) subunits. Prevention of the CaN-GABA(A) receptor complex formation by expression of an autoinhibitory domain of CaN in the hippocampus of transgenic mice blocks the induction of LTD. Conversely, genetic expression of the CaN catalytic domain in the hippocampus depresses inhibitory synaptic responses, occluding LTD. Thus, an activity-dependent physical and functional interaction between CaN and GABA(A) receptors is both necessary and sufficient for inducing LTD at CA1 individual inhibitory synapses.

    The Journal of neuroscience : the official journal of the Society for Neuroscience 2003;23;3;826-36

  • Coupling of agonist binding to channel gating in the GABA(A) receptor.

    Kash TL, Jenkins A, Kelley JC, Trudell JR and Harrison NL

    Graduate Program in Neuroscience, Weill Graduate School of Biomedical Sciences, Cornell University, New York, New York 10021, USA.

    Neurotransmitters such as acetylcholine and GABA (gamma-aminobutyric acid) mediate rapid synaptic transmission by activating receptors belonging to the gene superfamily of ligand-gated ion channels (LGICs). These channels are pentameric proteins that function as signal transducers, converting chemical messages into electrical signals. Neurotransmitters activate LGICs by interacting with a ligand-binding site, triggering a conformational change in the protein that results in the opening of an ion channel. This process, which is known as 'gating', occurs rapidly and reversibly, but the molecular rearrangements involved are not well understood. Here we show that optimal gating in the GABA(A) receptor, a member of the LGIC superfamily, is dependent on electrostatic interactions between the negatively charged Asp 57 and Asp 149 residues in extracellular loops 2 and 7, and the positively charged Lys 279 residue in the transmembrane 2-3 linker region of the alpha1-subunit. During gating, Asp 149 and Lys 279 seem to move closer to one another, providing a potential mechanism for the coupling of ligand binding to opening of the ion channel.

    Nature 2003;421;6920;272-5

  • Association analysis of exonic variants of the GABA(B)-receptor gene and alpha electroencephalogram voltage in normal subjects and alcohol-dependent patients.

    Winterer G, Mahlberg R, Smolka MN, Samochowiec J, Ziller M, Rommelspacher HP, Herrmann WM, Schmidt LG and Sander T

    Department of Psychiatry, Benjamin Franklin University Hospital, Free University of Berlin, Berlin, Germany. wintereg@intra.nimh.nih.gov

    Based on pharmacologic evidence, it has been suggested that GABA(B) receptors may play a crucial role in the generation of alpha-electroencephalogram (EEG) oscillations. We tested whether three exonic variants of the gene encoding the human GABA(B) receptor (GABA(B)R1) modify scalp-recorded alpha-EEG voltage. One hundred twenty-eight patients suffering from alcoholism and 114 normal subjects were investigated. Alcohol-dependent patients were included because evidence exists that the frequently observed alpha low voltage in these subjects is at least partly a trait variable. Logistic regression analyses revealed no associations between alpha-EEG voltage and polymorphic variations in exon 1a1 or exon 11. A significant interaction was observed for an exon 7 substitution polymorphism and diagnosis (P = 0.009). Post hoc analyses showed an association between EEG phenotype and exon 7 genotype in normal subjects only. It is concluded that this particular association may only be observable under physiologic conditions and that alpha low voltage in alcohol-dependent subjects is under the control of either different genes or that it is related to the disease process.

    Behavior genetics 2003;33;1;7-15

  • Gender-specific contribution of the GABA(A) subunit genes on 5q33 in methamphetamine use disorder.

    Lin SK, Chen CK, Ball D, Liu HC and Loh EW

    Department of Addiction Science, Taipei City Psychiatric Center, Taipei, Taiwan.

    Family and twins studies have suggested that genetic factors are involved in the development of substance use disorders. Several unrelated case/control association studies have reported associations of the GABA(A) subunit genes on 5q33 with the development of alcohol dependence. We hypothesized that these particular GABA(A) subunit genes also contribute to the development of methamphetamine use disorder. To test our hypothesis, we recruited cases using a series of questionnaires. Among the polymorphic SNPs, significant differences between cases and controls were identified in the female sample in the rs2279020 of the GABA(A)alpha1 subunit gene, and the novel SNP rs4480617 in the GABA(A)gamma2 subunit gene. No associations were found in the male sample. Further haplotype analysis identified several marker blocks significantly associated with methamphetamine use disorder in females; each block consists of the rs4480617. Our study provides preliminary evidence that the GABA(A) subunit genes on 5q33 may preferentially contribute to methamphetamine use disorder in females.

