text:
GABAergic promoter hypermethylation as a model to study the neurochemistry of schizophrenia vulnerability.
ABSTRACT The neuronal GABAergic mechanisms that mediate the symptomatic beneficial effects elicited by a combination of antipsychotics with valproate (a histone deacetylase inhibitor) in the treatment of psychosis (expressed by schizophrenia or bipolar disorder patients) are unknown. This prompted us to investigate whether the beneficial action of this combination results from a modification of histone tail covalent esterification or is secondary to specific chromatin remodeling. The results suggest that clozapine, or sulpiride associated with valproate, by increasing DNA demethylation with an unknown mechanism, causes a chromatin remodeling that brings about a beneficial change in the epigenetic GABAergic dysfunction typical of schizophrenia and bipolar disorder patients.
Conference Paper: Metabolic Imbalance Associated with DNA Hypomethylation and Oxidative DNA/Protein Damage In Children with Autism
S. J. James, S. Melnyk, G. J. Fuchs, M. Lopez, S. Kahler, J. Fussell, L. Seidel, O. Pavliv, B. J. Bellando[Show abstract] [Hide abstract]
ABSTRACT: Background: Oxidative stress and abnormal DNA methylation have been implicated in the pathophysiology of autism. The metabolic pathology of autism is relatively unexplored although metabolic imbalance is implicated in the pathogenesis of multiple other neurobehavioral disorders. An abnormal accumulation or deficit of specific metabolites in a defined pathway can provide clues into relevant candidate genes and/or environmental exposures. In addition, the identification of precursor-product metabolite imbalance can inform targeted intervention strategies to restore metabolic balance and potentially improve symptoms of autism. We have investigated metabolic pathways essential for cellular methylation and antioxidant capacity and the functional impact of metabolic imbalance on genome-wide DNA hypomethylation and protein/DNA oxidative damage in children with autism. These metabolic pathways regulate the distribution of precursors for DNA synthesis (proliferation), DNA methylation (epigenetic regulation of gene expression) and glutathione synthesis (redox/antioxidant defense capacity). Previously, we reported that the metabolic profile of many children with autism is consistent with reduced methylation capacity and a more oxidized microenvironment. Objectives: To determine whether methylation and antioxidant metabolic profile differs between case children, unaffected siblings, and age-matched control children and to determine whether the metabolic imbalance is accompanied by DNA hypomethylation and protein/DNA oxidative damage. Methods: Subjects included 162 children, ages 3-10, who were participants in the autism IMAGE study (Integrated Metabolic And Genomic Endeavor) at Arkansas Children's Hospital Research Institute. The IMAGE cohort is comprised of 162 children including of 68 case children, 54 age-matched controls and 40 unaffected siblings. Children with autistic disorder were diagnosed using DSM-IV (299.0), ADOS and/or CARS >30. Fasting plasma samples were analyzed for folate-dependent transmethylation and transsulfuration metabolites and 3-nitrotyrosine (oxidized protein derivative) using HPLC with electrochemical detection. Genome-wide DNA methylation (as %5-methylcytosine) and the oxidized DNA adduct 8-oxo-deoxyguanine were quantified with Dionex HPLC-UV system coupled to an electrospray ionization (ESI) tandem mass spectrometer. Results: In a pair-wise comparison, the overall metabolic profile of the unaffected siblings differed significantly from their autistic siblings but was not different from unrelated control children. In addition, we report new evidence of genome-wide DNA hypomethylation (epigenetic dysregulation) and oxidative protein/DNA damage in children with autism that was not present in their paired siblings or in unaffected control children. Conclusions: These data indicate that the deficit in antioxidant and methylation capacity is autism-specific and is associated with DNA hypomethylation (epigenetic dysregulation) and oxidative damage. Further, these results suggest a plausible mechanism by which environmental stressors might modulate the genetic predisposition to autism. Acknowledgement: This research was supported with funding from the National Institute of Child Health and Development (RO1 HD051873; SJJ) and Department of Defense (AS073218P1; SJJ)International Meeting for Autism Research 2011; 05/2011 -
Article: Differential regulation of MeCP2 and PP1 in passive or voluntary administration of cocaine or food.
