Impact of Bariatric Surgery on Subtilisin/Kexin Type 9 (PCSK9) Gene Expression and Inflammation in the Adipose Tissue of Obese Diabetic Rats.

Bariatric surgery improves dyslipidaemia and reduces body weight, but it remains unclear how bariatric surgery modulates gene expression in fat cells to influence the proprotein convertase subtilisin/kexin type 9 (PCSK-9) and low-density lipoprotein receptor (LDLR) gene expression. The expression of the PCSK9/LDLR/tumor necrosis factor-alpha (TNFα) gene in adipose tissue was measured in two groups of Zucker Diabetic Sprague Dawley (ZDSD) rats after Roux-en-Y gastric bypass (RYGB) surgery or 'SHAM' operation. There was lower PCSK9 (p = 0.02) and higher LDLR gene expression (p = 0.02) in adipose tissue in rats after RYGB. Weight change did not correlate with PCSK9 gene expression (r = -0.5, p = 0.08) or TNFα gene expression (r = -0.4, p = 0.1). TNFα gene expression was positively correlated with PCSK9 gene expression (r = 0.7, p = 0.001) but not correlated with LDLR expression (r = -0.3, p = 0.3). Circulating triglyceride levels were lower in RYGB compared to the SHAM group (1.1 (0.8-1.4) vs. 1.5 (1.0-4.2), p = 0.038) mmol/L with no difference in cholesterol levels. LDLR gene expression was increased post-bariatric surgery with the potential to reduce the number of circulating LDL particles. PCSK9 gene expression and TNFα gene expression were positively correlated after RYGB in ZDSD rats, suggesting that the modulation of pro-inflammatory pathways in adipose tissue after RYGB may partly relate to PCSK9 and LDLR gene expression.

Ceftriaxone pretreatment confers neuroprotection in rats with acute glaucoma and reduces the score of seizures induced by pentylenotetrazole in mice.

ß-Lactam antibiotics such as ceftriaxone, are potent stimulators of the expression of l-glutamate transporter GLT-1 and may exert neuroprotective effects when chronically used in rats and mice. In this study, we used two animal models to test the neurological effect of subchronic treatment with ceftriaxone: experimental acute glaucoma in Wistar rats and induction of acute seizures with pentylenetetrazole in mice. We also assessed the performance of mice in the rotarod to calculate therapeutic indexes and exploratory activity in the open field. Our results showed that subchronic use of ceftriaxone was neuroprotective in both models, reducing injury in acute ischemia and ischemia/reperfusion in specific layers of retina and leading to a decrease in the seizure severity score. In behavioral experiments, we observed that ceftriaxone increased hyperactivity followed by a decrease in exploratory behavior in the open field, and there was no motor impairment in the rotarod test. We conclude that ceftriaxone may be useful as a tool in the development of new neuroprotective drugs targeting diseases which present a possible dysfunction in the balance of glutamatergic neurotransmission.

Elevated Expression of Two Pore Potassium Channel THIK-1 in Alzheimer's Disease: An Inflammatory Mechanism.

INTRODUCTION: Tandem pore domain halothane-inhibited K+ channel 1 (THIK-1, coded by KCNK13) provides an upstream regulation of the activation of the NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome, which has been suggested as one of the key mechanisms of the pathological process in neurodegeneration mainly from in vitro and in vivo model systems studies. However, unequivocal evidence from neurodegenerative disorders has been lacking. OBJECTIVE: To investigate the involvement of the THIK-1/NLRP3 pathway in the pathological process of Alzheimer's disease (AD) and Parkinson's disease (PD). METHODS: This study investigated gene expression of markers in the THIK-1/NLRP3 pathway in an animal model representing AD as well as in human postmortem brains of AD and PD by quantitative real-time PCR. THIK-1 protein expression was determined using automated capillary electrophoresis immunoblotting. Furthermore, DNA methylation of KCNK13 was analysed in AD cohort by pyrosequencing. RESULTS: A substantial upregulation of KCNK13, glial activation markers, NLRP3 inflammasome components, and IL1B was observed in the animal study. Increased expression of KCNK13 support an inflammatory glial cell activation in both advanced AD and PD. The increase in KCNK13 expression was also supported by downregulation in DNA methylation of KCNK13 in AD. CONCLUSIONS: The association between THIK-1 K+ channels expression and pathology changes indicates a THIK-1-induced activation of this glial subtype in AD and PD. Therefore, specific blocks of the microglial THIK-1 K+ channels at the early stage of AD and PD may be beneficial for the patients.

