Loss of nuclear factor E2-related factor 1 in the brain leads to dysregulation of proteasome gene expression and neurodegeneration.

The ubiquitin-proteasome pathway plays an important role in the pathogenesis of neurodegeneration, but mechanisms controlling expression of components in this pathway remain poorly understood. Nuclear factor E2-related factor 1 (Nrf1) transcription factor has been shown to regulate expression of antioxidant and cytoprotective genes. To determine the function of Nrf1 in the brain, mice with a late-stage deletion of Nrf1 in neuronal cells were generated. Loss of Nrf1 leads to impaired proteasome function and neurodegeneration. Gene expression profiling and RT-PCR analysis revealed a coordinate down-regulation of various proteasomal genes including PsmB6, which encodes a catalytic subunit of the proteasome. Transcriptional analysis and chromatin immunoprecipitation experiments demonstrated that PsmB6 is an Nrf1 target gene. These findings reveal Nrf1 as a key transcriptional regulator required for the expression of proteasomal genes in neurons and suggest that perturbations of Nrf1 function may contribute to the pathogenesis of neurodegenerative diseases.

The first decade and beyond of transcriptional profiling in schizophrenia.

Gene expression changes in brains of individuals with schizophrenia (SZ) have been hypothesized to reflect possible pathways related to pathophysiology and/or medication. Other factors having robust effects on gene expression profiling in brain and possibly influence the schizophrenia transcriptome such as age and pH are examined. Pathways of curated gene expression or gene correlation networks reported in SZ (white matter, apoptosis, neurogenesis, synaptic plasticity, glutamatergic and GABAergic neurotransmission, immune and stress-response, mitochondrial, and neurodevelopment) are not unique to SZ and have been associated with other psychiatric disorders. Suggestions going forward to improve the next decade of profiling: consider multiple brain regions that are carefully dissected, release large datasets from multiple brain regions in controls to better understand neurocircuitry, integrate genetics and gene expression, measure expression variants on genome wide level, peripheral biomarker studies, and analyze the transcriptome across a developmental series of brains. Gene expression, while an important feature of the genomic landscape, requires further systems biology to advance from control brains to a more precise definition of the schizophrenia interactome.

Chronic antipsychotic treatment exerts limited effects on the mania-like behavior of dopamine transporter knockdown mice.

BACKGROUND: Bipolar disorder is a life-threatening disorder linked to dopamine transporter (DAT) polymorphisms, with reduced DAT levels seen in positron emission tomography and postmortem brains. AIMS: The purpose of this study was to examine the effects of approved antipsychotics on DAT dysfunction-mediated mania behavior in mice. METHODS: DAT knockdown mice received either D2-family receptor antagonist risperidone or asenapine and mania-related behaviors were assessed in the clinically-relevant behavioral pattern monitor to assess spontaneous exploration. RESULTS: Chronic risperidone did not reverse mania-like behavior in DAT knockdown mice. Chronic asenapine reduced mania behavior but this effect was more pronounced in wild-type littermates than in DAT knockdown mice. CONCLUSION: Taken together, these findings suggest that while acute antipsychotic treatment may be beneficial in management of bipolar mania, more targeted therapeutics may be necessary for long-term treatment. Specific investigation into DAT-targeting drugs could improve future treatment of bipolar mania.

Analysis of whole genome biomarker expression in blood and brain.

The consistency of peripheral gene expression data and the overlap with brain expression has not been evaluated in biomarker discovery, nor has it been reported in multiple tissues from the same subjects on a genome wide transcript level. The effects of processing whole blood, transformation, and passaged cell lines on gene expression profiling was studied in healthy subjects using Affymetrix arrays. Ficoll extracted peripheral blood mononuclear cells (PBMCs), Epstein-Barr virus (EBV) transformed lymphocytes, passaged lymphoblastic cell lines (LCLs), and whole blood from Tempus tubes were compared. There were 6,813 transcripts differentially expressed between different methods of blood preparation. Principal component analysis resolved two partitions involving pre- and post-transformation EBV effects. Combining results from Affymetrix arrays, postmortem subjects' brain and PBMC profiles showed co-expression levels of summarized transcripts for 4,103 of 17,859 (22.9%) RefSeq transcripts. In a control experiment, rat hemi-brain and blood showed similar expression levels for 19% of RefSeq transcripts. After filtering transcripts that were not significantly different in abundance between human cerebellum and PBMCs from the Affymetrix exon array the correlation in mean transcript abundance was high as expected (r = 0.98). Differences in the alternative splicing index in brain and blood were found for about 90% of all transcripts examined. This study demonstrates over 4,100 brain transcripts co-expressed in blood samples can be further examined by in vitro and in vivo experimental studies of blood and cell lines from patients with psychiatric disorders.

