mGluR5 hypofunction is integral to glutamatergic dysregulation in schizophrenia.

Multiple lines of evidence point to glutamatergic signaling in the postsynaptic density (PSD) as a pathophysiologic mechanism in schizophrenia. Integral to PSD glutamatergic signaling is reciprocal interplay between GluN and mGluR5 signaling. We examined agonist-induced mGluR5 signaling in the postmortem dorsolateral prefrontal cortex (DLPFC) derived from 17 patients and age-matched and sex-matched controls. The patient group showed a striking reduction in mGluR5 signaling, manifested by decreases in Gq/11 coupling and association with PI3K and Homer compared to controls (p < 0.01 for all). This was accompanied by increases in serine and tyrosine phosphorylation of mGluR5, which can decrease mGluR5 activity via desensitization (p < 0.01). In addition, we find altered protein-protein interaction (PPI) of mGluR5 with RGS4, norbin, Preso 1 and tamalin, which can also attenuate mGluR5 activity. We previously reported molecular underpinnings of GluN hypofunction (decreased GluN2 phosphorylation) and here we show those of reduced mGluR5 signaling in schizophrenia. We find that reduced GluN2 phosphorylation can be precipitated by attenuated mGluR5 activity and that increased mGluR5 phosphorylation can result from decreased GluN function, suggesting a reciprocal interplay between the two pathways in schizophrenia. Interestingly, the patient group showed decreased mGluR5-GluN association (p < 0.01), a mechanistic basis for the reciprocal facilitation. In sum, we present the first direct evidence for mGluR5 hypoactivity, propose a reciprocal interplay between GluN and mGluR5 pathways as integral to glutamatergic dysregulation and suggest protein-protein interactions in mGluR5-GluN complexes as potential targets for intervention in schizophrenia.

Abuse liability and therapeutic potential of the Mitragyna speciosa (kratom) alkaloids mitragynine and 7-hydroxymitragynine.

Kratom, derived from the plant Mitragyna speciosa, is receiving increased attention as an alternative to traditional opiates and as a replacement therapy for opiate dependence. Mitragynine (MG) and 7-hydroxymitragynine (7-HMG) are major psychoactive constituents of kratom. While MG and 7-HMG share behavioral and analgesic properties with morphine, their reinforcing effects have not been examined to date. 7-HMG, but not MG, substituted for morphine self-administration in a dose-dependent manner in the rat self-administration paradigm. Following the substitution procedure, re-assessment of morphine self-administration revealed a significant increase following 7-HMG and a significant decrease following MG substitution. In a separate cohort, 7-HMG, but not MG, engendered and maintained intravenous self-administration in a dose-dependent manner. The effects of pretreatment with nalxonaxine (NLXZ), a µ1 opiate receptor antagonist, and naltrindole (NTI), a δ opiate receptor antagonist, on 7-HMG and morphine self-administration were also examined. Both NLXZ and NTI reduced 7-HMG self-administration, whereas only NLXZ decreased morphine intake. The present results are the first to demonstrate that 7-HMG is readily self-administered, and the reinforcing effects of 7-HMG are mediated in part by µ and δ opiate receptors. In addition, prior exposure to 7-HMG increased subsequent morphine intake whereas prior exposure to MG decreased morphine intake. The present findings indicate that MG does not have abuse potential and reduces morphine intake, desired characteristics of candidate pharmacotherapies for opiate addiction and withdrawal, whereas 7-HMG should be considered a kratom constituent with high abuse potential that may also increase the intake of other opiates.

Brain-Wide Insulin Resistance, Tau Phosphorylation Changes, and Hippocampal Neprilysin and Amyloid-ß Alterations in a Monkey Model of Type 1 Diabetes.