Subchronic PCP effects on DNA methylation and protein expression of NMDA receptor subunit genes in the prefrontal cortex and hippocampus of female rats.

BACKGROUND: N-methyl-d-aspartate receptor (NMDAR) dysfunction is implicated in schizophrenia, and NMDAR antagonists, such as phencyclidine (PCP), can induce behaviours that mimic aspects of the disorder. AIMS: We investigated DNA methylation of Grin1, Grin2a and Grin2b promoter region and NR1 and NR2 protein expression in the prefrontal cortex (PFC) and hippocampus of adult female Lister-hooded rats following subchronic PCP (scPCP) administration. We also determined whether any alterations were tissue-specific. METHODS: Rats were divided into two groups that received vehicle (0.9% saline) or 2 mg/kg PCP twice a day for 7 days (n = 10 per group). After behavioural testing (novel object recognition), to confirm a cognitive deficit, brains were dissected and NMDAR subunit DNA methylation and protein expression were analysed by pyrosequencing and ELISA. Line-1 methylation was determined as a measure of global methylation. Data were analysed using Student's t-test and Pearson correlation. RESULTS: The scPCP administration led to Grin1 and Grin2b hypermethylation and reduction in NR1 protein in both PFC and hippocampus. No significant differences were observed in Line-1 or Grin2a methylation and NR2 protein. CONCLUSIONS: The scPCP treatment resulted in increased DNA methylation at promoter sites of Grin1 and Grin2b NMDAR subunits in two brain areas implicated in schizophrenia, independent of any global change in DNA methylation, and are similar to our observations in a neurodevelopmental animal model of schizophrenia - social isolation rearing post-weaning. Moreover, these alterations may contribute to the changes in protein expression for NMDAR subunits demonstrating the potential importance of epigenetic mechanisms in schizophrenia.

Methylation Status of Exon IV of the Brain-Derived Neurotrophic Factor (BDNF)-Encoding Gene in Patients with Non-Diabetic Hyperglycaemia (NDH) before and after a Lifestyle Intervention.

BDNF signalling in hypothalamic neuronal circuits is thought to regulate mammalian food intake. In light of this, we investigated how a lifestyle intervention influenced serum levels and DNA methylation of BDNF gene in fat tissue and buffy coat of NDH individuals. In total, 20 participants underwent anthropometric measurements/fasting blood tests and adipose tissue biopsy pre-/post-lifestyle (6 months) intervention. DNA was extracted from adipose tissue and buffy coat, bisulphite converted, and pyrosequencing was used to determine methylation levels in exon IV of the BDNF gene. RNA was extracted from buffy coat for gene expression analysis and serum BDNF levels were measured by ELISA. No differences were found in BDNF serum levels, but buffy coat mean BDNF gene methylation decreased post-intervention. There were correlations between BDNF serum levels and/or methylation and cardiometabolic markers. (i) Pre-intervention: for BDNF methylation, we found positive correlations between mean methylation in fat tissue and waist-hip ratio, and negative correlations between mean methylation in buffy coat and weight. (ii) Post-intervention: we found correlations between BDNF mean methylation in buffy coat and HbA1c, BDNF methylation in buffy coat and circulating IGFBP-2, and BDNF serum and insulin. Higher BDNF % methylation levels are known to reduce BNDF expression. The fall in buffy coat mean BDNF methylation plus the association between lower BDNF methylation (so potentially higher BDNF) and higher HbA1c and serum IGFBP-2 (as a marker of insulin sensitivity) and between lower serum BDNF and higher circulating insulin are evidence for the degree of BDNF gene methylation being implicated in insulinisation and glucose homeostasis, particularly after lifestyle change in NDH individuals.

Relationship between the Plasma Proteome and Changes in Inflammatory Markers after Bariatric Surgery.

