The sea urchin embryo as a model for mammalian developmental neurotoxicity: ontogenesis of the high-affinity choline transporter and its role in cholinergic trophic activity.

Embryonic development in the sea urchin requires trophic actions of the same neurotransmitters that participate in mammalian brain assembly. We evaluated the development of the high-affinity choline transporter, which controls acetylcholine synthesis. A variety of developmental neurotoxicants affect this transporter in mammalian brain. [3H]Hemicholinium-3 binding to the transporter was found in the cell membrane fraction at stages from the unfertilized egg to pluteus, with a binding affinity comparable with that seen in mammalian brain. Over the course of development, the concentration of transporter sites rose more than 3-fold, achieving concentrations comparable with those of cholinergically enriched mammalian brain regions. Dimethylaminoethanol (DMAE), a competitive inhibitor of choline transport, elicited dysmorphology beginning at the mid-blastula stage, with anomalies beginning progressively later as the concentration of DMAE was lowered. Pretreatment, cotreatment, or delayed treatment with acetylcholine or choline prevented the adverse effects of DMAE. Because acetylcholine was protective at a lower threshold, the DMAE-induced defects were most likely mediated by its effects on acetylcholine synthesis. Transient removal of the hyaline layer enabled a charged transport inhibitor, hemicholinium-3, to penetrate sufficiently to elicit similar anomalies, which were again prevented by acetylcholine or choline. These results indicate that the developing sea urchin possesses a high-affinity choline transporter analogous to that found in the mammalian brain, and, as in mammals, the functioning of this transporter plays a key role in the developmental, trophic activity of acetylcholine. The sea urchin model may thus be useful in high-throughput screening of suspected developmental neurotoxicants

An Invertebrate Model of the Developmental Neurotoxicity of Insecticides: Effects of Chlorpyrifos and Dieldrin in Sea Urchin Embryos and Larvae

Chlorpyrifos targets mammalian brain development through a combination of effects directed at cholinergic receptors and intracellular signaling cascades that are involved in cell differentiation. We used sea urchin embryos as an invertebrate model system to explore the cellular mechanisms underlying the actions of chlorpyrifos and to delineate the critical period of developmental vulnerability. Sea urchin embryos and larvae were exposed to chlorpyrifos at different stages of development ranging from early cell cleavages through the prism stage. Although early cleavages were unaffected even at high chlorpyrifos concentrations, micromolar concentrations added at the mid-blastula stage evoked a prominent change in cell phenotype and overall larval structure, with appearance of pigmented cells followed by their accumulation in an extralarval cap that was extruded from the animal pole. At higher concentrations (20-40 microM), these abnormal cells constituted over 90% of the total cell number. Studies with cholinergic receptor blocking agents and protein kinase C inhibitors indicated two distinct types of effects, one mediated through stimulation of nicotinic cholinergic receptors and the other targeting intracellular signaling. The effects of chlorpyrifos were not mimicked by chlorpyrifos oxon, the active metabolite that inhibits cholinesterase, nor by nonorganophosphate cholinesterase inhibitors. Dieldrin, an organochlorine that targets GABA(A )receptors, was similarly ineffective. The effects of chlorpyrifos and its underlying cholinergic and signaling-related mechanisms parallel prior findings in mammalian embryonic central nervous system. Invertebrate test systems may thus provide both a screening procedure for potential neuroteratogenesis by organophosphate-related compounds, as well as a system with which to uncover novel mechanisms underlying developmental vulnerability

The sea urchin embryo, an invertebrate model for mammalian developmental neurotoxicity, reveals multiple neurotransmitter mechanisms for effects of chlorpyrifos: Therapeutic interventions and a comparison with the monoamine depleter, reserpine

Lower organisms show promise for the screening of neurotoxicants that might target mammalian brain development. Sea urchins use neurotransmitters as embryonic growth regulatory signals, so that adverse effects on neural substrates for mammalian brain development can be studied in this simple organism. We compared the effects of the organophosphate insecticide, chlorpyrifos in sea urchin embryos with those of the monoamine depleter, reserpine, so as to investigate multiple neurotransmitter mechanisms involved in developmental toxicity and to evaluate different therapeutic interventions corresponding to each neurotransmitter system. Whereas reserpine interfered with all stages of embryonic development, the effects of chlorpyrifos did not emerge until the mid-blastula stage. After that point, the effects of the two agents were similar. Treatment with membrane permeable analogs of the monoamine neurotransmitters, serotonin and dopamine, prevented the adverse effects of either chlorpyrifos or reserpine, despite the fact that chlorpyrifos works simultaneously through actions on acetylcholine, monoamines and other neurotransmitter pathways. This suggests that different neurotransmitters, converging on the same downstream signaling events, could work together or in parallel to offset the developmental disruption caused by exposure to disparate agents. We tested this hypothesis by evaluating membrane permeable analogs of acetylcholine and cannabinoids, both of which proved effective against chlorpyrifos- or reserpine-induced teratogenesis. Invertebrate test systems can provide both a screening procedure for mammalian neuroteratogenesis and may uncover novel mechanisms underlying developmental vulnerability as well as possible therapeutic approaches to prevent teratogenesis