Spatial relationship between synapse loss and beta-amyloid deposition in Tg2576 mice.

Although there is evidence that beta-amyloid impairs synaptic function, the relationship between beta-amyloid and synapse loss is not well understood. In this study we assessed synapse density within the hippocampus and the entorhinal cortex of Tg2576 mice at 6-18 months of age using stereological methods at both the light and electron microscope levels. Under light microscopy we failed to find overall decreases in the density of synaptophysin-positive boutons in any brain areas selected, but bouton density was significantly decreased within 200 mum of compact beta-amyloid plaques in the outer molecular layer of the dentate gyrus and Layers II and III of the entorhinal cortex at 15-18 months of age in Tg 2576 mice. Under electron microscopy, we found overall decreases in synapse density in the outer molecular layer of the dentate gyrus at both 6-9 and 15-18 months of age, and in Layers II and III of the entorhinal cortex at 15-18 months of age in Tg 2576 mice. However, we did not find overall changes in synapse density in the stratum radiatum of the CA1 subfield. Furthermore, in the two former brain areas we found a correlation between lower synapse density and greater proximity to beta-amyloid plaques. These results provide the first quantitative morphological evidence at the ultrastructure level of a spatial relationship between beta-amyloid plaques and synapse loss within the hippocampus and the entorhinal cortex of Tg2576 mice.

Independent quantitative trait loci influence ventral and dorsal hippocampal volume in recombinant inbred strains of mice.

Anatomical and functional studies support segregation of the hippocampus into ventral and dorsal components along its septotemporal axis. However, it is unknown whether the development of these two components of the hippocampus is influenced by common or separate genetic factors. In this study, we used recombinant inbred strains of mice to determine whether the same or different quantitative trait loci (QTL) influence ventral and dorsal hippocampal volume. Using two sets of strains of recombinant inbred mice (BXD and AXB/BXA), we identified separate QTLs for ventral and dorsal hippocampal volume. In BXD mice, suggestive QTLs for ventral hippocampus were identified on chromosomes 2, 8 and 13, and a significant QTL for dorsal hippocampal volume was identified on chromosome 15. There was also a suggestive QTL for dorsal hippocampal volume on chromosome 13. In AXB/BXA mice, there were no significant or suggestive QTLs for ventral hippocampal volume, but a significant QTL for dorsal hippocampus was identified on chromosome 5. These findings suggest that the development of the ventral and dorsal components of the hippocampus is influenced by separate genetic loci.

Neuroprotective effects of olanzapine in a rat model of neurodevelopmental injury.

Recent clinical studies have suggested that treatment with atypical antipsychotic drugs, such as olanzapine, may slow progressive changes in brain structure in patients with schizophrenia. To investigate the possible neural basis of this effect, we sought to determine whether treatment with olanzapine would inhibit the loss of hippocampal neurons associated with the administration of the excitotoxin, kainic acid, in neonatal rats. At post-natal day 7 (P7), rats were exposed to kainic acid via intracerebroventricular administration. Neuronal loss within the CA2 and CA3 subfields of the hippocampus and neurogenesis within the dentate gyrus of the hippocampus were then assessed at P14 by Fluoro-Jade B and BrdU labeling, respectively. Daily doses of olanzapine (2, 6, or 12 mg/day), haloperidol (1.2 mg/kg), melatonin (10 mg/kg), or saline were administered between P7 and P14. Melatonin is an anti-oxidant drug and was included in this study as a positive control, since it has been observed to have neuroprotective effects in a variety of animal models. The highest dose of olanzapine and melatonin, but not haloperidol, ameliorated the hippocampal neuronal loss triggered by kainic acid administration. However, drug administration did not have a significant effect on the rate of neurogenesis. These results suggest that olanzapine has neuroprotective effects in a rat model of neurodevelopmental insult, and may be relevant to the observed effects of atypical antipsychotic drugs on brain structure in patients with schizophrenia.

Acetylcholinesterase inhibitors ameliorate behavioral deficits in the Tg2576 mouse model of Alzheimer's disease.