Sarah Pol Bodetto, Pascal Romieu, Maxime Sartori, Carolina Tesone-Coelho, Monique Majchrzak, Alexandra Barbelivien, Jean Zwiller, Patrick Anglard[Show abstract] [Hide abstract]
ABSTRACT: Cocaine exposure induces changes in the expression of numerous genes, in part through epigenetic modifications. We have initially shown that cocaine increases the expression of the chromatin remodeling protein methyl-CpG binding protein 2 (MeCP2) and characterized the protein phosphatase-1Cβ (PP1Cβ) gene, as repressed by passive i.p. cocaine injections through a Mecp2-mediated mechanism involving de novo DNA methylation. Both proteins being involved in learning and memory processes, we investigated whether voluntary cocaine administration would similarly affect their expression using an operant self-administration paradigm. Passive and voluntary i.v. cocaine intake was found to induce Mecp2 and to repress PP1Cβ in the prefrontal cortex and the caudate putamen. This observation is consistent with the role of Mecp2 acting as a transcriptional repressor of PP1Cβ and shows that passive intake was sufficient to alter their expression. Surprisingly, striking differences were observed under the same conditions in food-restricted rats tested for food pellet delivery. In the prefrontal cortex and throughout the striatum, both proteins were induced by food operant conditioning, but remained unaffected by passive food delivery. Although cocaine and food activate a common reward circuit, changes observed in the expression of other genes such as reelin and GAD67 provide new insights into molecular mechanisms differentiating neuroadaptations triggered by each reinforcer. The identification of hitherto unknown genes differentially regulated by drugs of abuse and a natural reinforcer should improve our understanding of how two rewarding stimuli differ in their ability to drive behavior.The International Journal of Neuropsychopharmacology 06/2014; 17(12):1-14. · 5.26 Impact Factor - SourceAvailable from: Chi-Yu Lai[Show abstract] [Hide abstract]
ABSTRACT: Psychiatric disorders are among the most debilitating of all medical illnesses. Whilst there are drugs that can be used to treat these disorders, they give sub-optimal recovery in many people and a significant number of individuals do not respond to any treatments and remain treatment resistant. Surprisingly, the mechanism by which psychotropic drugs cause their therapeutic benefits remain unknown but likely involves the underlying molecular pathways affected by the drugs. Hence, in this review, we have focused on recent findings on the molecular mechanism affected by antipsychotic, mood stabilizing and antidepressant drugs at the levels of epigenetics, intracellular signalling cascades and microRNAs. We posit that understanding these important interactions will result in a better understanding of how these drugs act which in turn may aid in considering how to develop drugs with better efficacy or increased therapeutic reach.Clinical Psychopharmacology and Neuroscience 08/2014; 12(2):94-110.
Page 1
© 2009 Expert Reviews Ltd
For several decades the biochemical targets to
monitor the brain molecular changes associated
with psychosis were the monoamines dopamine,
serotonin and norepinephrine [1]. Recently, an
emerging new target appears to be the GABAergic
neurotransmission [2]. Hence, the overarching
hypothesis guiding current research in schizophre-
nia (SZ) has been focused on a dysfunction of the
telencephalic GABAergic system, which is consid-
ered a key component in the pathophysiology of
this psychiatric disorder [2]. Consistent with this
new strategy, we and others have shown that the
expression of glutamic acid decarboxylase (GAD67),
reelin [2–6] and other genes [4,7–10] is downregu-
lated by aberrant epigenetic events in cortical, hip-
pocampal and basal ganglia GABAergic neurons
of SZ postmortem brains [9–15]. This downregu-
lation is among the most robust and consistent
findings reported by various groups in studies of
postmortem brains of SZ patients. Owing to the
prominent role of GABAergic neurotransmission
in the maintenance of the normal function of tel-
encephalic pyramidal circuits, we have inferred
that, very likely, reelin and GAD67 downregulation
reflects a brain molecular mechanism underlying
SZ and bipolar (BP) disorder symptomatology.