Epidemiological findings suggest that diabetic individuals are at a greater risk for developing Alzheimer's disease (AD). To examine the mechanisms by which diabetes mellitus (DM) may contribute to AD pathology in humans, we examined brain tissue from streptozotocin-treated type 1 diabetic adult male vervet monkeys receiving twice-daily exogenous insulin injections for 8-20 weeks. We found greater inhibitory phosphorylation of insulin receptor substrate 1 in each brain region examined of the diabetic monkeys when compared with controls, consistent with a pattern of brain insulin resistance that is similar to that reported in the human AD brain. Additionally, a widespread increase in phosphorylated tau was seen, including brain areas vulnerable in AD, as well as relatively spared structures, such as the cerebellum. An increase in active ERK1/2 was also detected, consistent with DM leading to changes in tau-kinase activity broadly within the brain. In contrast to these widespread changes, we found an increase in soluble amyloid-ß (Aß) levels that was restricted to the temporal lobe, with the greatest increase seen in the hippocampus. Consistent with this localized Aß increase, a hippocampus-restricted decrease in the protein and mRNA for the Aß-degrading enzyme neprilysin (NEP) was found, whereas various Aß-clearing and -degrading proteins were unchanged. Thus, we document multiple biochemical changes in the insulin-controlled DM monkey brain that can link DM with the risk of developing AD, including dysregulation of the insulin-signaling pathway, changes in tau phosphorylation, and a decrease in NEP expression in the hippocampus that is coupled with a localized increase in Aß. SIGNIFICANCE STATEMENT: Given that diabetes mellitus (DM) appears to increase the risk of developing Alzheimer's disease (AD), understanding the mechanisms by which DM promotes AD is important. We report that DM in a nonhuman primate brain leads to changes in the levels or posttranslational processing of proteins central to AD pathobiology, including tau, amyloid-ß (Aß), and the Aß-degrading protease neprilysin. Additional evidence from this model suggests that alterations in brain insulin signaling occurred that are reminiscent of insulin signaling pathway changes seen in human AD. Thus, in an in vivo model highly relevant to humans, we show multiple alterations in the brain resulting from DM that are mechanistically linked to AD risk.

Changes in dopamine transporter binding in nucleus accumbens following chronic self-administration cocaine: heroin combinations.

Concurrent use of cocaine and heroin (speedball) has been shown to exert synergistic effects on dopamine neurotransmission in the nucleus accumbens (NAc), as observed by significant increases in extracellular dopamine levels and compensatory elevations in the maximal reuptake rate of dopamine. The present studies were undertaken to determine whether chronic self-administration of cocaine, heroin or a combination of cocaine:heroin led to compensatory changes in the abundance and/or affinity of high- and low-affinity DAT binding sites. Saturation binding of the cocaine analog [(125) I] 3ß-(4-iodophenyl)tropan-2ß-carboxylic acid methyl ester ([(125) I]RTI-55) in rat NAc membranes resulted in binding curves that were best fit to two-site binding models, allowing calculation of dissociation constant (Kd ) and binding density (Bmax ) values corresponding to high- and low-affinity DAT binding sites. Scatchard analysis of the saturation binding curves clearly demonstrate the presence of high- and low- affinity binding sites in the NAc, with low-affinity sites comprising 85 to 94% of the binding sites. DAT binding analyses revealed that self-administration of cocaine and a cocaine:heroin combination increased the affinity of the low-affinity site for the cocaine congener RTI-55 compared to saline. These results indicate that the alterations observed following chronic speedball self-administration are likely due to the cocaine component alone; thus further studies are necessary to elaborate upon the synergistic effect of cocaine:heroin combinations on the dopamine system in the NAc.

Monkey alcohol tissue research resource: banking tissues for alcohol research.