RATIONALE: Acetylcholinesterase inhibitors are widely used for the treatment of patients with Alzheimer's disease (AD). However, the relationship between the capacity of such drugs to ameliorate the symptoms of AD and their ability to alter the underlying disease process is not well understood. Transgenic mice that overexpress the human form of amyloid precursor protein and develop deposits of beta-amyloid (Abeta) and behavioral deficits during adulthood are useful for investigating this question. OBJECTIVES: The effects of administration of two acetylcholinesterase inhibitors, physostigmine and donepezil, on Abeta plaque formation and memory-related behaviors were investigated in the Tg2576-transgenic mouse model of AD. At 9-10 months of age, Tg2576-transgenic [Tg(+)] mice develop Abeta plaques and impairments on paradigms related to learning and memory as compared to transgene-negative [Tg(-)] mice. METHODS: Beginning at 9 months of age, increasing doses of physostigmine (0.03, 0.1, and 0.3 mg/kg), donepezil (0.1, 0.3, and 1.0 mg/kg), or saline were administered over 6 weeks to cohorts of Tg(+) and Tg(-) mice. Performance on tests of spatial reversal learning and fear conditioning was evaluated at each drug dose throughout the period of drug administration. After drug administration was completed, the animals were sacrificed and Abeta plaque number was quantified. RESULTS: Administration of physostigmine and donepezil improved deficits in contextual and cued memory in Tg(+) mice so that their behaviors became more similar to Tg(-) mice. However, administration of physostigmine and donepezil tended to improve cued memory and deficits in spatial learning in both Tg(+) and Tg(-) mice. Physostigmine administration demonstrated more prominent effects in improving contextual memory than donepezil, while donepezil was more effective than physostigmine in improving deficits in the acquisition of the spatial memory paradigm. Administration of neither drug altered the deposition of Abeta plaques. CONCLUSIONS: These studies suggest that acetylcholinesterase inhibitors can ameliorate memory deficits in Tg(+) mice without necessarily altering the deposition of Abeta plaques. Tg2576 mice may be useful as an animal model to further investigate the mechanisms by which aceytlcholinesterase inhibitors improve cognitive deficits in patients with AD.

Low dose quetiapine reverses deficits in contextual and cued fear conditioning in rats with excitotoxin-induced hippocampal neuropathy.

Previous studies have demonstrated that adult rats with excitotoxic lesions of the hippocampus display deficits in memory-related behaviors similar to the memory deficits associated with schizophrenia. In this study, we assessed the sub-chronic effects of quetiapine, risperidone and haloperidol on performance deficits after intracerebroventricular administration of the excitotoxin, kainic acid, using paradigms for contextual and cued fear conditioning and spatial reversal learning in rats. The effects of three doses of quetiapine (5, 10 and 20 mg/kg) and single doses of risperidone (0.5 mg/kg) and haloperidol (0.15 mg/kg) were compared. Quetiapine administration at the lowest dose (5 mg/kg) reversed deficits in contextual and cued fear conditioning, but not deficits in spatial reversal learning, in kainic acid-treated animals. However, the two higher doses of quetiapine, and the single doses of risperidone and haloperidol, did not reverse any of the kainic acid-induced behavioral deficits. These results may be relevant to the effects of quetiapine and other antipsychotic drugs on memory deficits in patients with schizophrenia.

Olanzapine Reversed Brain Gene Expression Changes Induced by Phencyclidine Treatment in Non-Human Primates.

The NMDA receptor antagonist phencyclidine (PCP) creates schizophrenia-like symptoms in normal controls. The effect of PCP on non-human primate brain gene expression was examined and compared to changes induced by olanzapine treatment. Experimental studies of PCP and antipsychotic drugs have direct relevance to understanding the patho-physiology and treatment of schizophrenia. Genome-wide changes in prefrontal cortex gene expression revealed alterations of 146 transcripts in the PCP treatment group compared to vehicle controls. Dysregulated genes were enriched in identified classes implicated in neurological and genetic disorders, including schizophrenia genes from the Psychiatric Genomics Consortium 108 loci as well as cell death in PCP-treated primates. Canonical pathway analysis revealed a significant overrepresentation of several groups including synaptic long-term potentiation and calcium signaling. Olanzapine coadministered with PCP normalized 34% of the 146 PCP-induced probe set expression changes, and a network of 17 olanzapine-normalized genes was identified enriched in schizophrenia candidate genes containing RGS4, SYN1 and AKT as nodes. The results of this study support the use of PCP administration in non-human primates as a glutamatergic model of schizophrenia and suggest that a large number of PCP-induced expression differences can be reversed by olanzapine. The results of this study may be informative for identification of potential candidates for pharmacogenetics and biomarker research related to the treatment of schizophrenia.

Coding and noncoding gene expression biomarkers in mood disorders and schizophrenia.

