Anti Kalda

DNA Methylation Regulates Cocaine-Induced Behavioral Sensitization in Mice

The behavioral sensitization produced by repeated cocaine treatment represents the neural adaptations underlying some of the features of addiction in humans. Cocaine administrations induce neural adaptations through regulation of gene expression. Several studies suggest that epigenetic modifications, including DNA methylation, are the critical regulators of gene expression in the adult central nervous system. DNA methylation is catalyzed by DNA methyltransferases (DNMTs) and consequent promoter region hypermethylation is associated with transcriptional silencing. In this study a potential role for DNA methylation in a cocaine-induced behavioral sensitization model in mice was explored. We report that acute cocaine treatment caused an upregulation of DNMT3A and DNMT3B gene expression in the nucleus accumbens (NAc). Using methylated DNA immunoprecipitation, DNA bisulfite modification, and chromatin immunoprecipitation assays, we observed that cocaine treatment resulted in DNA hypermethylation and increased binding of methyl CpG binding protein 2 (MeCP2) at the protein phosphatase-1 catalytic subunit (PP1c) promoter. These changes are associated with transcriptional downregulation of PP1c in NAc. In contrast, acute and repeated cocaine administrations induced hypomethylation and decreased binding of MeCP2 at the fosB promoter, and these are associated with transcriptional upregulation of fosB in NAc. We also found that pharmacological inhibition of DNMT by zebularine treatment decreased cocaine-induced DNA hypermethylation at the PP1c promoter and attenuated PP1c mRNA downregulation in NAc. Finally, zebularine and cocaine co-treatment delayed the development of cocaine-induced behavioral sensitization. Together, these results suggest that dynamic changes of DNA methylation may be an important gene regulation mechanism underlying cocaine-induced behavioral sensitization.

Histone Deacetylase Inhibitors Modulates the Induction and Expression of Amphetamine-induced Behavioral Sensitization Partially Through an Associated Learning of the Environment in Mice

The behavioral sensitization produced by repeated amphetamine treatment may represent the neural adaptations underlying some of the features of psychosis and addiction in humans. Chromatin modification (specifically histone hyperacetylation) was recently recognized as an important regulator of psychostimulant-induced plasticity. We have investigated the effects of treatment with the histone deacetylase (HDAC) inhibitors butyric acid (BA, 630mg/kg, i.p.) and valproic acid (VPA, 175mg/kg, i.p.) on the psyhcostimulant locomotor sensitization induced by amphetamine (AMPH, 2.0mg/kg, i.p.). Neither BA nor VPA had locomotor effects alone, but both significantly potentiated the amphetamine-induced behavioral sensitization in mice. At the molecular level, VPA and amphetamine produced an increase of histone H4 acetylation in the striatum as detected by Western blot analysis, while co-treatment with VPA and AMPH produced an additive effect on histone H4 acetylation. We then administered the HDAC inhibitors after treatment with amphetamine for 8 days to establish locomotor sensitization. We found that repeated administration of VPA or BA for 6 days inhibited the expression of sensitized response following amphetamine challenge. Finally, in a context-specific model we studied the effect of HDAC inhibitors on amphetamine-induced association of the treatment environment (associative learning). We found that VPA and BA enhance the context-specificity of expression of amphetamine sensitization. Thus, HDAC inhibitors differentially modulate the induction and expression of amphetamine-induced effects. Together, these results suggest that dynamic changes in chromatin modification may be an important mechanism underlying amphetamine-induced neuronal plasticity and associative learning.

Additive effects of histone deacetylase inhibitors and amphetamine on histone H4 acetylation, cAMP responsive element binding protein phosphorylation and ΔFosB expression in the striatum and locomotor sensitization in mice

Histone deacetylase (HDAC) plays an important role in chromatin remodeling in response to a variety of neurochemical signalings and behavioral manipulations, and may be a therapeutic target for modulation of psychostimulant behavioral sensitization. In this study, we investigated the molecular interaction between histone deacetylase inhibitor (HDACi) and psychostimulant in vivo of mice after repeated treatment with the HDACi, butyric acid (BA) and valproic acid (VPA), alone or in combination with amphetamine. Repeated treatment with amphetamine produced HDACi-like effects: enhanced global histone H4 acetylation level by Western blot as well as specific histone H4 acetylation associated with fosB promoter by chromatin immunoprecipitation in the striatum. Conversely, repeated treatment with BA or VPA produced amphetamine-like effects: enhanced cAMP responsive element binding protein (CREB) phosphorylation at Ser(133) position and increased DeltaFosB protein levels in the striatum. Furthermore, co-administration of BA or VPA with amphetamine produced additive effects on histone H4 acetylation as well as CREB phosphorylation in the striatum. The interplay of HDAC and CREB was also supported by co-immunoprecipitation assays demonstrating that repeated treatment with VPA reduced the association of CREB and HDAC1 in the striatum. Finally, the additive effect of VPA/BA and amphetamine on histone H4 acetylation, phosphorylated CREB, and DeltaFosB was associated with potentiated amphetamine-induced locomotor activity. Thus, HDACi may interact additively with psychostimulants at both histone acetylation and CREB phosphorylation through the CREB:HDAC protein complex in the striatum to modulate DeltaFosB protein levels and psychomotor behavioral sensitization.