BACKGROUND: An estimated 18 million adults in the United States meet the clinical criteria for diagnosis of alcohol abuse or alcoholism, a disorder ranked as the third leading cause of preventable death. In addition to brain pathology, heavy alcohol consumption is comorbid with damage to major organs including heart, lungs, liver, pancreas, and kidneys. Much of what is known about risk for and consequences of heavy consumption derive from rodent or retrospective human studies. The neurobiological effects of chronic intake in rodent studies may not easily translate to humans due to key differences in brain structure and organization between species, including a lack of higher-order cognitive functions, and differences in underlying prefrontal cortical neural structures that characterize the primate brain. Further, rodents do not voluntarily consume large quantities of ethanol (EtOH) and they metabolize it more rapidly than primates. METHODS: The basis of the Monkey Alcohol Tissue Research Resource (MATRR) is that nonhuman primates, specifically monkeys, show a range of drinking excessive amounts of alcohol (>3.0 g/kg or a 12 drink equivalent per day) over long periods of time (12 to 30 months) with concomitant pathological changes in endocrine, hepatic, and central nervous system (CNS) processes. The patterns and range of alcohol intake that monkeys voluntarily consume parallel what is observed in humans with alcohol use disorders and the longitudinal experimental design spans stages of drinking from the EtOH-naïve state to early exposure through chronic abuse. Age- and sex-matched control animals self-administer an isocaloric solution under identical operant procedures. RESULTS: The MATRR is a unique postmortem tissue bank that provides CNS and peripheral tissues, and associated bioinformatics from monkeys that self-administer EtOH using a standardized experimental paradigm to the broader alcohol research community. CONCLUSIONS: This resource provides a translational platform from which we can better understand the disease processes associated with alcoholism.

Oxa-Iboga alkaloids lack cardiac risk and disrupt opioid use in animal models.

Ibogaine and its main metabolite noribogaine provide important molecular prototypes for markedly different treatment of substance use disorders and co-morbid mental health illnesses. However, these compounds present a cardiac safety risk and a highly complex molecular mechanism. We introduce a class of iboga alkaloids - termed oxa-iboga - defined as benzofuran-containing iboga analogs and created via structural editing of the iboga skeleton. The oxa-iboga compounds lack the proarrhythmic adverse effects of ibogaine and noribogaine in primary human cardiomyocytes and show superior efficacy in animal models of opioid use disorder in male rats. They act as potent kappa opioid receptor agonists in vitro and in vivo, but exhibit atypical behavioral features compared to standard kappa opioid agonists. Oxa-noribogaine induces long-lasting suppression of morphine, heroin, and fentanyl intake after a single dose or a short treatment regimen, reversal of persistent opioid-induced hyperalgesia, and suppression of opioid drug seeking in rodent relapse models. As such, oxa-iboga compounds represent mechanistically distinct iboga analogs with therapeutic potential.

Chronic haloperidol administration downregulates select BDNF transcript and protein levels in the dorsolateral prefrontal cortex of rhesus monkeys.

Post-mortem studies in the prefrontal cortex and hippocampal formation from schizophrenia patients have revealed significant disruptions in the expression molecules associated with cytoarchitecture, synaptic structure, function, and plasticity, known to be regulated in part by brain derived neurotrophic factor (BDNF). Interestingly, several studies using postmortem brain tissue from individuals diagnosed with schizophrenia have revealed a significant reduction in BDNF mRNA and protein levels in the dorsolateral prefrontal cortex (DLPFC), hippocampus and related areas; however, differentiating the effects of illness from antipsychotic history has remained difficult. We hypothesized that chronic antipsychotic treatment may contribute to the altered BDNF mRNA and protein expression observed in post-mortem brains of individuals diagnosed with schizophrenia. To address the influence of antipsychotic administration on BDNF expression in the primate brain, rhesus monkeys orally administered haloperidol, clozapine, or vehicle twice daily for 180 days. We found BDNF splice variants 4 and 5 in the DLPFC and variant 2 in the EC were significantly down-regulated following chronic administration of haloperidol. In addition, proBDNF and mature BDNF expression in the DLPFC, but not the EC, were significantly reduced. Based on the known regulation of BDNF expression by BDNF-AS, we assessed the expression of this lncRNA and found expression was significantly upregulated in the DLPFC, but not EC. The results of the present study provide evidence of haloperidol-induced regulation of BDNF mRNA and protein expression in the DLFPC and suggest an important role for BDNF-AS in this regulation. Given the role of BDNF in synaptic plasticity, neuronal survival and maintenance, aberrant expression induced by haloperidol likely has significant ramifications for neuronal populations and circuits in primate cortex.