Reelin, an extracellular matrix protein, is pref-
erentially synthesized and secreted by GABAergic
neurons in the cortex, hippocampus and caudate
putamen. Upon secretion into the extracellular
matrix, reelin adheres to the dendritic shafts and
surrounds the dendritic spines of cortical pyrami-
dal neurons [2,16]. This protein, perhaps by imping-
ing on postsynaptically located apolipoprotein E2,
very large-density lipoprotein or integrin receptors
[17,18], modulates event-related protein synthesis,
including that of the immediate-early gene activ-
ity regulated cytoskeletal protein (Arc) [18], and
induces tyrosine phosphorylation of NMDA-
selective glutamate receptor subunit-modulating
NMDA receptor activity, markedly enhancing
long-term potentiation associated with cognitive
function [17]. Additionally, a reelin deficiency, such
as that observed in the brain of the heterozygous
reeler mouse, results in a net decrease of pyramidal
neuron dendritic spine expression in the frontal
cortex and hippocampus [2,19].
Reelin and GAD67 promoter regions are
embedded in large CpG islands and express
methylation that regulates the transcriptional
activity of these genes [13,14]. The efficiency of
this regulation can be underscored by stating
Erminio Costa†,
Ying Chen, Erbo Dong,
Dennis R Grayson,
Marija Kundakovic,
Ekrem Maloku,
William (Brad) Ruzicka,
Rosalba Satta,
Marin Veldic,
Adrian Zhubi and
Alessandro Guidotti
†Author for correspondence
Department of Psychiatry,
University of Illinois at Chicago,
1601 Taylor, Chicago,
Tel.: +1 312 413 4591
Fax: +1 312 413 4569
The neuronal GABAergic mechanisms that mediate the symptomatic beneficial effects elicited
by a combination of antipsychotics with valproate (a histone deacetylase inhibitor) in the
treatment of psychosis (expressed by schizophrenia or bipolar disorder patients) are unknown.
This prompted us to investigate whether the beneficial action of this combination results from
a modification of histone tail covalent esterification or is secondary to specific chromatin
remodeling. The results suggest that clozapine, or sulpiride associated with valproate, by
increasing DNA demethylation with an unknown mechanism, causes a chromatin remodeling
that brings about a beneficial change in the epigenetic GABAergic dysfunction typical of
schizophrenia and bipolar disorder patients.
Keywords: antipsychotic • chromatin remodeling • DNA demethylase • DNA methyltransferase inhibitor
• GABAergic neurotransmission • GAD67 • histone deacetylase inhibitor • psychosis • reelin • valproate
GABAergic promoter
hypermethylation as a model
to study the neurochemistry of
schizophrenia vulnerability
Expert Rev. Neurother. 9(1), 87–98 (2009)
For reprint orders, please contact email@example.com
Page 2
Expert Rev. Neurother. 9(1), (2009)
Cortical layers
Neurons (mm3 × 103)
NPS DNMT1 mRNA
NPS DNMT3a mRNA
SZP DNMT1 mRNA
SZP DNMT3a mRNA
Costa, Chen, Dong et al.
that, when the human reelin promoter is hypomethylated in vitro,
reelin expression levels can increase by up to 80-fold; on the con-
trary, when the reelin promoter is hyper methylated, for example,
by the addition of methionine to the culture medium, the reelin
expression level is decreased significantly [14]. Noh et al. demon-
strated that, in primary cortical cultures of GABAergic neurons,
a reduction of DNA methyltransferase (DNMT; cytosine-5)
expression facilitated by antisense technology blocks methionine-
induced reelin and GAD67 promoter hypermethylation and reelin
mRNA downregulation [20].