Dehydroepiandrosterone sulphate prevents oxygen–glucose deprivation-induced injury in cerebellar granule cell culture

Decreased levels of dehydroepiandrosterone sulphate have been hypothesized to contribute to increased vulnerability of the ageing or stressed human brain to ischemia. To help to address the question of whether of dehydroepiandrosterone sulphate has a possible neuroprotective effect against ischemic neuronal injury, we tested its effect on the neurodegeneration induced by oxygen-glucose deprivation in rat cultured cerebellar granule cells. Dehydroepiandrosterone sulphate added to the medium after injury demonstrated a neuroprotective effect with a median inhibitory concentration of 0.5 microM. At 10 microM concentration almost full neuroprotection was observed. Even more pronounced neuroprotective effect was found when dehydroepiandrosterone sulphate was added for 48h before injury. Furthermore, partial neuroprotection of dehydroepiandrosterone sulphate was also found against 1-methyl-4-phenylpyridinium, colchicine, glutamate and N-methyl-D-aspartate-induced toxicity. Further analysis demonstrated that dehydroepiandrosterone sulphate eliminated the apoptotic features of the oxygen-glucose deprivation-induced neuronal death: DNA fragmentation and nuclear condensation/fragmentation.Thus, our data suggest that dehydroepiandrosterone sulphate may have therapeutic potential in the prevention and treatment of ischemic/hypoxic neuronal damage. The neuroprotective action of dehydroepiandrosterone sulphate was inhibited by both a GABA(A) receptor-linked chloride channel agonist and an antagonist, pentobarbital and picrotoxin, respectively. It seems that GABA(A) receptor-mediated neuronal inhibition as well as neuronal excitation can reduce the neuroprotective action of dehydroepiandrosterone sulphate.

Medium transitory oxygen-glucose deprivation induced both apoptosis and necrosis in cerebellar granule cells

Recent experiments have shown that an ischemic insult can induce both necrosis and apoptosis. A series of experiments were designed to examine the potential induction of apoptosis by oxygen-glucose deprivation (OGD) in cerebellar granule cell culture. A medium OGD (90 min) induced apoptosis in cell culture, with maximal effect 12 h after exposure, as indicated by following morphological (TdT-mediated dUTP-biotin nick end-labeling) and biochemical markers (DNA oligonucleosomal fragmentation). Mitochondrial injury (MTT assay) was among the early effects we detected during and after OGD and it was correlated with the dynamics of TUNEL positive cells. The amount of LDH release from damaged cells, associated with necrosis was increased significantly 12 h after exposure. These results indicate that medium OGD induced a rapid (<12 h) mixture of apoptosis and necrosis, followed by mainly secondary necrosis.

Maternal separation is associated with DNA methylation and behavioural changes in adult rats

Early life stress is known to promote long-term neurobiological changes, which may underlie the increased risk of psychopathology. Maternal separation (MS) is used as an early life stressor that causes profound neurochemical and behavioural changes in the pups that persist into adulthood. However, the exact mechanism of how MS alters these behavioural changes is not yet understood. Epigenetic modifications, such as DNA methylation, are critical regulators of persistent gene expression changes and may be related to behavioural disorders. The aim of the present study was to investigate whether early life stress on rats could alter cocaine-induced behavioural sensitisation in adulthood via aberrant DNA methylation. We have three main findings: (1) MS increased DNA methyltransferases (DNMTs) expression in the nucleus accumbens (NAc) of infant and adult rats; (2) MS induced DNA hypomethylation on a global level in the NAc, and hypermethylation of the promoter regions of the protein phosphatase 1 catalytic subunit (PP1C) and adenosine A2Areceptor (A2AR) genes, which was associated with their transcriptional downregulation in the NAc; (3) MS-induced molecular changes paralleled an increased response to cocaine-induced locomotor activity and exploratory behaviour in adult rats. Thus, our results suggest that stressful experiences in early life may create a background, via aberrant DNA methylation, which promotes the development of cocaine-induced behavioural sensitisation in adulthood.

Neuroprotective action of group I metabotropic glutamate receptor agonists against oxygen-glucose deprivation-induced neuronal death.