Severe obesity is a disease associated with multiple adverse effects on health. Metabolic bariatric surgery (MBS) can have significant effects on multiple body systems and was shown to improve inflammatory markers in previous short-term follow-up studies. We evaluated associations between changes in inflammatory markers (CRP, IL6 and TNFα) and circulating proteins after MBS. METHODS: Sequential window acquisition of all theoretical mass spectra (SWATH-MS) proteomics was performed on plasma samples taken at baseline (pre-surgery) and 6 and 12 months after MBS, and concurrent analyses of inflammatory/metabolic parameters were carried out. The change in absolute abundances of those proteins, showing significant change at both 6 and 12 months, was tested for correlation with the absolute and percentage (%) change in inflammatory markers. RESULTS: We found the following results: at 6 months, there was a correlation between %change in IL-6 and fold change in HSPA4 (rho = -0.659; p = 0.038) and in SERPINF1 (rho = 0.714, p = 0.020); at 12 months, there was a positive correlation between %change in IL-6 and fold change in the following proteins-LGALS3BP (rho = 0.700, p = 0.036), HSP90B1 (rho = 0.667; p = 0.05) and ACE (rho = 0.667, p = 0.05). We found significant inverse correlations at 12 months between %change in TNFα and the following proteins: EPHX2 and ACE (for both rho = -0.783, p = 0.013). We also found significant inverse correlations between %change in CRP at 12 months and SHBG (rho = -0.759, p = 0.029), L1CAM (rho = -0.904, p = 0.002) and AMBP (rho = -0.684, p = 0.042). CONCLUSION: Using SWATH-MS, we identified several proteins that are involved in the inflammatory response whose levels change in patients who achieve remission of T2DM after bariatric surgery in tandem with changes in IL6, TNFα and/or CRP. Future studies are needed to clarify the underlying mechanisms in how MBS decreases low-grade inflammation.

Changes in the Proteome Profile of People Achieving Remission of Type 2 Diabetes after Bariatric Surgery.

Bariatric surgery (BS) results in metabolic pathway recalibration. We have identified potential biomarkers in plasma of people achieving type 2 diabetes mellitus (T2DM) remission after BS. Longitudinal analysis was performed on plasma from 10 individuals following Roux-en-Y gastric bypass (n = 7) or sleeve gastrectomy (n = 3). Sequential window acquisition of all theoretical fragment ion spectra mass spectrometry (SWATH-MS) was done on samples taken at 4 months before (baseline) and 6 and 12 months after BS. Four hundred sixty-seven proteins were quantified by SWATH-MS. Principal component analysis resolved samples from distinct time points after selection of key discriminatory proteins: 25 proteins were differentially expressed between baseline and 6 months post-surgery; 39 proteins between baseline and 12 months. Eight proteins (SHBG, TF, PRG4, APOA4, LRG1, HSPA4, EPHX2 and PGLYRP) were significantly different to baseline at both 6 and 12 months post-surgery. The panel of proteins identified as consistently different included peptides related to insulin sensitivity (SHBG increase), systemic inflammation (TF and HSPA4-both decreased) and lipid metabolism (APOA4 decreased). We found significant changes in the proteome for eight proteins at 6- and 12-months post-BS, and several of these are key components in metabolic and inflammatory pathways. These may represent potential biomarkers of remission of T2DM.

The relationship of childhood trauma and DNA methylation of NMDA receptor genes in first-episode schizophrenia.

Aim: We investigated GRIN1, GRIN2A, GRIN2B and LINE-1 DNA methylation in first-episode schizophrenia patients, their nonaffected siblings and age- and sex-matched controls testing for associations between DNA methylation and exposition to childhood trauma. Materials & methods: The Childhood Trauma Questionnaire evaluated the history of childhood trauma. Genomic DNA was bisulfite converted and pyrosequencing was employed to quantify DNA methylation. Results:GRIN2A, GRIN2B and LINE-1 DNA methylation was not associated with childhood trauma in patients, siblings and controls. Siblings with childhood trauma had hypermethylation at CpG1 of GRIN1 compared with siblings without trauma. Conclusion: Childhood trauma may influence GRIN1 methylation in subjects with liability to psychosis, but not in frank schizophrenia or controls.

Lay abstract Schizophrenia results from a combination of genetic and environmental influences. We investigated how some changes in genes can be silenced by a process named DNA methylation and may be linked to schizophrenia. For this reason, we hypothesized that childhood trauma, an environmental risk factor, would be associated with DNA methylation in schizophrenia patients compared with their unaffected siblings and controls. Our research has shown that altered blood DNA methylation of one candidate gene for psychiatric disorders may be associated with childhood trauma in the unaffected siblings of schizophrenia patients, but not in frank schizophrenia or controls. We believe that this gene plays an important role in helping identify vulnerable as well as resilient individuals to schizophrenia disorder.