Mood disorders and schizophrenia are common and complex disorders with consistent evidence of genetic and environmental influences on predisposition. It is generally believed that the consequences of disease, gene expression, and allelic heterogeneity may be partly the explanation for the variability observed in treatment response. Correspondingly, while effective treatments are available for some patients, approximately half of the patients fail to respond to current neuropsychiatric treatments. A number of peripheral gene expression studies have been conducted to understand these brain-based disorders and mechanisms of treatment response with the aim of identifying suitable biomarkers and perhaps subgroups of patients based upon molecular fingerprint. In this review, we summarize the results from blood-derived gene expression studies implemented with the aim of discovering biomarkers for treatment response and classification of disorders. We include data from a biomarker study conducted in first-episode subjects with schizophrenia, where the results provide insight into possible individual biological differences that predict antipsychotic response. It is concluded that, while peripheral studies of expression are generating valuable results in pathways involving immune regulation and response, larger studies are required which hopefully will lead to robust biomarkers for treatment response and perhaps underlying variations relevant to these complex disorders.

Mitochondrial mutations and polymorphisms in psychiatric disorders.

Mitochondrial deficiencies with unknown causes have been observed in schizophrenia (SZ) and bipolar disorder (BD) in imaging and postmortem studies. Polymorphisms and somatic mutations in mitochondrial DNA (mtDNA) were investigated as potential causes with next generation sequencing of mtDNA (mtDNA-Seq) and genotyping arrays in subjects with SZ, BD, major depressive disorder (MDD), and controls. The common deletion of 4,977 bp in mtDNA was compared between SZ and controls in 11 different vulnerable brain regions and in blood samples, and in dorsolateral prefrontal cortex (DLPFC) of BD, SZ, and controls. In a separate analysis, association of mitochondria SNPs (mtSNPs) with SZ and BD in European ancestry individuals (n = 6,040) was tested using Genetic Association Information Network (GAIN) and Wellcome Trust Case Control Consortium 2 (WTCCC2) datasets. The common deletion levels were highly variable across brain regions, with a 40-fold increase in some regions (nucleus accumbens, caudate nucleus and amygdala), increased with age, and showed little change in blood samples from the same subjects. The common deletion levels were increased in the DLPFC for BD compared to controls, but not in SZ. Full mtDNA genome resequencing of 23 subjects, showed seven novel homoplasmic mutations, five were novel synonymous coding mutations. By logistic regression analysis there were no significant mtSNPs associated with BD or SZ after genome wide correction. However, nominal association of mtSNPs (p < 0.05) to SZ and BD were found in the hypervariable region of mtDNA to T195C and T16519C. The results confirm prior reports that certain brain regions accumulate somatic mutations at higher levels than blood. The study in mtDNA of common polymorphisms, somatic mutations, and rare mutations in larger populations may lead to a better understanding of the pathophysiology of psychiatric disorders.

Genetic influences on hippocampal structure and function in recombinant inbred mice.

Previously, we identified separate genetic influences on ventral versus dorsal hippocampal volume in BXD recombinant inbred mice [Martin MV, Dong HX, Vallera D, Lu L, Williams RW, Rosen GD, et al. Independent quantitative trait loci influence ventral and dorsal hippocampal volume in recombinant inbred strains of mice. Genes Brain Behav 2006;5:614-23]. Based on genotype at genetic markers associated with ventral hippocampal volume, we evaluated BXD mouse strains with relatively small versus large ventral hippocampal volumes using numerous behavioral paradigms known to rely upon hippocampal function and several other tasks that tap into behaviors analogous to those often impaired in schizophrenia. We observed a relationship between genotype at markers known to influence ventral hippocampal volume and working memory at an intermediate memory load. There was no association between genotype at markers known to influence ventral hippocampal volume and spatial reference memory, prepulse inhibition, or elevated plus maze performance. The relevance of these findings for understanding the pathophysiology of schizophrenia are discussed, including the possibility that genetic predisposition toward anterior hippocampal volume reductions and working memory deficits in schizophrenia may be related through a shared genetic locus.

Neuropsychological performance as endophenotypes in extended schizophrenia families from the Central Valley of Costa Rica.

