- Email: [email protected]
P.1.01 An association of the glial cell line-derived neurotrophic factor family receptor alpha-1 gene with schizophrenia
Molecular neuropsychopharmacology the Purkinje cell output (unpublished). By contrast, we also made conventional (“static”) Purkinje cell-specific gamma2 knockouts and these mice are not ataxic on the rotarod; thus from the “static knockout”, using one particular behavioural criterion (rotarod), we would conclude that the molecular layer interneuron input onto Purkinje cells is dispensable for motor co-ordination, an opposite conclusion from that reached with the “zolpidem method”. These two approaches on the same cell type, the static knockout of GABAergic input, and the reversible modulation of GABAergic input, illustrate the desirability of pursuing both strategies to get a full picture (Wulff et al 2007). The “zolpidem method” will be applicable to a wide range of synapses throughout the brain that use GABAA receptor inputs. Funding source: Work in my lab is supported by the Medical Research Council (grant G0800399). Reference(s) [1] Gosgnach, S., Lanuza, G.M., Butt, S.J., Saueressig, H., Zhang, Y., Velasquez, T., Riethmacher, D., Callaway, E.M., Kiehn, O., Goulding, M., 2006, V1 spinal neurons regulate the speed of vertebrate locomotor outputs. Nature 440: 215–219. [2] Wulff, P., Wisden, W., 2005, Dissecting neural circuitry by combining genetics and pharmacology. Trends Neurosci 28: 44−50. [3] Wulff, P., Goetz, T., Lepp¨a, E., Linden, A.M., Renzi, M., Swinny, J.D., Vekovischeva, O.Y., Sieghart, W., Somogyi, P., Korpi, E.R., Farrant, M., Wisden, W., 2007, From synapse to behavior: rapid modulation of defined neuronal types with engineered GABAA receptors. Nat Neurosci 10: 923–929.
Posters P.1.01 An association of the glial cell linederived neurotrophic factor family receptor alpha-1 gene with schizophrenia A. Alkelai1 ° , Y. Kohn1 , S. Lupoli2 , T. Olender3 , K. SarnerKanyas1 , E. Ben-Asher3 , D. Lancet3 , F. Macciardi2 , B. Lerer1 . 1 Hadassah-Hebrew University Medical Center, Psychiatry, Jerusalem, Israel; 2 University of Milan, Department of Sciences and Biomedical Technologies, Milan, Italy; 3 Weizmann Institute of Science, Department of Molecular Genetics, Rehovot, Israel Purpose of the study: Schizophrenia is a severe, chronic neuropsychiatric disorder with poorly defined etiology and pathophysiology. No diagnostic neuropathology has
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been identified and available treatment is symptomatic and only partially successful. There is strong support for a significant role of genetic factors in the etiology of schizophrenia. Identifying schizophrenia related genes an important priority since it will help to understand the pathophysiology of the disorder and give provide the prospect of more effective treatment. A genome scan for schizophrenia related loci in Arab Israeli families by Lerer et al. [1] detected significant evidence for linkage at chromosome 6q23. Association [2] and replication studies [3] identified AHI1 as a susceptibility gene. The same genome scan revealed suggestive evidence for a schizophrenia susceptibility locus in the 10q23−26 region. Since the mode of inheritance of schizophrenia is most likely polygenic, multiple genes may be involved in its pathogenesis even in an ethnically homogeneous population. Expanding the initial phase of the project beyond the 6q23 region, we sought further evidence for linkage of 10q23-q26 with