Early-life stress effects on BDNF DNA methylation in first-episode psychosis and in rats reared in isolation.

Stressful events during early-life are risk factors for psychiatric disorders. Brain-derived neurotrophic factor (BDNF) is implicated in psychosis pathophysiology and deficits in BDNF mRNA in animal models of psychiatric disease are reported. DNA methylation can control gene expression and may be influenced by environmental factors such as early-life stress. We investigated BDNF methylation in first-episode psychosis (FEP) patients (n = 58), their unaffected siblings (n = 29) and community-based controls (n = 59), each of whom completed the Childhood Trauma Questionnaire (CTQ); BDNF methylation was also tested in male Wistar rats housed isolated or grouped from weaning. DNA was extracted from human blood and rat brain (prefrontal cortex and hippocampus), bisulphite-converted and the methylation of equivalent sequences within BDNF exon IV determined by pyrosequencing. BDNF methylation did not differ significantly between diagnostic groups; however, individuals who had experienced trauma presented higher levels of methylation. We found association between the mean BDNF methylation and total CTQ score in FEP, as well as between individual CpG sites and subtypes of trauma. No significant correlations were found for controls or siblings with child trauma. These results were independent of age, gender, body mass index, BDNF genotype or LINE-1, a measure of global methylation, which showed no significant association with trauma. Isolation rearing resulted in increased BDNF methylation in both brain regions compared to group-housed animals, a correlate of previously reported changes in gene expression. Our results suggest that childhood maltreatment may result in increased BDNF methylation, providing a mechanism underlying the association between early-life stress and psychosis.

CACNA1C methylation: association with cortisol, perceived stress, rs1006737 and childhood trauma in males.

Aim: We investigated morning cortisol, stress, rs1006737 and childhood trauma relationship with CACNA1C methylation. Materials & methods: Morning cortisol release, childhood trauma and perceived stress were collected and genotyping for rs1006737 conducted in 103 adult males. Genomic DNA extracted from saliva was bisulphite converted and using pyrosequencing methylation determined at 11 CpG sites within intron 3 of CACNA1C. Results: A significant negative correlation between waking cortisol and overall mean methylation was found and a positive correlation between CpG5 methylation and perceived stress. Conclusion:CACNA1C methylation levels may be related to cortisol release and stress perception. Future work should evaluate the influence of altered CACNA1C methylation on stress reactivity to investigate this as a potential mechanism for mental health vulnerability.

Circulating microRNA changes in patients with impaired glucose regulation.

We analysed if levels of four miRNAs would change after a lifestyle intervention involving dietary and exercises in prediabetes. MiRNAs previously shown to be associated with diabetes (Let-7a, Let-7e, miR-144 and miR-92a) were extracted from serum pre- and post-intervention. mRNA was extracted from fat-tissue for gene expression analyses. The intervention resulted in increased Let-7a and miR-92a. We found correlations between miRNAs and clinical variables (triglycerides, cholesterol, insulin, weight and BMI). We also found correlations between miRNAs and target genes, revealing a link between miR-92a and IGF system. A lifestyle intervention resulted in marked changes in miRNAs. The association of miRNAs with insulin and the IGF system (both receptors and binding proteins) may represent a mechanism of regulating IGFs metabolic actions.

Lifestyle intervention in individuals with impaired glucose regulation affects Caveolin-1 expression and DNA methylation.

AIMS: We investigated whether a lifestyle intervention could influence expression and DNA methylation of diabetes-related genes in patients with impaired glucose regulation (IGR), the results were compared to bariatric surgery, considering it an intensive change. METHODS: Twenty participants with IGR had adipose tissue biopsy and blood collected pre- and post-lifestyle (6 months) intervention; 12 obese patients had subcutaneous fat taken before and after bariatric surgery. RNA/DNA was extracted from all samples and underwent qPCR. DNA was bisulphite converted and 12 CpG sites of Caveolin-1 (CAV1) promoter were pyrosequenced. RESULTS: lifestyle intervention resulted in opposite direction changes in fat tissue and blood for CAV1 expression and DNA methylation and these changes were correlated between tissues, while no significative differences were found in CAV1 expression after bariatric surgery. CONCLUSIONS: Our findings suggest a role for CAV1 in modulating adipocyte function as a consequence of lifestyle changes, as exercises and diet. These results may provide insights into new therapeutic targets for diabetes prevention.