    The pharmacogenomics journal 2003;3;6;349-55

  • Unique assignment of inter-subunit association in GABA(A) alpha 1 beta 3 gamma 2 receptors determined by molecular modeling.

    Trudell J

    Department of Anesthesia and Beckman Program for Molecular and Genetic Medicine, Stanford School of Medicine, Stanford, CA 94305-5117, USA. trudell@stanford.edu

    Recent publications defined requirements for inter-subunit contacts in a benzodiazepine-sensitive GABA(A) receptor (GABA(A)R alpha 1 beta 3 gamma 2). There is strong evidence that the heteropentameric receptor contains two alpha 1, two beta 3, and one gamma 2 subunit. However, the available data do not distinguish two possibilities: When viewed clockwise from an extracellular viewpoint the subunits could be arranged in either gamma 2 beta 3 alpha 1 beta 3 alpha 1 or gamma 2 alpha 1 beta 3 alpha 1 beta 3 configurations. Here we use molecular modeling to thread the relevant GABA(A)R subunit sequences onto a template of homopentameric subunits in the crystal structure of the acetylcholine binding protein (AChBP). The GABA(A) sequences are known to have 15-18% identity with the acetylcholine binding protein and nearly all residues that are conserved within the nAChR family are present in AChBP. The correctly aligned GABA(A) sequences were threaded onto the AChBP template in the gamma 2 beta 3 alpha 1 beta 3 alpha 1 or gamma 2 alpha 1 beta 3 alpha 1 beta 3 arrangements. Only the gamma 2 alpha 1 beta 3 alpha 1 beta 3 arrangement satisfied three known criteria: (1) alpha 1 His(102) binds at the gamma 2 subunit interface in proximity to gamma 2 residues Thr(142), Phe(77), and Met(130); (2) alpha 1 residues 80-100 bind near gamma 2 residues 91-104; and (3) alpha 1 residues 58-67 bind near the beta 3 subunit interface. In addition to predicting the most likely inter-subunit arrangement, the model predicts which residues form the GABA and benzodiazepine binding sites.

    Funded by: NIGMS NIH HHS: 1R01 GM64371

    Biochimica et biophysica acta 2002;1565;1;91-6

  • Homologous sites of GABA(A) receptor alpha(1), beta(3) and gamma(2) subunits are important for assembly.

    Sarto I, Wabnegger L, Dögl E and Sieghart W

    Division of Biochemistry and Molecular Biology, Brain Research Institute, University of Vienna, Spitalgasse 4, Austria.

    GABA(A) receptors are the major inhibitory transmitter receptors in the central nervous system. The majority of these receptors is composed of two alpha, two beta and one gamma subunit that assemble around an aqueous pore and form an intrinsic chloride ion channel. Using full-length or truncated chimeric subunits it was demonstrated that homologous sequences from different subunit classes, alpha(1)(54-68), beta(3)(52-66), and gamma(2)(67-81), are important for assembly of GABA(A) receptors composed of alpha(1), beta(3), and gamma(2) subunits. In addition, evidence was provided that these sequences all are located in topologically homologous regions of the different subunits. Finally, it was demonstrated that the sequences investigated cause a selective assembly with certain subunits only and thus influence subunit arrangement within GABA(A) receptors.

    Neuropharmacology 2002;43;4;482-91

  • Tryptophan scanning mutagenesis in TM4 of the GABA(A) receptor alpha1 subunit: implications for modulation by inhaled anesthetics and ion channel structure.