Pharmacology and Therapeutic Potential of Benzothiazole Analogues for Cocaine Use Disorder.

Pharmacological targeting of the dopamine D4 receptor (D4R)─expressed in brain regions that control cognition, attention, and decision-making─could be useful for several neuropsychiatric disorders including substance use disorders (SUDs). This study focused on the synthesis and evaluation of a novel series of benzothiazole analogues designed to target D4R. We identified several compounds with high D4R binding affinity (Ki ≤ 6.9 nM) and >91-fold selectivity over other D2-like receptors (D2R, D3R) with diverse partial agonist and antagonist profiles. Novel analogue 16f is a potent low-efficacy D4R partial agonist, metabolically stable in rat and human liver microsomes, and has excellent brain penetration in rats (AUCbrain/plasma > 3). 16f (5-30 mg/kg, i.p.) dose-dependently decreased iv cocaine self-administration in rats, consistent with previous results produced by D4R-selective antagonists. Off-target antagonism of 5-HT2A or 5-HT2B may also contribute to these effects. Results with 16f support further efforts to target D4R in SUD treatment.

Differential regulation of accumbal dopamine transmission in rats following cocaine, heroin and speedball self-administration.

Cocaine/heroin combinations (speedball) exert synergistic neurochemical and behavioral effects that are thought to contribute to the increased abuse potential and subjective effects reported by polydrug users. In vivo fast-scan cyclic voltammetry was used to examine the effects of chronic intravenous self-administration (25 consecutive sessions) of cocaine (250 µg/inf), heroin (4.95 µg/inf) and speedball (250/4.95 µg/inf cocaine/heroin) on changes in electrically evoked dopamine (DA) efflux, maximal rate of DA uptake (V(max)) and the apparent affinity (K(m)) of the DA transporter in the nucleus accumbens. The increase in electrically evoked DA was comparable following cocaine and speedball injection; however, heroin did not increase evoked DA. DA transporter K(m) values were similarly elevated following cocaine and speedball, but unaffected by heroin. However, speedball self-administration significantly increased baseline V(max), while heroin and cocaine did not change baseline V(max), compared with the baseline V(max) values of drug-naïve animals. Overall, elevated DA clearance is a likely consequence of synergistic elevations of nucleus accumbens extracellular DA concentrations by chronic speedball self-administration, as reported previously in microdialysis studies. The present results indicate neuroadaptive processes that are unique to cocaine/heroin combinations and cannot be readily explained by simple additivity of changes observed with cocaine and heroin alone.

Three diketomorpholines from a Penicillium sp. (strain G1071).

Three previously undescribed diketomorpholine natural products, along with the known phenalenones, herqueinone and norherqueinone, were isolated from the mycoparasitic fungal strain G1071, which was identified as a Penicillium sp. in the section Sclerotiora. The structures were established by analyzing NMR data and mass spectrometry fragmentation patterns. The absolute configurations of deacetyl-javanicunine A, javanicunine C, and javanicunine D, were assigned by examining ECD spectra and Marfey's analysis. The structural diversity generated by this fungal strain was interesting, as only a few diketomorpholines (~17) have been reported from nature.

Incubation of nicotine seeking is associated with enhanced protein kinase A-regulated signaling of dopamine- and cAMP-regulated phosphoprotein of 32 kDa in the insular cortex.