OBJECTIVE: The understanding of complex heritable psychiatric disorders such as schizophrenia could be clarified by examining endophenotypes within genetically isolated populations, such as the one found in the Central Valley of Costa Rica. The reduction of familial variability within a sample could allow the relationship between the cognitive and symptomatic manifestations of the illness and the genetic underpinnings to become more observable. This study investigates the neuropsychological test performances of 41 family members from four extended multiplex families within the Spanish origin population of the Central Valley of Costa Rica as potential endophenotypes for genetic studies. METHODS: Individuals with a diagnosis of schizophrenia or schizoaffective disorder were compared with unaffected relatives and 15 unrelated controls with no family history of schizophrenia. RESULTS: Although the sample size is small, the results confirm previous reports in the literature of deficits in working memory, executive function, processing speed, and verbal fluency in individuals with schizophrenia compared with controls and intermediate performance in nonpsychotic family members compared with controls. We also found several suggestive quantitative cognitive trait loci with log of the odds greater than 1.75. CONCLUSION: These findings suggest that the cognitive deficits in schizophrenia are consistent aspects of the illness, although their usefulness as endophenotypes for genetic studies remains unclear.

Mitochondrial variants in schizophrenia, bipolar disorder, and major depressive disorder.

BACKGROUND: Mitochondria provide most of the energy for brain cells by the process of oxidative phosphorylation. Mitochondrial abnormalities and deficiencies in oxidative phosphorylation have been reported in individuals with schizophrenia (SZ), bipolar disorder (BD), and major depressive disorder (MDD) in transcriptomic, proteomic, and metabolomic studies. Several mutations in mitochondrial DNA (mtDNA) sequence have been reported in SZ and BD patients. METHODOLOGY/PRINCIPAL FINDINGS: Dorsolateral prefrontal cortex (DLPFC) from a cohort of 77 SZ, BD, and MDD subjects and age-matched controls (C) was studied for mtDNA sequence variations and heteroplasmy levels using Affymetrix mtDNA resequencing arrays. Heteroplasmy levels by microarray were compared to levels obtained with SNaPshot and allele specific real-time PCR. This study examined the association between brain pH and mtDNA alleles. The microarray resequencing of mtDNA was 100% concordant with conventional sequencing results for 103 mtDNA variants. The rate of synonymous base pair substitutions in the coding regions of the mtDNA genome was 22% higher (p = 0.0017) in DLPFC of individuals with SZ compared to controls. The association of brain pH and super haplogroup (U, K, UK) was significant (p = 0.004) and independent of postmortem interval time. CONCLUSIONS: Focusing on haplogroup and individual susceptibility factors in psychiatric disorders by considering mtDNA variants may lead to innovative treatments to improve mitochondrial health and brain function.

Exon expression in lymphoblastoid cell lines from subjects with schizophrenia before and after glucose deprivation.

BACKGROUND: The purpose of this study was to examine the effects of glucose reduction stress on lymphoblastic cell line (LCL) gene expression in subjects with schizophrenia compared to non-psychotic relatives. METHODS: LCLs were grown under two glucose conditions to measure the effects of glucose reduction stress on exon expression in subjects with schizophrenia compared to unaffected family member controls. A second aim of this project was to identify cis-regulated transcripts associated with diagnosis. RESULTS: There were a total of 122 transcripts with significant diagnosis by probeset interaction effects and 328 transcripts with glucose deprivation by probeset interaction probeset effects after corrections for multiple comparisons. There were 8 transcripts with expression significantly affected by the interaction between diagnosis and glucose deprivation and probeset after correction for multiple comparisons. The overall validation rate by qPCR of 13 diagnosis effect genes identified through microarray was 62%, and all genes tested by qPCR showed concordant up- or down-regulation by qPCR and microarray. We assessed brain gene expression of five genes found to be altered by diagnosis and glucose deprivation in LCLs and found a significant decrease in expression of one gene, glutaminase, in the dorsolateral prefrontal cortex (DLPFC). One SNP with previously identified regulation by a 3' UTR SNP was found to influence IRF5 expression in both brain and lymphocytes. The relationship between the 3' UTR rs10954213 genotype and IRF5 expression was significant in LCLs (p = 0.0001), DLPFC (p = 0.007), and anterior cingulate cortex (p = 0.002). CONCLUSION: Experimental manipulation of cells lines from subjects with schizophrenia may be a useful approach to explore stress related gene expression alterations in schizophrenia and to identify SNP variants associated with gene expression.

Mitochondrial involvement in psychiatric disorders.

Recent findings of mitochondrial abnormalities in brains from subjects with neurological disorders have led to a renewed search for mitochondrial abnormalities in psychiatric disorders. A growing body of evidence suggests that there is mitochondrial dysfunction in schizophrenia, bipolar disorder, and major depressive disorder, including evidence from electron microscopy, imaging, gene expression, genotyping, and sequencing studies. Specific evidence of dysfunction such as increased common deletion and decreased gene expression in mitochondria in psychiatric illnesses suggests that direct examination of mitochondrial DNA from postmortem brain cells may provide further details of mitochondrial alterations in psychiatric disorders.