    Jenkins A, Andreasen A, Trudell JR and Harrison NL

    Department of Anesthesiology, Weill Medical College of Cornell University, New York, NY 10021, USA. anj2005@med.cornell.edu

    Previous studies have shown that amino acid residues in trans-membrane (TM) segments 1, 2 and 3 of the alpha subunit are critical for the enhancement of GABA(A) receptor function by inhaled anesthetics. In this study we used tryptophan (Trp) scanning mutagenesis between Ile 406 and Asn 417 in the alpha1 subunit to determine the effects of Trp substitution in the fourth transmembrane segment (TM4) on receptor gating and anesthetic modulation. Wild-type and mutant alpha1 subunits were transiently expressed in HEK 293 cells with wild-type beta2 and gamma2s subunits and GABA-activated currents were recorded using whole-cell voltage clamp. The potentiation by three inhaled anesthetics (isoflurane, halothane and chloroform) of responses elicited by a submaximal concentration of GABA were also examined.EC(50) values for GABA at the mutant receptors were in the range 4-60 microM (wild-type=20 microM), indicating that Trp substitution can alter the apparent affinity of the receptor for GABA positively or negatively, dependent on position. The variation of the calculated EC(50) value for GABA exhibited an interesting periodicity, with the cycle length for each repeat corresponding to approximately 3.6 amino acids. These data are consistent with an alpha-helical structure for the TM4 segment of the alpha subunit. Several of these Trp point mutations altered the ability of one or more of the three inhaled anesthetics to modulate receptor function; four of the 12 mutations abolished receptor modulation by one or more of the anesthetics tested. These data are consistent with a role for these residues at the extracellular end of TM4 in anesthetic modulation of GABA(A) receptors.

    Funded by: NIGMS NIH HHS: GM45129, GM61925, GM64371

    Neuropharmacology 2002;43;4;669-78

  • Association analysis of an (AC)n repeat polymorphism in the GABA(B) receptor gene and schizophrenia.

    Imai K, Harada S, Kawanishi Y, Tachikawa H, Okubo T and Asada T

    Department of Psychiatry, Institute of Clinical Medicine, University of Tsukuba, Ibaraki, Japan. zc6k-imi@asahi-net.or.jp

    The human gamma-aminobutyric acid type B (GABA(B)) receptor gene is a candidate gene for schizophrenia due to its chromosomal location and neurobiologic roles. In the present study, association analyses of genetic polymorphisms of the GABA(B) receptor gene with schizophrenia were carried out in 102 unrelated schizophrenic patients and 100 healthy controls, using a polymerase chain reaction-based, single-strand conformational polymorphism analysis. Although the Ala20Val and Gly489Ser mutations were not found in our samples, we found a novel polymorphism of (AC)n dinucleotide repeats located approximately 1.6 kb upstream from the translational start site. No significant difference in allele frequencies was found between controls and patients with schizophrenia (P = 0.0587) using the Monte Carlo method. Significant differences were found between controls and patients with continuous-course schizophrenia (P = 0.0019), and between controls and patients with a positive family history of psychoses (P = 0.0015). These differences, however, were not significant after Bonferroni correction. These data did not support our hypothesis that polymorphisms of the GABA(B) receptor gene may confer vulnerability for schizophrenia.

    American journal of medical genetics 2002;114;6;605-8

  • Association of protein kinase C with GABA(A) receptors containing alpha1 and alpha4 subunits in the cerebral cortex: selective effects of chronic ethanol consumption.

    Kumar S, Sieghart W and Morrow AL

    Bowles Center for Alcohol Studies, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7178, USA.

    Previous studies have suggested that protein kinase C (PKC) isoforms differentially influence the sensitivity of gamma-aminobutyric acid(A) (GABA(A) ) receptor responses in brain. Both PKCgamma and PKCepsilon knock-out mice exhibit altered ethanol potentiation of GABA(A) receptor mediated Cl(-) flux. Furthermore, chronic ethanol consumption alters GABA(A) receptor function and receptor subunit peptide expression by mechanisms that are not yet understood. The present study explored the possibility that PKC isoforms are directly associated with GABA(A) receptors, and this association is influenced by chronic ethanol exposure. GABA(A) receptors containing alpha1 or alpha4 subunits were immunoprecipitated from solubilized protein derived from the membrane fraction of rat cerebral cortex using selective antibodies. Immunoprecipitated receptors were screened by western blot analysis for the presence of PKCdelta, gamma and epsilon isoforms. We found pronounced labeling of PKCgamma but not PKCdelta or PKCepsilon in the alpha1 and alpha4 subunit immunoprecipitates. Immunoprecipitation with PKCgamma, but not with IgG antibody also yielded GABA(A) receptor alpha1 and alpha4 subunits in the immunoprecipitate. The association of PKCgamma with alpha1-containing receptors was decreased 44 +/- 11% after chronic ethanol consumption. In contrast, PKCgamma associated with alpha4-containing receptors was increased 32 +/- 7% after chronic ethanol consumption. These results suggest that PKCgamma may be involved in GABA(A) receptor adaptations following chronic ethanol consumption.