A recent clinical study demonstrated that damage to the insular cortex can disrupt tobacco addiction. The neurobiological mechanisms for this effect are not yet understood. In this study we used an animal model of nicotine addiction to examine the possibility that changes in insular cortex levels of dopamine (DA)- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32), a phosphoprotein enriched in DA neurons containing DA D1 receptors, may be associated with changes in vulnerability to nicotine addiction. Once rats acquired self-administration, they were given unlimited access to nicotine (0.01 mg/kg/infusion) for 23 h/day for a total of 10 days. Each infusion was paired with a visual cue (stimulus light) and auditory cue (sound of pump). Nicotine seeking, as assessed under a cue-induced reinstatement paradigm, and markers of DARPP-32 signaling, as assessed using western blot analysis, were examined in separate groups of rats at two different abstinent intervals: 1 and 7 days. Consistent with findings with other drugs of abuse, rats in the 7-day abstinence group took longer to extinguish and responded at higher levels during reinstatement testing as compared with rats in the 1-day reinstatement group. Relative to saline controls, rats in the 7-day but not the 1-day abstinence group had higher levels of DARPP-32 phosphorylated at the protein kinase A site in the insular cortex. These results demonstrate incubation of drug seeking following extended access to nicotine self-administration and suggest that enhanced protein kinase A signaling in the insular cortex via phosphorylation of DARPP-32 at Thr34 is associated with this effect.

Modeling substance abuse for applications in proteomics.

The ability to model aspects of human addictive behaviors in laboratory animals provides an important avenue for gaining insight into the biochemical alterations associated with drug intake and the identification of targets for medication development to treat addictive disorders. The intravenous self-administration procedure provides the means to model the reinforcing effects of abused drugs and to correlate biochemical alterations with drug reinforcement. In this chapter, we provide a detailed methodology for rodent intravenous self-administration and the isolation and preparation of proteins from dissected brain regions for Western blot analysis and high-throughput proteomic analysis. Examples of cocaine-induced alterations in the abundances of ionotropic glutamate receptor subunits in reinforcement-related brain regions are provided.

De novo protein sequence analysis of Macaca mulatta.

BACKGROUND: Macaca mulatta is one of the most utilized non-human primate species in biomedical research offering unique behavioral, neuroanatomical, and neurobiochemcial similarities to humans. This makes it a unique organism to model various diseases such as psychiatric and neurodegenerative illnesses while also providing insight into the complexities of the primate brain. A major obstacle in utilizing rhesus monkey models for human disease is the paucity of protein annotations for this species (~42,000 protein annotations) compared to 330,210 protein annotations for humans. The lack of available information limits the use of rhesus monkey for proteomic scale studies which rely heavily on database searches for protein identification. While characterization of proteins of interest from Macaca mulatta using the standard database search engines (e.g., MASCOT) can be accomplished, searches must be performed using a 'broad species database' which does not provide optimal confidence in protein annotation. Therefore, it becomes necessary to determine partial or complete amino acid sequences using either manual or automated de novo peptide sequence analysis methods. RESULTS: The recently popularized MALDI-TOF-TOF mass spectrometer yields a complex MS/MS fragmentation pattern difficult to characterize by manual de novo sequencing method on a proteomics scale. Therefore, PEAKS assisted de novo sequencing was performed on nucleus accumbens cytosolic proteins from Macaca mulatta. The most abundant peptide fragments 'b-ions and y-ions', the less abundant peptide fragments 'a-ions' as well as the immonium ions were utilized to develop confident and complete peptide sequences de novo from MS/MS spectra. The generated sequences were used to perform homology searches to characterize the protein identification. CONCLUSION: The current study validates a robust method to confidently characterize the proteins from an incomplete sequence database of Macaca mulatta, using the PEAKS de novo sequencing software, facilitating the use of this animal model in various neuroproteomics studies.

Region specific regulation of NR1 in rhesus monkeys following chronic antipsychotic drug administration.

BACKGROUND: Altered NMDA receptor subunit protein levels have been reported in various regions of the schizophrenic brain; however, chronic antipsychotic administration in schizophrenic subjects may confound interpretation. METHODS: The effects of chronic antipsychotic drug administration (haloperidol and clozapine) on protein levels of NR1, NR2A and NR2B proteins were evaluated in the nucleus accumbens (NAc), putamen (PUT), dorsolateral prefrontal cortex (DLPFC), superior temporal gyrus (STG), and entorhinal cortex (EC) of rhesus monkeys using Western blot analysis. RESULTS: Haloperidol administration significantly decreased NR1 expression in the DLPFC. In contrast, NR2B expression was not affected by antipsychotic administration in any brain region examined. NR2A was not reliably detected in any of the brain regions. CONCLUSIONS: Results indicate that the NR1 subunit in the DLPFC may be a substrate for antipsychotic action and that glutamatergic hypofunction in the DLPFC commonly associated with cognitive dysfunction in schizophrenia may be associated with haloperidol administration.