Neuromyelitis optica spectrum disorder in three generations of a Chinese family.

Neuromyelitis optica spectrum disorder is an inflammatory demyelinating disease that is largely sporadic. Familial disease has been reported in one or two generations, although its basis remains unknown. We report here three subjects meeting diagnostic criteria for NMOSD in one family: a father and son, and the maternal aunt of the father. Anticipation, of 27 years, was apparent in transmission from father to son. Aquaporin-4 antibodies were observed in the aunt but not the father and son, nor in other family members. A putative pathogenic mutation in the NECL2 gene was not found in this pedigree. This first report of NMOSD in three generations of one family underlines the heterogeneity of familial NMOSD.

GRIN2B promoter methylation deficits in early-onset schizophrenia and its association with cognitive function.

AIM: We investigated GRIN1 and GRIN2B promoter methylation in first-episode schizophrenia patients compared with siblings and controls, testing for correlations between DNA methylation, cognitive performance and clinical variables. MATERIALS & METHODS: Blood-derived DNA from all groups underwent bisulfite conversion and pyrosequencing to determine methylation at CpG sites within the GRIN1 and GRIN2B promoters and results were compared with the measure of global methylation LINE-1. RESULTS: We found hypomethylation among all CpGs analyzed within GRIN2B promoter in patients and greater LINE-1 methylation in patients and siblings. CpG4 was correlated to a measure of intellectual function. CONCLUSION: Changes in GRIN2B promoter methylation may represent an environmental influence contributing to glutamatergic dysfunction in psychosis and relate to lower cognitive performance in subjects with first-episode schizophrenia.

Data Independent Acquisition Mass Spectrometry Can Identify Circulating Proteins That Predict Future Weight Loss with a Diet and Exercise Programme.

We investigated biological determinants that would associate with the response to a diet and weight loss programme in impaired glucose regulation (IGR) people using sequential window acquisition of all theoretical fragment ion spectra (SWATH) mass spectrometry (MS), a data acquisition method which complement traditional mass spectrometry-based proteomics techniques. Ten women and 10 men with IGR underwent anthropometric measurements and fasting blood tests. SWATH MS was carried out with subsequent immunoassay of specific peptide levels. After a six-month intervention, 40% of participants lost 3% or more in weight, 45% of patients remained within 3% of their starting weight and 15% increased their weight by 3% or more. Hemoglobin A1c (HbA1C) level was reduced with weight loss with improvements in insulin sensitivity. SWATH MS on pre-intervention samples and subsequent principal component analysis identified a cluster of proteins associated with future weight loss, including insulin-like growth factor-II (IGF-II) and Vitamin D binding protein. Individuals who lost 3% in weight had significantly higher baseline IGF-II levels than those who did not lose weight. SWATH MS successfully discriminated between individuals who were more likely to lose weight and potentially improve their sensitivity to insulin. A higher IGF-II baseline was predictive of success with weight reduction, suggesting that biological determinants are important in response to weight loss and exercise regimes. This may permit better targeting of interventions to prevent diabetes in the future.

Parvalbumin promoter hypermethylation in postmortem brain in schizophrenia.

Deficits of brain parvalbumin (PV) are a consistent finding in schizophrenia and models of psychosis. We investigated whether this is associated with abnormal PV gene (PVALB) methylation in the brain in schizophrenia. Bisulfite pyrosequencing was used to determine cytosine (CpG) methylation in a PVALB promoter sequence. Greater PVALB methylation was found in schizophrenia hippocampus, while no differences were observed in prefrontal cortex. LINE-1 methylation, a measure of global methylation, was also elevated in both regions in schizophrenia, although the PVALB change was independent of this effect. These results provide the first evidence that PVALB promoter methylation is abnormal in schizophrenia and suggest that this epigenetic finding may relate to the reduction of PV expression seen in the disease.

Angiotensin AT1 receptors modulate the anxiogenic effects of angiotensin (5-8) injected into the rat ventrolateral periaqueductal gray.