    Funded by: NIAAA NIH HHS: AA 09013, AA 11605

    Journal of neurochemistry 2002;82;1;110-7

  • Mutation of GABRA1 in an autosomal dominant form of juvenile myoclonic epilepsy.

    Cossette P, Liu L, Brisebois K, Dong H, Lortie A, Vanasse M, Saint-Hilaire JM, Carmant L, Verner A, Lu WY, Wang YT and Rouleau GA

    Centre for Research in Neuroscience, McGill University Health Center Research Institute and McGill University, 1650 Cedar Avenue, Montréal, Québec H3G 1A4, Canada.

    Although many genes that predispose for epilepsy in humans have been determined, those that underlie the classical syndromes of idiopathic generalized epilepsy (IGE) have yet to be identified. We report that an Ala322Asp mutation in GABRA1, encoding the alpha1 subunit of the gamma-aminobutyric acid receptor subtype A (GABA(A)), is found in affected individuals of a large French Canadian family with juvenile myoclonic epilepsy. Compared with wildtype receptors, GABA(A) receptors that contain the mutant subunit show a lesser amplitude of GABA-activated currents in vitro, indicating that seizures may result from loss of function of this inhibitory ligand-gated channel. Our results confirm that mutation of GABRA1 predisposes towards a common idiopathic generalized epilepsy syndrome in humans.

    Nature genetics 2002;31;2;184-9

  • Association of the gamma-aminobutyric acid A receptor gene cluster with alcohol dependence in Taiwanese Han.

    Chang YT, Sun HS, Fann CS, Chang CJ, Liao ZH, Huang JL, Loh EW, Yu WY and Cheng AT

    Molecular psychiatry 2002;7;8;828-9

  • A multivariate analysis of 59 candidate genes in personality traits: the temperament and character inventory.

    Comings DE, Gade-Andavolu R, Gonzalez N, Wu S, Muhleman D, Blake H, Mann MB, Dietz G, Saucier G and MacMurray JP

    Department of Medical Genetics, City of Hope Medical Center, Duarte, CA 91010, USA.

    Cloninger (Cloninger CR. Neurogenetic adaptive mechanisms in alcoholism. Science 1987: 236: 410-416) proposed three basic personality dimensions for temperament: novelty seeking, harm avoidance, and reward dependence. He suggested that novelty seeking primarily utilized dopamine pathways, harm avoidance utilized serotonin pathways, and reward dependence utilized norepinephrine pathways. Subsequently, one additional temperament dimension (persistence) and three character dimensions (cooperativeness, self-directedness, and self-transcendence) were added to form the temperament and character inventory (TCI). We have utilized a previously described multivariate analysis technique (Comings DE, Gade-Andavolu R, Gonzalez N et al. Comparison of the role of dopamine, serotonin, and noradrenergic genes in ADHD, ODD and conduct disorder. Multivariate regression analysis of 20 genes. Clin Genet 2000: 57: 178-196; Comings DD, Gade-Andavolu R, Gonzalez N et al. Multivariate analysis of associations of 42 genes in ADHD, ODD and conduct disorder. Clin Genet 2000: in press) to examine the relative role of 59 candidate genes in the seven TCI traits and test the hypothesis that specific personality traits were associated with specific genes. While there was some tendency for this to be true, a more important trend was the involvement of different ratios of functionally related groups of genes, and of different genotypes of the same genes, for different traits.

    Funded by: NIDA NIH HHS: R01-DA08417

    Clinical genetics 2000;58;5;375-85

  • theta, a novel gamma-aminobutyric acid type A receptor subunit.