Self-administered heroin and cocaine combinations in the rat: additive reinforcing effects-supra-additive effects on nucleus accumbens extracellular dopamine.

The concurrent use of cocaine and opiate combinations (speedball) has increased since the 1970s and now represents a growing subset of intravenous drug abusers. An isobolographic analysis was applied to the ascending limb of the dose-effect curves for rat self-administration of cocaine, heroin, and their combination to determine the nature of the interaction. The addition of heroin to cocaine shifted the dose-effect curve for self-administration to the left, and the modulation in reinforcing efficacy of the combination of cocaine and heroin was found to be additive. A second experiment used microdialysis to determine the effects of this drug combination on nucleus accumbens (NAc) extracellular levels of dopamine ([DA](e)) in rats self-administering low doses of cocaine, heroin, or cocaine/heroin combinations. These doses of cocaine and cocaine/heroin combinations significantly increased NAc [DA](e), while heroin alone did not. The ratio of the % baseline of [DA](e) (or the dialysate concentrations of DA) to cocaine in the dialysate was higher during self-administration of cocaine/heroin combinations than with cocaine alone. These data indicate that although the interaction between cocaine and heroin in maintaining self-administration is additive, a potentiation of NAc dopaminergic neurotransmission is present, suggesting that NAc [DA](e) may not be a direct measure of reinforcing efficacy and/or it is not central to the mediation of the self-administration of this drug combination.

Assessment of genome and proteome profiles in cocaine abuse.

Until recently, knowledge of the impact of abuse drugs on gene and protein expression in the brain was limited to less than 100 targets. With the advent of high-throughput genomic and proteomic techniques investigators are now able to evaluate changes across the entire genome and across thousands of proteins in defined brain regions and generate expression profiles of vulnerable neuroanatomical substrates in rodent and non-human primate drug abuse models and in human post-mortem brain tissue from drug abuse victims. The availability of gene and protein expression profiles will continue to expand our understanding of the short- and long-term consequences of drug addiction and other addictive disorders and may provide new approaches or new targets for pharmacotherapeutic intervention. This chapter will review gene expression data from rodent, non-human primate and human post-mortem studies of cocaine abuse and will provide a preliminary proteomic profile of human cocaine abuse and explore how these studies have advanced our understanding of addiction.

Methods for proteomics in neuroscience.

Proteomics reveals complex protein expression, function, interactions and localization in different phenotypes of neuron. As proteomics, regarded as a highly complex screening technology, moves from a theoretical approach to practical reality, neuroscientists have to determine the most-appropriate applications for this technology. Even though proteomics compliments genomics, it is in sheer contrast to the basically constant genome due to its dynamic nature. Neuroscientists have to surmount difficulties particular to the research in neuroscience; such as limited sample amounts, heterogeneous cellular compositions in samples and the fact that many proteins of interest are hydrophobic proteins. The necessity of exclusive technology, sophisticated software and skilled manpower tops the challenge. This review examines subcellular organelle isolation, protein fractionation and separation using two-dimensional gel electrophoresis (2-DGE) as well as multi-dimensional liquid chromatography (LC) followed by mass spectrometry (MS). The methods for quantifying relative gene product expression between samples (e.g., two-dimensional difference in gel electrophoresis (2D-DIGE), isotope-coded affinity tag (ICAT) and iTRAQ) are elaborated. An overview of the techniques used currently to assign post-translational modification status on a proteomics scale is also evaluated. The feasible coverage of the proteome, ability to detect unique cell components such as post-synaptic densities and membrane proteins, resource requirements and quantitative as well as qualitative reliability of different approaches is also discussed. While there are many challenges in neuroproteomics, this field promises many returns in the future.

