Neuregulin 1 and susceptibility to schizophrenia

Abstract

The cause of schizophrenia is unknown, but it has a significant genetic component. Pharmacologic studies, studies of gene expression in man, and studies of mouse mutants suggest involvement of glutamate and dopamine neurotransmitter systems. However, so far, strong association has not been found between schizophrenia and variants of the genes encoding components of these systems. Here, we report the results of a genomewide scan of schizophrenia families in Iceland; these results support previous work, done in five populations, showing that schizophrenia maps to chromosome 8p. Extensive fine-mapping of the 8p locus and haplotype-association analysis, supplemented by a transmission/disequilibrium test, identifies neuregulin 1 (NRG1) as a candidate gene for schizophrenia. NRG1 is expressed at central nervous system synapses and has a clear role in the expression and activation of neurotransmitter receptors, including glutamate receptors. Mutant mice heterozygous for either NRG1 or its receptor, ErbB4, show a behavioral phenotype that overlaps with mouse models for schizophrenia. Furthermore, NRG1 hypomorphs have fewer functional NMDA receptors than wild-type mice. We also demonstrate that the behavioral phenotypes of the NRG1 hypomorphs are partially reversible with clozapine, an atypical antipsychotic drug used to treat schizophrenia.

Figure 1

Genomewide scan of 110 patients with schizophrenia from 33 families. The 110 patients included 106 who fulfilled the Research Diagnostic Criteria (RDC) schizophrenia diagnosis, three patients diagnosed with unspecified functional psychosis, and one diagnosed with schizoaffective disorder. With the assistance of our genealogical database, the patients were clustered into 33 families, and relations between patients are as distant as second cousins. The patients and their relatives were genotyped using a framework set of 950 microsatellite markers. The X-axis gives the genetic distance (in cM) along the chromosome, and the Y-axis gives the LOD score. The data were analyzed using multipoint allele-sharing, without specification of an inheritance model, using Allegro (Gudbjartsson et al. 2000).

Figure 2

Fine mapping of the schizophrenia locus on chromosome 8p, with 50 additional markers mapped to the 30-cM locus. Since correct marker order and genetic distance is essential for multipoint genetic analysis, we determined the order of markers by generating a physical map of 30 cM in 8p12-p21, using BAC clones, and we determined the genetic distances on our high-resolution genetic map, on the basis of 1,200 meiotic events (Kong et al. 2002). The solid line represents the allele-sharing LOD score, and the dashed line represents the parametric LOD score. The additional markers increased the information content from 0.7 to >0.9. The X-axis gives the genetic distance (in cM) along the chromosome, and the Y-axis gives the LOD score.

Figure 3

Microsatellite at-risk haplotypes on chromosome 8p and known genes within a region showing haplotype sharing between 9 of the 33 linkage families. Extensive sharing of two microsatellite haplotypes between patients from the linkage families is shown at the top. Haplotypes were reconstructed by application of the Allegro program (Gudbjartsson et al. 2000), using available family members to derive the phase. Key markers in the haplotypes are shown, and the size of the region is indicated. Families carrying the haplotypes are labeled F1–F9, and the number of affected individuals in each family carrying that haplotype is given in parentheses. Maximum haplotype sharing between families is 9.5 Mb for haplotype I and 11.4 Mb for haplotype II. Shared haplotypes between families narrow the region of interest to 600 kb between markers 29H12-7320 and 473C15-439, indicated by a bar (microsatellite haplotype I). The location of a BAC contig covering 1.5 Mb of the locus region is indicated. The sequence of GS1-57G24 was obtained from the public domain (GenBank accession number AF128834), but we sequenced the other BACs shown. The positions of NRG1 and an EST cluster of unknown function (Hs.97362) are schematically shown in relation to the BACs. Exons are indicated by vertical bars.

Figure 4

Microsatellite and SNP at-rick haplotypes at the 5′ end of the NRG1 gene. Four haplotypes, defined by 12 SNPs and four microsatellite markers, were individually found in excess in the schizophrenia patients with similar relative risk. The common core for these haplotypes, defined by five SNPs and two microsatellite markers, is shown at the bottom. The frequencies for each haplotype in all affected individuals, independent affected individuals, and control individuals are indicated in the panel on the right. The only known exon within the boundaries of the core at-risk haplotype is the 5′ exon of GGF2, as indicated in the figure. The distance between the markers flanking possible recombination breakpoints (arrows) is 290 kb, corresponding to 0.16–0.45 Mb on the scale at the bottom of figure 3.

Figure 5

Locomotion and PPI in NRG1 and ErbB4 hypomorphic mice. NRG1 and ErbB4 hypomorphic mice were significantly more active, according to different measures that reflect locomotion and exploration. Here, we show distance traveled in a novel open-field test for the two lines of mice: NRG1 hypomorphic mice (A) and ErbB4 hypomorphic mice (B). Data have been binned into 5-min intervals over the 30-min observation period. Distance traveled was significantly increased in both NRG1 and ErbB4 male mice in comparison to litter-mate control mice (n=21 NRG1 mice and 22 litter-mate control mice, P=.04; n=39 ErbB4 mice and 22 litter-mate control mice, P=.03). Open-field activity was monitored as described. Error bars indicate the SEM. A, insert, hyperactivity of the NRG1 mutant mice was reversed by clozapine. The NRG1 mice and litter-mate control mice were injected i.p. with either clozapine (1 mg/kg) or vehicle, 25 min prior to testing (n=10 NRG1 mice and 10 control mice, P=.02). Open-field activity was monitored as described in the “Subjects and Methods” section. PPI was recorded for NRG1 (C) and ErbB4 (D) hypomorphic mice, using a conditioning, prepulse noise burst of 72, 74, or 78 dB. NRG1 hypomorphic mice have impaired PPI in comparison to normal, litter-mate control mice (F-test, P=.03), whereas ErbB4 mutant mice were not significantly impaired (F-test, P=.056).