Grayson et al. reported that, in the frontal cortex of SZ
patients, the reelin promoter regions
containing PAX6 binding sites are
hypermethylated [13]. Abdolmaleky et
al. found a similar hypermethylation
further upstream proximal to the CREB
regulatory element [15]. These data are
in keeping with the hypothesis that the
decrease of reelin and GAD67 or other
genes expressed in cortical GABAergic
neurons of psychotic patients may be
mediated by cytosine-5 hypermethyla-
tion in the respective promoters elicited
by the increased expression of DNMTs
(Table 1) [21].
Genetic versus epigenetic
mechanisms in the GABAergic
dysfunction expressed by SZ & BP
disorder patients
Although a downregulation of reelin,
GAD67 and other genes expressed in
GABAergic neurons of psychotic patients
could reflect a genetic linkage, it has been
suggested that the complex inheritance
and selective pathology in GABAergic
neurons may be better explained by the
action of epigenetic factors on susceptible
candidate genes [10,21,22].
There is substantial evidence suggesting that the downregula-
tion of reelin, GAD67 and other promoters expressed in GABAergic
neurons of SZ and BP disorder patients is not caused by:
A genetic defect. The reelin and •?
some 7q22 [23] and 2q31 [24], respectively; neither region has
been clearly implicated in SZ pathogenesis;
GAD67 genes map to chromo-
A highly conserved single nucleotide polymorphism within
the regulatory regions of the genes. In fact, only in few cases
has this polymorphism been associated with an increased risk
of psychotic symptoms [25–27] but, in a number of other stud-
ies [28,29], no association has been reported. Although the
polymorphism detected in the reelin and GAD67 regulatory
regions may be insufficient per se to alter gene expression, it
may sensitize these genes to the influence of an epigenetic
regulation by environmental factors, toxins, drugs or
viruses [9,25];
The loss of GABAergic neurons. In fact, studies describing the
downregulation of reelin and GAD67 have found no changes in
the expression of other specific genes of GABAergic neurons,
for example GAD65 [3,4].
On the contrary, accumulating evidence suggests that the
downregulation of reelin, GAD67 and other genes in SZ and BP
patients is very likely mediated by epigenetic hypermethylation
of the related promoters. In fact, it has been shown in SZ and BP
patients that there is:
Table 1. DNA methyltransferase 1 mRNA is
overexpressed but glutamic acid decarboxylase
and reelin mRNAs are underexpressed in layer I
cortical GABAergic interneurons of schizophrenic
patients' brains.
10 ± 1.6
45 ± 11
3.3 ± 0.07 0.14 ± 0.03
7 ± 0.9
47 ± 15
24 ± 4.3*
14 ± 4.7*
1.2 ± 0.2*
8.6 ± 7.4
48 ± 9.5
0.15 ± 0.07
Values are expressed as the ratio of fmol of DNA methyltransferase 1, glutamic
acid decarboxylase or reelin mRNA/pmol G3PDH mRNA. Each value is the mean
± standard deviation of three to six samples.
*p < 0.05 compared with the respective samples of the NPS group.
L: Layer; NPS: Nonpsychotic patients; SZ: Schizophrenia.
Reprinted with permission from [12].
Figure 1. Density of DNMT1 and DNMT3a mRNA-positive neurons in Brodmann's
area 10 from NPS and SZPs. Each bar represents the mean ± standard error of eight
NPS (filled bars) or seven SZPs (checkered bars). The difference between SZPs and NPS
was calculated by t-test.
*Statistically significant difference when SZPs are compared with NPS.
DNMT1 mRNA-positive neurons:
LI: p = 0.02; LII: p < 0.01; LIII: p = 0.04; LIV: p = 0.03; LV: p = not significant;
LVI:p = not significant.
DNMT3a mRNA-positive neurons:
LI: p = 0.01; LII: p < 0.01; LIII: p = not significant; LIV: p < 0.01; LV: p = not significant;
LVI: p = not significant.
DNMT: DNA methyltransferase; L: Layer; NPS: Nonpsychotic patients;
SZP: Schizophrenic patients.
For experimental details see [11].
