Amygdala and anterior cingulate transcriptomes from individuals with bipolar disorder reveal downregulated neuroimmune and synaptic pathways

Abstract

Recent genetic studies have identified variants associated with bipolar disorder (BD), but it remains unclear how brain gene expression is altered in BD and how genetic risk for BD may contribute to these alterations. Here, we obtained transcriptomes from subgenual anterior cingulate cortex and amygdala samples from post-mortem brains of individuals with BD and neurotypical controls, including 511 total samples from 295 unique donors. We examined differential gene expression between cases and controls and the transcriptional effects of BD-associated genetic variants. We found two coexpressed modules that were associated with transcriptional changes in BD: one enriched for immune and inflammatory genes and the other with genes related to the postsynaptic membrane. Over 50% of BD genome-wide significant loci contained significant expression quantitative trait loci (QTL) (eQTL), and these data converged on several individual genes, including SCN2A and GRIN2A. Thus, these data implicate specific genes and pathways that may contribute to the pathology of BP.

Figure 1.

Summary of FDR<5% significant differentially expressed features overall and by brain region. A total of 25,136 genes, 73,214 transcripts, 396,818 exons, and 266,197 junctions were tested in the sACC and amygdala. Results shown include: a) Venn diagram of the overlap in the number of unique genes with a significant feature by brain region; b) breakdown of gene types for the unique set of genes represented in a); and c) breakdown of the significant features (gene, transcript, exon or junction) in the implicated genes.

Figure 2.

Results of WGCNA analysis: a) boxplot of red module eigengene values for each sample by case-control status controlling for age and brain region; b) top 15 enriched pathways in the red module containing genes with FDR<5% significant differentially expressed features; c) volcano plot of fold change in gene level expression by −log10pvalue for genes in the red module, with those genes in the top 15 pathways that have a significant differentially expressed features highlighted in red (note some may have differentially expressed features other than at the gene level which is shown here); d) boxplot of pink module eigengene values for each sample by case-control status, controlling for age and brain region; e) top 15 enriched pathways in the pink module containing genes with FDR<5% significant differentially expressed features; and f) volcano plot of fold change in gene level expression by −log10pvalue for genes in the pink module, with those genes in the top 15 pathways that have a significant differentially expressed features highlighted in red (note some may have differentially expressed features other than at the gene level which is shown here). Boxplots in a) and d) show data on 126 BD cases and 142 controls in the sACC (green circles) together with 121 BD cases and 122 controls in the amygdala (blue circles). The boxplots display the median as the center line, the interquartile range (IQR; 25^th – 75^th percentile) as the box range, and 1.5 times the IQR as the whiskers unless a minimum/maximum is reached.

Figure 3.

Summary of FDR<1% expression (eQTL) and splicing (sQTL) quantitative trait loci in loci suggested by the latest Psychiatric Genomics Consortium (PGC) genome-wide association study of bipolar disorder. Tests were carried out for QTL associations between 10,777 SNPs and gene (n=4,647), transcript (n=14,434), exon (n=76,589) and junction (n=49,188) features in these loci, and results summarized for the lead SNP in each locus. a) Overlap of loci with significant eQTL/sQTL by brain region among the 30 genome-wide significant loci. b) and c) Breakdown of gene types for the implicated genes in a) and the features that are associated in the genome-wide significant loci. d) Overlap of loci with significant eQTL/sQTL by brain region among the 850 suggestive genome-wide loci. e) and f) Breakdown of gene types for the implicated genes in d) and the features that are associated in these loci.

Figure 4.

Gene visualization plots and accompanying SNP-feature scatterplots for the lead SNP with significant e/sQTLs (FDR<1%). The gene visualization plots show the location of the lead SNP in blue and exon/intron models in black for all protein coding transcripts and the union gene model for each gene. Specific gene features are shown in red if they are down-regulated in the e/sQTL and in green if they are up-regulated. Red/green highlighted boxes over introns represent down/up regulated junctions between corresponding exons. The SNP-feature boxplots show residualized expression levels (from the eQTL linear regression models described in the Methods) of the most significant feature with the lead SNP, with SNP genotypes shown from most to least common: a) SCN2A (β=−1.12, p=5.08×10⁻²⁹) and b) GRIN2A (β=−0.07, p=1.32×10⁻⁵). Boxplots for a) and b) show results based on 262 samples (126 BD cases and 142 controls) from the sACC and display the median as the center line, the IQR as the box range, and 1.5 times the IQR as the whiskers unless a minimum/maximum is reached.

Figure 5.

Scatterplot of Z-score test statistics from TWAS of genes in the sACC versus amygdala. A total of 13,822 genes were tested across both brain regions. Points highlighted in colors are FDR<5% significant in the amygdala (n=125), sACC(n=156), or both regions (n=34). Points along the X=0 or Y=0 axes were not estimated in the amygdala or sACC, respectively, typically because heritability estimates failed in one of the regions and subsequent models could not be estimated.