NeuN+ neuronal nuclei in non-human primate prefrontal cortex and subcortical white matter after clozapine exposure.

Increased neuronal densities in subcortical white matter have been reported for some cases with schizophrenia. The underlying cellular and molecular mechanisms remain unresolved. We exposed 26 young adult macaque monkeys for 6 months to either clozapine, haloperidol or placebo and measured by structural MRI frontal gray and white matter volumes before and after treatment, followed by observer-independent, flow-cytometry-based quantification of neuronal and non-neuronal nuclei and molecular fingerprinting of cell-type specific transcripts. After clozapine exposure, the proportion of nuclei expressing the neuronal marker NeuN increased by approximately 50% in subcortical white matter, in conjunction with a more subtle and non-significant increase in overlying gray matter. Numbers and proportions of nuclei expressing the oligodendrocyte lineage marker, OLIG2, and cell-type specific RNA expression patterns, were maintained after antipsychotic drug exposure. Frontal lobe gray and white matter volumes remained indistinguishable between antipsychotic-drug-exposed and control groups. Chronic clozapine exposure increases the proportion of NeuN+ nuclei in frontal subcortical white matter, without alterations in frontal lobe volumes or cell type-specific gene expression. Further exploration of neurochemical plasticity in non-human primate brain exposed to antipsychotic drugs is warranted.

Gene expression elucidates functional impact of polygenic risk for schizophrenia.

Over 100 genetic loci harbor schizophrenia-associated variants, yet how these variants confer liability is uncertain. The CommonMind Consortium sequenced RNA from dorsolateral prefrontal cortex of people with schizophrenia (N = 258) and control subjects (N = 279), creating a resource of gene expression and its genetic regulation. Using this resource, ∼20% of schizophrenia loci have variants that could contribute to altered gene expression and liability. In five loci, only a single gene was involved: FURIN, TSNARE1, CNTN4, CLCN3 or SNAP91. Altering expression of FURIN, TSNARE1 or CNTN4 changed neurodevelopment in zebrafish; knockdown of FURIN in human neural progenitor cells yielded abnormal migration. Of 693 genes showing significant case-versus-control differential expression, their fold changes were ≤ 1.33, and an independent cohort yielded similar results. Gene co-expression implicates a network relevant for schizophrenia. Our findings show that schizophrenia is polygenic and highlight the utility of this resource for mechanistic interpretations of genetic liability for brain diseases.

Differential regulation of ionotropic glutamate receptor subunits following cocaine self-administration.

Previous examination of binge cocaine self-administration and 2 week withdrawal from cocaine self-administration on ionotropic glutamate receptor subunit (iGluRs) protein levels revealed significant alterations in iGluR protein levels that differed between the mesocorticolimbic and nigrostriatal pathways. The present study was undertaken to extend the examination of cocaine-induced alterations in iGluR protein expression by assessing the effects of acute withdrawal (15-16 h) from limited access cocaine self-administration (8 h/day, 15 days). Western blotting was used to compare levels of iGluR protein expression (NR1-3B, GluR1-7, KA2) in the mesolimbic (ventral tegmental area, VTA; nucleus accumbens, NAc; and prefrontal cortex, PFC) and nigrostriatal pathways (substantia nigra, SN and dorsal caudate-putamen, CPu). Within the mesolimbic pathway, reductions were observed in NR1 and GluR5 immunoreactivity in the VTA although no significant alterations were observed in any iGluR subunits in the NAc. In the PFC, NR1 was significantly upregulated while GluR2/3, GluR4, GluR5, GluR6/7, and KA2 were decreased. Within the nigrostriatal pathway, NR1, NR2A, NR2B, GluR1, GluR6/7 and KA2 were increased in the dorsal CPu, whereas no significant changes were observed in the SN. The results demonstrate region- and pathway-specific alterations in iGluR subunit expression following limited cocaine self-administration and suggest the importance for the activation of pathways that are substrates of the reinforcing and motoric effects of cocaine.