    Bonnert TP, McKernan RM, Farrar S, le Bourdellès B, Heavens RP, Smith DW, Hewson L, Rigby MR, Sirinathsinghji DJ, Brown N, Wafford KA and Whiting PJ

    Neuroscience Research Centre, Merck Sharp and Dohme Research Laboratories, Eastwick Road, Harlow, Essex CM20 2QR, England, UK.

    gamma-Aminobutyric acid type A (GABA-A) receptors are a major mediator of inhibitory neurotransmission in the mammalian central nervous system, and the site of action of a number of clinically important drugs. These receptors exist as a family of subtypes with distinct temporal and spatial patterns of expression and distinct properties that presumably underlie a precise role for each subtype. The newest member of this gene family is the theta subunit. The deduced polypeptide sequence is 627 amino acids long and has highest sequence identity (50.5%) with the beta1 subunit. Within the rat striatum, this subunit coassembles with alpha2, beta1, and gamma1, suggesting that gamma-aminobutyric acid type A receptors consisting of arrangements other than alpha beta + gamma, delta, or epsilon do exist. Expression of alpha2beta1gamma1theta in transfected mammalian cells leads to the formation of receptors with a 4-fold decrease in the affinity for gamma-aminobutyric acid compared with alpha2beta1gamma1. This subunit has a unique distribution, with studies so far suggesting significant expression within monoaminergic neurons of both human and monkey brain.

    Proceedings of the National Academy of Sciences of the United States of America 1999;96;17;9891-6

  • Dopamine receptor D2 and D4 genes, GABA(A) alpha-1 subunit genes and response to lithium prophylaxis in mood disorders.

    Serretti A, Lilli R, Lorenzi C, Franchini L, Di Bella D, Catalano M and Smeraldi E

    Istituto Scientifico Ospedale San Raffaele, Department of Neuropsychiatric Sciences, University of Milan School of Medicine, Italy. serretti.alessandro@hsr.it

    Lithium is an effective prophylactic agent in mood disorders, and genetic factors are likely to modulate individual susceptibility to lithium treatment. The aim of this study is to investigate the influence of dopamine receptor D2 (DRD2), D4 exon 3 (DRD4), and gamma-aminobutyric acid type A (GABA(A)) receptor alpha-1 subunit (GABRA1) gene variants on the efficacy of lithium prophylaxis in mood disorders. Patients with mood disorders (N = 125: bipolar subtype, n = 100; major depressive disorder subtype, n = 25) were followed prospectively for an average of 53 months and were typed for DRD2 (Ser311/Cys311: n = 121, VNTR: n = 63), DRD4 (n = 125) and GABRA1 (n = 61) variants using polymerase chain reaction (PCR) techniques. DRD2, DRD4 and GABRA1 variants were not associated with response to lithium. A trend was observed toward a better outcome of DRD4* 2/4 subjects, but it was due to only two subjects. Consideration of possible stratification effects like gender, polarity, family history, age at onset and duration of lithium treatment did not reveal any association either. DRD2, DRD4 and GABRA1 variants therefore do not appear to be associated with the outcome of lithium prophylaxis in mood disorders.

    Psychiatry research 1999;87;1;7-19

  • No interaction of GABA(A) alpha-1 subunit and dopamine receptor D4 exon 3 genes in symptomatology of major psychoses.

    Serretti A, Macciardi F, Cusin C, Lattuada E, Lilli R, Di Bella D, Catalano M and Smeraldi E

    Department of Neuroscience, Istituto Scientifico H. San Raffaele, University of Milan School of Medicine, Italy. serretti.alessandro@mail.hsr.it