No Reliable Association between Runs of Homozygosity and Schizophrenia in a Well-Powered Replication Study

It is well known that inbreeding increases the risk of recessive monogenic diseases, but it is less certain whether it contributes to the etiology of complex diseases such as schizophrenia. One way to estimate the effects of inbreeding is to examine the association between disease diagnosis and genome-wide autozygosity estimated using runs of homozygosity (ROH) in genome-wide single nucleotide polymorphism arrays. Using data for schizophrenia from the Psychiatric Genomics Consortium (n = 21,868), Keller et al. (2012) estimated that the odds of developing schizophrenia increased by approximately 17% for every additional percent of the genome that is autozygous (β = 16.1, CI(β) = [6.93, 25.7], Z = 3.44, p = 0.0006). Here we describe replication results from 22 independent schizophrenia case-control datasets from the Psychiatric Genomics Consortium (n = 39,830). Using the same ROH calling thresholds and procedures as Keller et al. (2012), we were unable to replicate the significant association between ROH burden and schizophrenia in the independent PGC phase II data, although the effect was in the predicted direction, and the combined (original + replication) dataset yielded an attenuated but significant relationship between Froh and schizophrenia (β = 4.86,CI(β) = [0.90,8.83],Z = 2.40,p = 0.02). Since Keller et al. (2012), several studies reported inconsistent association of ROH burden with complex traits, particularly in case-control data. These conflicting results might suggest that the effects of autozygosity are confounded by various factors, such as socioeconomic status, education, urbanicity, and religiosity, which may be associated with both real inbreeding and the outcome measures of interest

Gene expression imputation across multiple brain regions provides insights into schizophrenia risk

Transcriptomic imputation approaches combine eQTL reference panels with large-scale genotype data in order to test associations between disease and gene expression. These genic associations could elucidate signals in complex genome-wide association study (GWAS) loci and may disentangle the role of different tissues in disease development. We used the largest eQTL reference panel for the dorso-lateral prefrontal cortex (DLPFC) to create a set of gene expression predictors and demonstrate their utility. We applied DLPFC and 12 GTEx-brain predictors to 40,299 schizophrenia cases and 65,264 matched controls for a large transcriptomic imputation study of schizophrenia. We identified 413 genic associations across 13 brain regions. Stepwise conditioning identified 67 non-MHC genes, of which 14 did not fall within previous GWAS loci. We identified 36 significantly enriched pathways, including hexosaminidase-A deficiency, and multiple porphyric disorder pathways. We investigated developmental expression patterns among the 67 non-MHC genes and identified specific groups of pre- and postnatal expression

Genetic correlation between amyotrophic lateral sclerosis and schizophrenia

A. Palotie on työryhmän Schizophrenia Working Grp Psychiat jäsen.We have previously shown higher-than-expected rates of schizophrenia in relatives of patients with amyotrophic lateral sclerosis (ALS), suggesting an aetiological relationship between the diseases. Here, we investigate the genetic relationship between ALS and schizophrenia using genome-wide association study data from over 100,000 unique individuals. Using linkage disequilibrium score regression, we estimate the genetic correlation between ALS and schizophrenia to be 14.3% (7.05-21.6; P = 1 x 10(-4)) with schizophrenia polygenic risk scores explaining up to 0.12% of the variance in ALS (P = 8.4 x 10(-7)). A modest increase in comorbidity of ALS and schizophrenia is expected given these findings (odds ratio 1.08-1.26) but this would require very large studies to observe epidemiologically. We identify five potential novel ALS-associated loci using conditional false discovery rate analysis. It is likely that shared neurobiological mechanisms between these two disorders will engender novel hypotheses in future preclinical and clinical studies.Peer reviewe

Estimation of Genetic Correlation via Linkage Disequilibrium Score Regression and Genomic Restricted Maximum Likelihood