Losartan and PD 123,319 are non-peptide angiotensin (Ang) receptor antagonists for the AT1 and AT2 subtypes of Ang II receptors, respectively. The tetrapeptide Ang (5-8) is the smallest Ang-peptide that elicits anxiogenic effects on unconditioned and conditioned experimental models upon injection into the ventrolateral column of the periaqueductal gray (vlPAG), and Ang (5-8) can be synthesized (from Ang II or Ang III) and inactivated in this mesencephalic structure. The vlPAG is also known to play a central role in mechanisms of fear and anxiety. We therefore utilized male Wistar rats to examine the effects of losartan and PD 123,319 injections, selective antagonists of the AT1 and AT2 receptors, respectively, into the vlPAG in the elevated plus-maze, a classic rat model of anxiety, and against the anxiogenic effect of Ang (5-8) (0.4 nmol/0.25µL) upon injection into the same region. The anxiolytic profile was dependent on the dose of intra-vlPAG losartan, whereas no effects on experimental anxiety were observed in the plus-maze following PD 123,319 injection. The anxiogenic effect of Ang (5-8) injection into the vlPAG remained unchanged in the PD 123,319-pretreated rats, but the effect did not occur in losartan-pretreated rats. The results led us to suggest that the anxiogenic effect of Ang (5-8) injection into the vlPAG may depend on the local activation of AT1, but not AT2 receptors. Activation of AT1 receptors in structures nearby vlPAG may be tonically involved in fear and experimental anxiety.

Increased DNA methylation in the parvalbumin gene promoter is associated with methamphetamine dependence.

AIM: The parvalbumin (PV)-containing subgroup of GABAergic neurons is particularly affected in schizophrenia and animal models of psychosis, including after methamphetamine (METH) administration. We investigated whether METH dependence and METH-induced psychosis may involve an effect on DNA methylation of the PVALB promoter. MATERIALS & METHODS: The methylation of a PVALB promoter sequence was determined in 100 METH-dependent and 102 control subjects using pyrosequencing. RESULTS: A significant increase in PVALB methylation was observed in METH dependence and METH-induced psychosis. No significant effect on long interspersed nucleotide element-1 methylation, a measure of global DNA methylation, was observed. CONCLUSION: These results demonstrate a specific association between elevated PVALB methylation and METH-induced psychosis. This finding may contribute to the GABAergic deficits associated with METH dependence.

Differential Expression of AMPA Subunits Induced by NMDA Intrahippocampal Injection in Rats.

Glutamate is involved in excitotoxic mechanisms by interacting with different receptors. Such interactions result in neuronal death associated with several neurodegenerative disorders of the central nervous system (CNS). The aim of this work was to study the time course of changes in the expression of GluR1 and GluR2 subunits of glutamate amino-acid-3-hydroxy-5-methyl-isoxazol-4-propionic acid (AMPA) receptors in rat hippocampus induced by NMDA intrahippocampal injection. Rats were submitted to stereotaxic surgery for NMDA or saline (control) microinjection into dorsal hippocampus and the parameters were evaluated 24 h, 1, 2, and 4 weeks after injection. The extension and efficacy of the NMDA-induced injury were evaluated by Morris water maze (MWM) behavioral test and Nissl staining. The expression of GluR1 and GluR2 receptors, glial fibrillary acidic protein (GFAP), and neuronal marker (NeuN) was analyzed by immunohistochemistry. It was observed the impairment of learning and memory functions, loss of neuronal cells, and glial proliferation in CA1 area of NMDA compared with control groups, confirming the injury efficacy. In addition, NMDA injection induced distinct changes in GluR1 and GluR2 expression over the time. In conclusion, such changes may be related to the complex mechanism triggered in response to NMDA injection resulting in a local injury and in the activation of neuronal plasticity.

Subchronic administration of phencyclidine produces hypermethylation in the parvalbumin gene promoter in rat brain.

AIM: A deficit in parvalbumin neurons is found in schizophrenia and several animal models of the disease. In this preliminary study, we determined whether one such model, phencyclidine (PCP) administration, results in changes in DNA methylation in the rat Pvalb promoter. MATERIALS & METHODS: DNA from hippocampus and prefrontal cortex from rats, which 6 weeks previously received either 2 mg/kg PCP or vehicle for 7 days, underwent bisulphite pyrosequencing to determine methylation. RESULTS: PCP administration induced significantly greater methylation at one of two Pvalb CpG sites in both prefrontal cortex and hippocampus, while no significant difference was found in long interspersed nucleotide element-1, a global measure of DNA methylation. CONCLUSION: Subchronic PCP administration results in a specific hypermethylation in the Pvalb promoter which may contribute to parvalbumin deficits in this animal model of psychosis.