    Previously, we reported on an association of the dopamine receptor D4 (DRD4) gene with delusional symptomatology of major psychoses. However, despite the strength of the association, it only accounted for 2% of the variance, indicating that contributions from other genes were probable. In the present study, we investigated the original cohort of subjects to evaluate the gene for the gamma-aminobutyric acid type A (GABA(A)) receptor alpha-1 subunit (GABRA1). The possible association of GABRA1 with the psychopathology of major psychoses was tested both alone and in interaction with DRD4. Four hundred and sixty-one inpatients affected by major psychoses were assessed by the operational criteria checklist for psychotic illness (OPCRIT) and were also typed for the DRD4 and GABRA1 variants using PCR techniques. Mania, depression, delusion, and disorganization were the four symptomatologic factors used as phenotype definitions. GABRA1 variants were not associated with these symptomatologic factors, and consideration of possible stratification effects such as sex and psychiatric diagnosis also did not reveal any association. GABRA1 variants did not significantly influence the association of DRD4 with delusional symptoms. No interaction was observed on the other symptom factors. The GABA(A) alpha-1 subunit gene does not, therefore, interact with DRD4 in the symptomatology of major psychoses.

    American journal of medical genetics 1999;88;1;44-9

  • Maintenance of recombinant type A gamma-aminobutyric acid receptor function: role of protein tyrosine phosphorylation and calcineurin.

    Huang RQ and Dillon GH

    Department of Pharmacology, University of North Texas Health Science Center at Fort Worth, USA.

    In the present study, rundown of gamma-aminobutyric acid (GABA)-activated Cl- channels was studied in recombinant GABAA receptors stably expressed in human embryonic kidney cells (HEK 293), with conventional whole-cell and amphotericin B-perforated patch recording. When [ATP]i was lowered to 1 mM and resting [Ca++]i was buffered to a relatively high level, the response of alpha 3 beta 2 gamma 2 GABAA receptors to relatively low [GABA] (up to 50 microM) did not show rundown in the whole-cell configuration. However, high [GABA] (greater than 200 microM) induced significant rundown, which was observed by decreases in both the maximum GABA-induced current and GABA EC50. Rundown was prevented completely with a solution containing 4 mM Mg(++)-ATP and low resting [Ca++]i, or during perforated patch recording. The magnitude of rundown was comparable in alpha 1 beta 2 gamma 2 and beta 2 gamma 2 receptors. Neither stimulation nor inhibition of protein kinase A or protein kinase C had a significant effect on rundown. However, sodium metavanadate, an inhibitor of protein tyrosine phosphatase, significantly reduced rundown. In addition, inhibition of protein tyrosine kinase activity by either genistein or lavendustin A induced rundown of the GABA response. Inhibition of the Ca++/calmodulin-dependent phosphatase calcineurin with fenvalerate also prevented rundown of the response to GABA. Our results demonstrate that rundown of GABAA receptor function is concentration-dependent, due to depletion of ATP and/or unbuffered [Ca++]i, and does not depend on the presence or subtype of the alpha subunit. We propose that protein phosphorylation at a tyrosine kinase-dependent site, and a distinct unidentified site, which is dephosphorylated by calcineurin, maintains the function of GABAA receptors.

    Funded by: NIEHS NIH HHS: ES07904

    The Journal of pharmacology and experimental therapeutics 1998;286;1;243-55

  • Construction and characterization of a full length-enriched and a 5'-end-enriched cDNA library.

    Suzuki Y, Yoshitomo-Nakagawa K, Maruyama K, Suyama A and Sugano S

    International and Interdisciplinary Studies, The University of Tokyo, Japan.

    Using 'oligo-capped' mRNA [Maruyama, K., Sugano, S., 1994. Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides. Gene 138, 171-174], whose cap structure was replaced by a synthetic oligonucleotide, we constructed two types of cDNA library. One is a 'full length-enriched cDNA library' which has a high content of full-length cDNA clones and the other is a '5'-end-enriched cDNA library', which has a high content of cDNA clones with their mRNA start sites. The 5'-end-enriched library was constructed especially for isolating the mRNA start sites of long mRNAs. In order to characterize these libraries, we performed one-pass sequencing of randomly selected cDNA clones from both libraries (84 clones for the full length-enriched cDNA library and 159 clones for the 5'-end-enriched cDNA library). The cDNA clones of the polypeptide chain elongation factor 1 alpha were most frequently (nine clones) isolated, and more than 80% of them (eight clones) contained the mRNA start site of the gene. Furthermore, about 80% of the cDNA clones of both libraries whose sequence matched with known genes had the known 5' ends or sequences upstream of the known 5' ends (28 out of 35 for the full length-enriched library and 51 out of 62 for the 5'-end-enriched library). The longest full-length clone of the full length-enriched cDNA library was about 3300 bp (among 28 clones). In contrast, seven clones (out of the 51 clones with the mRNA start sites) from the 5'-end-enriched cDNA library came from mRNAs whose length is more than 3500 bp. These cDNA libraries may be useful for generating 5' ESTs with the information of the mRNA start sites that are now scarce in the EST database.