J. Lönnqvist on työryhmän Psychiat Genomics Consortium jäsen.Genetic correlation is a key population parameter that describes the shared genetic architecture of complex traits and diseases. It can be estimated by current state-of-art methods, i.e., linkage disequilibrium score regression (LDSC) and genomic restricted maximum likelihood (GREML). The massively reduced computing burden of LDSC compared to GREML makes it an attractive tool, although the accuracy (i.e., magnitude of standard errors) of LDSC estimates has not been thoroughly studied. In simulation, we show that the accuracy of GREML is generally higher than that of LDSC. When there is genetic heterogeneity between the actual sample and reference data from which LD scores are estimated, the accuracy of LDSC decreases further. In real data analyses estimating the genetic correlation between schizophrenia (SCZ) and body mass index, we show that GREML estimates based on similar to 150,000 individuals give a higher accuracy than LDSC estimates based on similar to 400,000 individuals (from combinedmeta-data). A GREML genomic partitioning analysis reveals that the genetic correlation between SCZ and height is significantly negative for regulatory regions, which whole genome or LDSC approach has less power to detect. We conclude that LDSC estimates should be carefully interpreted as there can be uncertainty about homogeneity among combined meta-datasets. We suggest that any interesting findings from massive LDSC analysis for a large number of complex traits should be followed up, where possible, with more detailed analyses with GREML methods, even if sample sizes are lesser.Peer reviewe

Interaction Testing and Polygenic Risk Scoring to Estimate the Association of Common Genetic Variants with Treatment Resistance in Schizophrenia

Importance: About 20% to 30% of people with schizophrenia have psychotic symptoms that do not respond adequately to first-line antipsychotic treatment. This clinical presentation, chronic and highly disabling, is known as treatment-resistant schizophrenia (TRS). The causes of treatment resistance and their relationships with causes underlying schizophrenia are largely unknown. Adequately powered genetic studies of TRS are scarce because of the difficulty in collecting data from well-characterized TRS cohorts. Objective: To examine the genetic architecture of TRS through the reassessment of genetic data from schizophrenia studies and its validation in carefully ascertained clinical samples. Design, Setting, and Participants: Two case-control genome-wide association studies (GWASs) of schizophrenia were performed in which the case samples were defined as individuals with TRS (n = 10501) and individuals with non-TRS (n = 20325). The differences in effect sizes for allelic associations were then determined between both studies, the reasoning being such differences reflect treatment resistance instead of schizophrenia. Genotype data were retrieved from the CLOZUK and Psychiatric Genomics Consortium (PGC) schizophrenia studies. The output was validated using polygenic risk score (PRS) profiling of 2 independent schizophrenia cohorts with TRS and non-TRS: a prevalence sample with 817 individuals (Cardiff Cognition in Schizophrenia [CardiffCOGS]) and an incidence sample with 563 individuals (Genetics Workstream of the Schizophrenia Treatment Resistance and Therapeutic Advances [STRATA-G]). Main Outcomes and Measures: GWAS of treatment resistance in schizophrenia. The results of the GWAS were compared with complex polygenic traits through a genetic correlation approach and were used for PRS analysis on the independent validation cohorts using the same TRS definition. Results: The study included a total of 85490 participants (48635 [56.9%] male) in its GWAS stage and 1380 participants (859 [62.2%] male) in its PRS validation stage. Treatment resistance in schizophrenia emerged as a polygenic trait with detectable heritability (1% to 4%), and several traits related to intelligence and cognition were found to be genetically correlated with it (genetic correlation, 0.41-0.69). PRS analysis in the CardiffCOGS prevalence sample showed a positive association between TRS and a history of taking clozapine (r2 = 2.03%; P =.001), which was replicated in the STRATA-G incidence sample (r2 = 1.09%; P =.04). Conclusions and Relevance: In this GWAS, common genetic variants were differentially associated with TRS, and these associations may have been obscured through the amalgamation of large GWAS samples in previous studies of broadly defined schizophrenia. Findings of this study suggest the validity of meta-analytic approaches for studies on patient outcomes, including treatment resistance

Genomic Dissection of Bipolar Disorder and Schizophrenia, Including 28 Subphenotypes

Schizophrenia and bipolar disorder are two distinct diagnoses that share symptomology. Understanding the genetic factors contributing to the shared and disorder-specific symptoms will be crucial for improving diagnosis and treatment. In genetic data consisting of 53,555 cases (20,129 bipolar disorder [BD], 33,426 schizophrenia [SCZ]) and 54,065 controls, we identified 114 genome-wide significant loci implicating synaptic and neuronal pathways shared between disorders. Comparing SCZ to BD (23,585 SCZ, 15,270 BD) identified four genomic regions including one with disorder-independent causal variants and potassium ion response genes as contributing to differences in biology between the disorders. Polygenic risk score (PRS) analyses identified several significant correlations within case-only phenotypes including SCZ PRS with psychotic features and age of onset in BD. For the first time, we discover specific loci that distinguish between BD and SCZ and identify polygenic components underlying multiple symptom dimensions. These results point to the utility of genetics to inform symptomology and potential treatment