schizophrenia, set out to refine this candidate region, tested for possible genetic interaction with the 6q23 region and sought specific susceptibility gene(s). Methods: To refine the candidate region and test for genetic interaction, 35 additional microsatellite markers were genotyped on an ABI3700 Automated DNA Analyzer (Applied Biosystems) in the 10q23-q26 candidate region in the same samples that were used for the whole genome scan [1]. Further single-locus and two-locus linkage analyses were performed with GENEHUNTER software. To seek genes associated with schizophrenia in the refined 10q24.33-q26.13 region a total of 2046 SNP were genotyped by using of the Illumina’s HumanCNV370Duo BeadChip and analyzed with PBAT Version 3.6. The family sample that was genotyped was the Expanded Arab Israeli Family Sample previously described by AmannZalcenstein et al. [2]. Results: A final candidate region of 19.9Mb was found between markers D10S222 and D10S587 (10q24.33q26.13) by nonparametric and parametric singe-locus linkage analyses which revealed a suggestive MOD score of 2.56 and an NPL of 3.34 (p = 0.0006). Twolocus multipoint linkage analysis yielded a significant LOD score of 7.45 under the multiplicative model, suggesting overlapping function of the genes at these two loci. Significance was assessed by permutation analysis. Extensive SNP genotyping of the refined candidate region revealed two SNPs associated with schizophrenia after Bonferroni correction for multiple testing (2046 tests, p = 0.000024), the most intriguing of which resides in the GFRA1 gene which encodes the glial cell line-derived neurotrophic factor (GDNF) family receptor alpha-1. Conclusions: This study suggests overlapping function of genes at the 6q23 and refined 10q23-q26 loci and shows
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Molecular neuropsychopharmacology
an association of GFRA1 gene with schizophrenia. The protein encoded by GFRA1 is a member of the GDNF receptor family. It is a receptor for both GDNF and neurturin (NTN). GDNF and NTN are two structurally related, potent neurotrophic factors that play key roles in the control of neuron survival and differentiation. These roles are potentially relevant to the pathogenesis of schizophrenia. Reference(s) [1] Lerer, B., Segman, R.H., Hamdan, A., Kanyas, K., Karni, O., Kohn, Y., Korner, M., Lanktree, M., Kaadan, M., Turetsky, N., Yakir, A., Kerem, B., Macciardi, F., 2003, Genome scan of Arab Israeli families maps a schizophrenia susceptibility gene to chromosome 6q23 and supports a locus at chromosome 10q24. Mol Psychiatry 8: 488–498. [2] Amann-Zalcenstein, D., Avidan, N., Kanyas, K., Ebstein, R.P., Kohn, Y., Hamdan, A., Ben-Asher, E., Karni, O., Mujaheed, M., Segman, R.H., Maier, W., Macciardi, F., Beckmann, J.S., Lancet, D., Lerer, B., 2006, AHI1, a pivotal neurodevelopmental gene, and C6orf217 are associated with susceptibility to schizophrenia. Eur J Hum Genet 14: 1111–1119. [3] Ingason, A., Sigmundsson, T., Steinberg, S., Sigurdsson, E., Haraldsson, M., Magnusdottir, B.B., Frigge, M.L., Kong, A., Gulcher, J., Thorsteinsdottir, U., Stefansson, K., Petursson, H., Stefansson, H., 2007, Support for involvement of the AHI1 locus in schizophrenia. Eur J Hum Genet 15, 988–991. P.1.02 M- and T-tropic HIVs cause apoptosis in rat neuronal cell culture F. Biggio1 ° , A. Bachis2 , I. Mocchetti2 . 