Title:
Genomic and animal model approaches to schizophrenia susceptibility

Yasuyuki Fukumaki
Division of Human Molecular Genetics, Research Center for Genetic Information, Medical Institute of Bioregulation, Kyushu University , Fukuoka 812-8582, Japan

Abstract:
Schizophrenia is a devastating neuropsychiatric disorder with a multifactorial background. To search for schizophrenia susceptibility loci, we have been taking three approaches as follows.
(I) Candidate gene approach: i) Based on the glutamatergic dysfunction hypothesis for schizophrenia pathophysiology, we conducted a systematic study of associations of glutamate receptor genes and glutamate transporter genes with schizophrenia. We selected a number of common SNPs to cover the entire gene region with linkage disequilibrium. We found associations of GRIN2D, GRIA4, GRM3, GRM8 , SLC1A2, SLC1A6 and SLC6A5 with schizophrenia. ii) Since schizophrenia-like symptoms are produced by administration of phencyclidine (PCP), a noncompetitive antagonist of NMDA receptors. PCP-responsive genes could be involved in pathophysiology of schizophrenia. We analyzed gene expression of brain tissues of rats treated by PCP administration, on the AGILENT microarray platform. We identified 7 genes showing > 2-fold expressional change evaluated by qRT-PCR, as subjects for the locus-wide association study. We observed haplotype associations in PDE4A and PLAT and single-point associations of one SNP inPDE4A and two SNPs in PLAT .
(II) Genome-wide approach: We conducted the case-control study using about 30,000 STR markers distributed across the entire genome. To overcome the type I error due to multiple tests, we planed three sequential steps of screening using independent pooling sample sets. After the third screening step, 374 markers remained significantly associated with the disorder. Fifty-nine of them were selected for SNP typing through evaluation of reproducibility of associated alleles for each marker. We individually genotyped all screening samples using a total of 1,536 tag SNPs located in the vicinity of ~200 kb from the 59 positive microsatellite markers. Out of 167 SNPs that replicated the significance, we selected 31 SNPs based on the levels of P values for the confirmatory association test using an independent large sample set. Finally one SNP located within C2orf24 , and two SNPs in the downstream region of the gene showed significant association with the disorder, suggesting that C2orf24 is a candidate for schizophrenia susceptibility in the Japanese population.
(III) Model animal approach: GRM3 showing significant association with schizophrenia in the first approach is of interest because LY354740, a highly selective and potent agonist of group II mGluR (mGluR2/3) attenuates schizophrenia-like symptoms induced by administration of PCP. To explore functional roles of GRM3 in schizophrenia pathogenesis, we conducted comprehensive behavioral analyses of mGluR3 knockout (KO) mice. The KO mice showed hyperactivity in the open field and light/dark transition tests , impaired reference memory for stressful events in the Porsolt forced swim test and impaired working memory in the T-maze forced alternation task test . The hyperactivity and impaired working memory are known as endophenotypes of schizophrenia. We performed in vivo microdialysis measurements of extracellular dopamine in the central nervous system of the KO mice. We observed enhancement of methamphetamine-induced release of dopamine in the nucleus accumbens of the KO mice, suggesting that GRM3 may be involved in schizophrenia pathogenesis through the dopaminergic and glutamatergic interaction . Combination of genomic and animal model approaches could elucidate susceptibility genes for schizophrenia.

References:
Deng, X., Takaki, H., Wang, L., Kuroki, T., Nakahara, T., Hashimoto, K., Ninomiya, H., Arinami, T., Inada, T., Ujike, H., Itokawa, M., Tochigi, M., Watanabe, Y., Someya, T., Kunugi, H., Iwata, N., Ozaki, N., Shibata, H., Fukumaki, Y.: Positive association of phencyclidine-responsive genes, PDE4A andPLAT , with schizophrenia.
Am J Med Genet B Neuropsychiatr Genet. 156B: 850-858, 2011.

Shibata, H, Yamamotob, K., Suna, Z., Okac, A., Inokoc, H., Arinamid,T., Inadae, T., Ujike, H., Itokawag, M., Tochigih, M., Watanabei,Y., Someyai, T., Kunugij, H., Suzukik, T., Iwatak, N., Ozakil, N., Fukumaki, Y.: Genome-wide association study of schizophrenia using microsatellite markers in the Japanese population. Psychiatr Genet. 2012 in press.

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