    Gene 1997;200;1-2;149-56

  • gamma-Aminobutyric acidA receptors displaying association of gamma 3-subunits with beta 2/3 and different alpha-subunits exhibit unique pharmacological properties.

    Tögel M, Mossier B, Fuchs K and Sieghart W

    Department of Biochemical Psychiatry, University Clinic for Psychiatry, Vienna, Austria.

    Two polyclonal antibodies were raised against the gamma 3-subunit of gamma-aminobutyric acidA (GABAA) receptors. These antibodies were able to precipitate GABAA receptors from brain membrane extracts, and the precipitated receptors exhibited benzodiazepine binding properties that were dramatically different from those of receptors precipitated by anti-gamma 2 antibodies. The anti-gamma 3 antibodies were used for immunopurification of GABAA receptors containing gamma 3-subunits. Western blot analysis of the immunopurified GABAA receptors indicated that the gamma 3-subunit exhibits an apparent molecular mass of 43-46 kDa. Furthermore, in addition to gamma 3-subunits, beta 2/3-, alpha 1-, alpha 2-, alpha 3-, alpha 4-, and alpha 6-subunits could be detected in immunoaffinity column eluates from total brain and cerebellum, respectively. These data indicate that gamma 3-subunits can combine with most alpha-subunits to form a multiplicity of GABAA receptors with distinct pharmacology.

    The Journal of biological chemistry 1994;269;17;12993-8

  • Isolation and characterization of the promoter of the human GABAA receptor alpha 1 subunit gene.

    Kang I, Lindquist DG, Kinane TB, Ercolani L, Pritchard GA and Miller LG

    Department of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, Boston, Massachusetts 02111.

    The GABAA receptor, as assessed by ligand binding and chloride flux measurement in vivo and in vitro, is down-regulated in response to chronic benzodiazepine exposure. The mRNA levels of the alpha 1 and gamma 2 subunits of the receptor are also reduced. We have isolated the promoter of the gene encoding the alpha 1 subunit of the GABAA receptor to elucidate the regulatory mechanism of its expression. A DNA segment 650 bp long has been isolated that includes 151 bp of untranslated 5' end of the cDNA sequence and 500 bp of potential promoter-enhancer region. The transcriptional activity of this DNA segment linked to the firefly luciferase gene showed a strong orientation specificity. The promoter activity was localized to a 60-bp segment by deletion mapping. Mobility shift binding assay results suggest that this segment may interact with one or more factors in HeLa cell nuclear extracts to form a transcriptional complex. Primary cultures of embryonic chick cortical cells transfected with the promoter-luciferase construct were treated chronically with lorazepam. Transcriptional activity of this promoter construct was strongly repressed by chronic administration of lorazepam.

    Funded by: NIDA NIH HHS: DA-05258; NIMH NIH HHS: MH-47598

    Journal of neurochemistry 1994;62;4;1643-6

  • Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides.

    Maruyama K and Sugano S

    Institute of Medical Science, University of Tokyo, Japan.

    We have devised a method to replace the cap structure of a mRNA with an oligoribonucleotide (r-oligo) to label the 5' end of eukaryotic mRNAs. The method consists of removing the cap with tobacco acid pyrophosphatase (TAP) and ligating r-oligos to decapped mRNAs with T4 RNA ligase. This reaction was made cap-specific by removing 5'-phosphates of non-capped RNAs with alkaline phosphatase prior to TAP treatment. Unlike the conventional methods that label the 5' end of cDNAs, this method specifically labels the capped end of the mRNAs with a synthetic r-oligo prior to first-strand cDNA synthesis. The 5' end of the mRNA was identified quite simply by reverse transcription-polymerase chain reaction (RT-PCR).