1 University of Cagliari, Department of Exp. Biology, Monserrato (Cagliari), Italy; 2 Georgetown University, Department of Neuroscience, Washington D.C., USA Human immunodeficiency virus type 1 (HIV-1) causes neuronal degeneration and, at a late stage, creates HIV-associated dementia (HAD) and other neurological abnormalities characterized by a severe loss of neurons and fibers throughout the brain that involved a broad spectrum of motor impairments and cognitive deficits. Neuronal loss, reactive astrocytes and other abnormalities are seen in the brain of individuals with Acquired Immune Deficiency Syndrome (AIDS)-associated Dementia Complex (ADC). A better understanding of the pathogenic mechanisms mediating HIV-1 neurotoxicity is vital for developing effective neuro-protective therapies against ADC. HIV-1 is believed to be the main agent causing ADC. However, little
is known about the molecular and cellular mechanisms of HIV-1 neurotoxicity considering that HIV-1 does not infect post-mitotic neurons and that viral load does not necessarily correlate with ADC. In the brain HIV-1 infects macrophages and lymphocyties after binding of the envelope glycoprotein 120 (gp-120) to CD4 in conjunction with several chemokine receptors family, such as CCR5 and CXCR4. These receptors act as co-receptors for viral entry and, consequently, regulate the lethal infection of the cells. Most HIV strains use CCR5 for HIV transmission and infection. Moreover progeny HIV-1 virions can also cause neuronal death. Several viral proteins, such as the envelope protein gp120 and the transcription activator Tat, have been shown to induce neuronal apoptosis through direct and indirect mechanisms both in vitro and in vivo. However, it has not been fully established yet whether HIV-1 promotes neuronal apoptosis by a direct mechanism. We have previously demonstrate that gpIIIB, a tropic (X4) strain, causes neuronal apoptosis through activation of the chemokine receptor CXCR4 both in vitro and in vivo. Our experiments were performed in Cortical neurons and Cerebellar granule cells prepared from 1 day old and 8 days old Sprague Dawley rat pups, respectively. To explore the neurotoxic effect of HIV-1, we exposed rat cerebellar granule cells and cortical neurons in culture to two different strains of HIV-1, IIIB and BaL, T- and M-tropic strains that utilize CXCR4 and CCR5 co-receptors, respectively, to infect cells. We observed that both viruses elicit a time-dependent apoptotic cell death in these cultures without inducing a productive infection as determined by the absence of the core protein of HIV-1, p24, in cell lysates. Instead, neurons were gp120 positive, suggesting that the envelope protein is shed by the virus and then subsequently internalized by neurons. The CXCR4 receptor antagonist AMD3100 or the CCR5 receptor inhibitor D-Ala-peptide T-amide blocked HIV IIIB and HIV Bal neurotoxicity, respectively. In contrast, the NMDA receptor blocker MK801 failed to protect neurons from HIV-mediated apoptosis, suggesting that HIV-1 neurotoxicity can be initiated by the viral protein gp120 binding to neuronal chemokine receptors. Our data suggest that HIV-1 neurotoxicity can be initiated by the viral protein gp120 binding to neuronal CXCR4 and that these two strains exhibit a different neurotoxic profile as well as distinct molecular mechanisms. Reference(s) [1] Bachis, A., Major, E.O., Mocchetti, I., 2003, Brain-derived neurotrophic factor inhibits human immunodeficiency virus-1/gp120-mediated cerebellar granule cell death by preventing gp120 internalization. J Neurosci Jul 2; 23(13): 5715–5722.