The metabotropic glutamate receptor (mGluR) non-selective agonist (1S,3R)-1-aminocycloheptane-trans-1,3-dicarboxylic acid [(1S, 3R)ACPD] and group I selective receptor agonist 3, 5-dihydrophenylglycine (DHPG) effectively attenuated oxygen-glucose deprivation (OGD)-induced death of the cultured cerebellar granule cells. Furthermore, (1S,3R)ACPD (100 microM) reduced the number of apoptotic cells. Antiapoptotic action of (1S,3R)ACPD was prevented by the group I selective antagonist (RS)-1-aminoindan-1, 5-dicarboxylic acid (AIDA, 100 microM) and protein kinase C (PKC) inhibitor bisindolylmaleimide (BMI, 1 microM).

Dehydroepiandrosterone with other neurosteroids preserve neuronal mitochondria from calcium overload

This current study was designed to test whether the dehydroepiandrosterone (DHEA) and other neurosteroids could improve mitochondrial resistance to ischemic damage and cytoplasmic Ca(2+) overload. To imitate these mechanisms at mitochondrial level we treated the saponin permeabilized neurons either with the respiratory chain inhibitor, 1-methyl-4-phenylpyridinium or raised free extra-mitochondrial [Ca(2+)]. Loss of mitochondrial membrane potential (as an indicator of loss of function) was detected by JC-1. The results demonstrate that DHEA partly prevented Ca(2+) overload induced loss of mitochondrial membrane potential but not the loss of potential induced by the inhibitor of the respiratory chain. A similar effect was observed in the presence of other neurosteroids, pregnenolone, pregnanolone and allopregnanolone. DHEA inhibited also the Ca(2+) accumulation to the mitochondria in the presence of Ca(2+) efflux inhibitors. Thus, in the present work we provide evidence that DHEA with several other neurosteroids protect the mitochondria against intracellular Ca(2+) overload by inhibiting Ca(2+) influx into the mitochondrial matrix.

Schizophrenia-like phenotype of polysialyltransferase ST8SIA2-deficient mice

Posttranslational modification of the neural cell adhesion molecule (NCAM) by polysialic acid (polySia) is crucial for nervous system development and brain plasticity. PolySia attachment is catalyzed by the polysialyltransferases (polySTs) ST8SIA2 and ST8SIA4, two enzymes with distinct but also common functions during neurodevelopment and in the adult brain. A growing body of evidence links aberrant levels of NCAM and polySia as well as variation in the ST8SIA2 gene to neuropsychiatric disorders, including schizophrenia. To investigate whether polyST deficiency might cause a schizophrenia-like phenotype, St8sia2−/− mice, St8sia4−/− mice and their wildtype littermates were assessed neuroanatomically and subjected to tests of cognition and sensorimotor functions. St8sia2−/− but not St8sia4−/− mice displayed enlarged lateral ventricles and a size reduction of the thalamus accompanied by a smaller internal capsule and a highly disorganized pattern of fibers connecting thalamus and cortex. Reduced levels of the vesicular glutamate transporter VGLUT2 pointed towards compromised glutamatergic thalamocortical input into the frontal cortex of St8sia2−/− mice. Both polyST-deficient lines were impaired in short- and long-term recognition memory, but only St8sia2−/− mice displayed impaired working memory and deficits in prepulse inhibition. Furthermore, only the St8sia2−/− mice exhibited anhedonic behavior and increased sensitivity to amphetamine-induced hyperlocomotion. These results reveal that reduced polysialylation in St8sia2−/− mice leads to pathological brain development and schizophrenia-like behavior. We therefore propose that genetic variation in ST8SIA2 has the potential to confer a neurodevelopmental predisposition to schizophrenia.

S-adenosylmethionine modifies cocaine-induced DNA methylation and increases locomotor sensitization in mice

Several studies suggest that individual variability is a critical component underlying drug addiction as not all members of a population who use addictive substance become addicted. There is evidence that the overall epigenetic status of a cell (epigenome) can be modulated by a variety of environmental factors, such as nutrients and chemicals. Based on these data, our aim was to investigate whether environmental factors like S-adenosylmethionine (SAM) via affecting epigenome could alter cocaine-induced gene expression and locomotor sensitization in mice. Our results demonstrate that repeated SAM (10 mm/kg) pretreatment significantly potentiated cocaine-induced locomotor sensitization. Using mouse nucleus accumbens (NAc) tissue, whole-genome gene expression profiling revealed that repeated SAM treatment affected a limited number of genes, but significantly modified cocaine-induced gene expression by blunting non-specifically the cocaine response. At the gene level, we discovered that SAM modulated cocaine-induced DNA methylation by inhibiting both promoter-associated CpG-island hyper- and hypomethylation in the NAc but not in the reference tissue cerebellum. Finally, our in vitro and in vivo data show that the modulating effect of SAM is in part due to decreased methyltransferase activity via down-regulation of Dnmt3a mRNA. Taken together, our results suggest that environmental factors that affect the NAc-cell epigenome may alter the development of psychostimulant-induced addiction and this may explain, at least partly, why some individuals are more vulnerable to drug addiction.