    Gene 1994;138;1-2;171-4

  • Confirmation of the localization of the human GABAA receptor alpha 1-subunit gene (GABRA1) to distal 5q by linkage analysis.

    Johnson KJ, Sander T, Hicks AA, van Marle A, Janz D, Mullan MJ, Riley BP and Darlison MG

    Department of Biochemistry and Molecular Genetics, St. Mary's Hospital Medical School, Imperial College, London, United Kingdom.

    The GABAA receptor is the major inhibitory neurotransmitter receptor in the mammalian brain. To date, 14 genes that encode subunits of this receptor have been identified; these appear to be scattered throughout the human genome and are under investigation as candidate loci for a number of neurological and psychiatric disorders. We report here a highly polymorphic (dC-dA)n repeat within the human alpha 1-subunit gene (GABRA1). Typing of this marker in the Centre d'Etude du Polymorphisme Humain (CEPH) panel of families confirms the previous assignment of the GABRA1 locus to the distal portion of chromosome 5q by demonstrating linkage to the markers CRI-L45 (D5S61) (Zmax = 11.00, theta max = 0.15), CRI-V1022 (D5S54) (Zmax = 7.25, theta max = 0.20), and CRI-P148 (D5S72) (Zmax = 5.71, theta max = 0.24).

    Genomics 1992;14;3;745-8

  • Sequence and expression of human GABAA receptor alpha 1 and beta 1 subunits.

    Schofield PR, Pritchett DB, Sontheimer H, Kettenmann H and Seeburg PH

    Zentrum für Molekulare Biologie, University of Heidelberg, FRG.

    The deduced amino acid sequences of cDNA clones encoding human GABAA receptor alpha 1 and beta 1 subunits are presented. The human subunits display very high levels of sequence identity with the corresponding bovine receptor subunits. The cloned human GABAA receptor subunits induce the formation of GABA-gated chloride channels when expressed in mammalian cells.

    FEBS letters 1989;244;2;361-4

  • Isolation of a cDNA clone for the alpha subunit of the human GABA-A receptor.

    Garrett KM, Duman RS, Saito N, Blume AJ, Vitek MP and Tallman JF

    Department of CNS Research, Lederle Laboratories, Pearl River, NY.

    A cDNA clone of an alpha subunit of the human GABA-A receptor has been isolated. The human clone (pCLL800) contains 1055 nucleotides in an open reading frame and 260 nucleotides in the 5' non-coding region. The 351 amino acid sequence of this human alpha subunit shows 97% homology with its bovine counterpart. Hybridization of pCLL800 to Northern blots shows a 3.9/4.3 Kb RNA doublet in human cortex, rat whole brain, cortex, hippocampus, midbrain, olfactory bulb and cerebellum. Developmental studies show that the levels of the rat alpha mRNA increase between one and three weeks of age in a manner similar to the development of the benzodiazepine binding sites.

    Biochemical and biophysical research communications 1988;156;2;1039-45

Gene lists (6)

Gene List Source Species Name Description Gene count
L00000009 G2C Homo sapiens Human PSD Human orthologues of mouse PSD adapted from Collins et al (2006) 1080
L00000016 G2C Homo sapiens Human PSP Human orthologues of mouse PSP adapted from Collins et al (2006) 1121
L00000059 G2C Homo sapiens BAYES-COLLINS-HUMAN-PSD-CONSENSUS Human cortex PSD consensus 748
L00000061 G2C Homo sapiens BAYES-COLLINS-MOUSE-PSD-CONSENSUS Mouse cortex PSD consensus (ortho) 984
L00000069 G2C Homo sapiens BAYES-COLLINS-HUMAN-PSD-FULL Human cortex biopsy PSD full list 1461
L00000071 G2C Homo sapiens BAYES-COLLINS-MOUSE-PSD-FULL Mouse cortex PSD full list (ortho) 1556
© G2C 2014. The Genes to Cognition Programme received funding from The Wellcome Trust and the EU FP7 Framework Programmes:
EUROSPIN (FP7-HEALTH-241498), SynSys (FP7-HEALTH-242167) and GENCODYS (FP7-HEALTH-241995).

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