Posters P.1.01 An association of the glial cell linederived neurotrophic factor family receptor alpha-1 gene with schizophrenia A. Alkelai1 ° , Y. Kohn1 , S. Lupoli2 , T. Olender3 , K. SarnerKanyas1 , E. Ben-Asher3 , D. Lancet3 , F. Macciardi2 , B. Lerer1 . 1 Hadassah-Hebrew University Medical Center, Psychiatry, Jerusalem, Israel; 2 University of Milan, Department of Sciences and Biomedical Technologies, Milan, Italy; 3 Weizmann Institute of Science, Department of Molecular Genetics, Rehovot, Israel Purpose of the study: Schizophrenia is a severe, chronic neuropsychiatric disorder with poorly defined etiology and pathophysiology. No diagnostic neuropathology has
S3
been identified and available treatment is symptomatic and only partially successful. There is strong support for a significant role of genetic factors in the etiology of schizophrenia. Identifying schizophrenia related genes an important priority since it will help to understand the pathophysiology of the disorder and give provide the prospect of more effective treatment. A genome scan for schizophrenia related loci in Arab Israeli families by Lerer et al. [1] detected significant evidence for linkage at chromosome 6q23. Association [2] and replication studies [3] identified AHI1 as a susceptibility gene. The same genome scan revealed suggestive evidence for a schizophrenia susceptibility locus in the 10q23−26 region. Since the mode of inheritance of schizophrenia is most likely polygenic, multiple genes may be involved in its pathogenesis even in an ethnically homogeneous population. Expanding the initial phase of the project beyond the 6q23 region, we sought further evidence for linkage of 10q23-q26 with schizophrenia, set out to refine this candidate region, tested for possible genetic interaction with the 6q23 region and sought specific susceptibility gene(s). Methods: To refine the candidate region and test for genetic interaction, 35 additional microsatellite markers were genotyped on an ABI3700 Automated DNA Analyzer (Applied Biosystems) in the 10q23-q26 candidate region in the same samples that were used for the whole genome scan [1]. Further single-locus and two-locus linkage analyses were performed with GENEHUNTER software. To seek genes associated with schizophrenia in the refined 10q24.33-q26.13 region a total of 2046 SNP were genotyped by using of the Illumina’s HumanCNV370Duo BeadChip and analyzed with PBAT Version 3.6. The family sample that was genotyped was the Expanded Arab Israeli Family Sample previously described by AmannZalcenstein et al. [2]. Results: A final candidate region of 19.9Mb was found between markers D10S222 and D10S587 (10q24.33q26.13) by nonparametric and parametric singe-locus linkage analyses which revealed a suggestive MOD score of 2.56 and an NPL of 3.34 (p = 0.0006). Twolocus multipoint linkage analysis yielded a significant LOD score of 7.45 under the multiplicative model, suggesting overlapping function of the genes at these two loci. Significance was assessed by permutation analysis. Extensive SNP genotyping of the refined candidate region revealed two SNPs associated with schizophrenia after Bonferroni correction for multiple testing (2046 tests, p = 0.000024), the most intriguing of which resides in the GFRA1 gene which encodes the glial cell line-derived neurotrophic factor (GDNF) family receptor alpha-1. Conclusions: This study suggests overlapping function of genes at the 6q23 and refined 10q23-q26 loci and shows
S4
Molecular neuropsychopharmacology
an association of GFRA1 gene with schizophrenia. The protein encoded by GFRA1 is a member of the GDNF receptor family. It is a receptor for both GDNF and neurturin (NTN). GDNF and NTN are two structurally related, potent neurotrophic factors that play key roles in the control of neuron survival and differentiation. These roles are potentially relevant to the pathogenesis of schizophrenia. Reference(s) [1] Lerer, B., Segman, R.H., Hamdan, A., Kanyas, K., Karni, O., Kohn, Y., Korner, M., Lanktree, M., Kaadan, M., Turetsky, N., Yakir, A., Kerem, B., Macciardi, F., 2003, Genome scan of Arab Israeli families maps a schizophrenia susceptibility gene to chromosome 6q23 and supports a locus at chromosome 10q24. Mol Psychiatry 8: 488–498. [2] Amann-Zalcenstein, D., Avidan, N., Kanyas, K., Ebstein, R.P., Kohn, Y., Hamdan, A., Ben-Asher, E., Karni, O., Mujaheed, M., Segman, R.H., Maier, W., Macciardi, F., Beckmann, J.S., Lancet, D., Lerer, B., 2006, AHI1, a pivotal neurodevelopmental gene, and C6orf217 are associated with susceptibility to schizophrenia. Eur J Hum Genet 14: 1111–1119. [3] Ingason, A., Sigmundsson, T., Steinberg, S., Sigurdsson, E., Haraldsson, M., Magnusdottir, B.B., Frigge, M.L., Kong, A., Gulcher, J., Thorsteinsdottir, U., Stefansson, K., Petursson, H., Stefansson, H., 2007, Support for involvement of the AHI1 locus in schizophrenia. Eur J Hum Genet 15, 988–991. P.1.02 M- and T-tropic HIVs cause apoptosis in rat neuronal cell culture F. Biggio1 ° , A. Bachis2 , I. Mocchetti2 . 1 University of Cagliari, Department of Exp. Biology, Monserrato (Cagliari), Italy; 2 Georgetown University, Department of Neuroscience, Washington D.C., USA Human immunodeficiency virus type 1 (HIV-1) causes neuronal degeneration and, at a late stage, creates HIV-associated dementia (HAD) and other neurological abnormalities characterized by a severe loss of neurons and fibers throughout the brain that involved a broad spectrum of motor impairments and cognitive deficits. Neuronal loss, reactive astrocytes and other abnormalities are seen in the brain of individuals with Acquired Immune Deficiency Syndrome (AIDS)-associated Dementia Complex (ADC). A better understanding of the pathogenic mechanisms mediating HIV-1 neurotoxicity is vital for developing effective neuro-protective therapies against ADC. HIV-1 is believed to be the main agent causing ADC. However, little
is known about the molecular and cellular mechanisms of HIV-1 neurotoxicity considering that HIV-1 does not infect post-mitotic neurons and that viral load does not necessarily correlate with ADC. In the brain HIV-1 infects macrophages and lymphocyties after binding of the envelope glycoprotein 120 (gp-120) to CD4 in conjunction with several chemokine receptors family, such as CCR5 and CXCR4. These receptors act as co-receptors for viral entry and, consequently, regulate the lethal infection of the cells. Most HIV strains use CCR5 for HIV transmission and infection. Moreover progeny HIV-1 virions can also cause neuronal death. Several viral proteins, such as the envelope protein gp120 and the transcription activator Tat, have been shown to induce neuronal apoptosis through direct and indirect mechanisms both in vitro and in vivo. However, it has not been fully established yet whether HIV-1 promotes neuronal apoptosis by a direct mechanism. We have previously demonstrate that gpIIIB, a tropic (X4) strain, causes neuronal apoptosis through activation of the chemokine receptor CXCR4 both in vitro and in vivo. Our experiments were performed in Cortical neurons and Cerebellar granule cells prepared from 1 day old and 8 days old Sprague Dawley rat pups, respectively. To explore the neurotoxic effect of HIV-1, we exposed rat cerebellar granule cells and cortical neurons in culture to two different strains of HIV-1, IIIB and BaL, T- and M-tropic strains that utilize CXCR4 and CCR5 co-receptors, respectively, to infect cells. We observed that both viruses elicit a time-dependent apoptotic cell death in these cultures without inducing a productive infection as determined by the absence of the core protein of HIV-1, p24, in cell lysates. Instead, neurons were gp120 positive, suggesting that the envelope protein is shed by the virus and then subsequently internalized by neurons. The CXCR4 receptor antagonist AMD3100 or the CCR5 receptor inhibitor D-Ala-peptide T-amide blocked HIV IIIB and HIV Bal neurotoxicity, respectively. In contrast, the NMDA receptor blocker MK801 failed to protect neurons from HIV-mediated apoptosis, suggesting that HIV-1 neurotoxicity can be initiated by the viral protein gp120 binding to neuronal chemokine receptors. Our data suggest that HIV-1 neurotoxicity can be initiated by the viral protein gp120 binding to neuronal CXCR4 and that these two strains exhibit a different neurotoxic profile as well as distinct molecular mechanisms. Reference(s) [1] Bachis, A., Major, E.O., Mocchetti, I., 2003, Brain-derived neurotrophic factor inhibits human immunodeficiency virus-1/gp120-mediated cerebellar granule cell death by preventing gp120 internalization. J Neurosci Jul 2; 23(13): 5715–5722.