Page 3
promoter CpG
promoter CpG
GABAergic promoter hypermethylation
An increased number of cortical
or caudate–putamen GABAergic neurons
staining positive for DNMT1 and
DNMT3a mRNA and proteins [11,12], the
enzymes that catalyze the methylation of
promoter cytosine (Figure 2);
(Figure 1)
An increased level of
onine (SAM) in the prefrontal cortex
(PFC) [30]. SAM is the universal methyl
donor utilized by the catalytic activity
of DNMTs;
The presence of reelin and other
GABAergic gene promoters that are
hypermethylated in the brain of SZ and
BP patients [10,13–15].
Neuronal molecular mechanisms in
the epigenetic regulation of
gene expression
In multicellular organisms, the cells of various tissues are spe-
cialized to perform distinct functions. They are differentiated
for this purpose and this process is accompanied by distinct
patterns of gene expression. Thus, despite the fact that, with
few exceptions, every cell in an organism has identical DNA, a
huge number of distinct transcriptional profiles are created and
maintained. Among the most important in controlling these
processes are the epigenetic factors that establish and maintain
a metastable and non-Mendelian heritable gene expression in
the absence of DNA sequence changes (Figure 3).
In psychiatric patients, the importance of epigenetic factors
in non-Mendelian inheritance of complex disorders such as
SZ, BP disorder and autism is exemplified by the 50% morbid-
ity discordance observed in monozygotic twins, despite their
identical DNA sequences [22]. This phenomenon has been
explained as an effect of epigenetic (environmental) factors
that likely induce an aberrant epigenetic chromatin remodeling,
causing morbidity in one of the two genetically predisposed
monozygotic twins [22].
Fundamental to the concept of epigenetic modulation of
gene expression is the fact that, in the nucleus, DNA is packed
around a histone octamer known as a nucleosome that forms a
highly regulated and complex DNA–protein structure known
as chromatin. Along a single chromosome, various regions of
DNA can be packed in two different extreme configurations
of chromatin:
A highly condensed tightly coiled chromatin referred to as het-•?
erochromatin. In this structural conformation, DNA is inac-
cessible to transcription factor regulation, therefore can be
considered transcriptionally inactive;
A relaxed, loosely coiled chromatin, referred to as euchromatin.
The DNA in euchromatin is readily accessible to transcription
factors and is transcriptionally active.
The molecular mechanisms contributing to functional epige-
netic events include DNA promoter methylation and chromatin
remodeling, including various covalent histone tail modifications
that contribute to specific aspects of DNA function.
DNA promoter methylation
In the mammalian genome, DNA methylation specifically occurs
on carbon in position 5 of the cytosine residues embedded in
CpG dinucleotides. This methylation is a process catalyzed by a
family of brain DNMT enzymes including DNMT1, 3a and 3b.
These enzymes transfer a methyl group from SAM to the carbon
5 of cytosine. Hypermethylation of cytosines embedded in CpG
dinucleotides of reelin, GAD67 and other promoters is associated
with gene expression downregulation (Figure 3), whereas cyto-
sine promoter demethylation is associated with gene expression
upregulation (Figure 3).
It was thought that, in neurons, DNA methylation patterns were
finalized during development and remained stable thereafter [31].
However, there is now increasing evidence that, in adult neurons,
DNA methylation patterns change rapidly and, thus, DNA methyla-
tion provides a platform on which the environment can sculpt the
genome and affect neuronal phenotype profiles throughout life with-
out altering the genotype [32,33]. Recent data suggest that, in addi-
tion to DNMTs, neuronal promoter methylation can be regulated
by the activity of a putative DNA demethylase (see later) that can
remove a methyl group from carbon 5 of cytosine (Figure 3). Hence,
this evidence suggests that, in neurons, promoter methylation is a
dynamic process that can be altered in response to environmental
factors such as stress, drugs and various psychopathologies.
Histone tail covalent modifications
In addition to DNMT and DNA demethylase, DNA methylation
can be regulated indirectly by changes in the covalent modifications
of nucleosomal histone tails. For example, covalent histone-3 lysine
9,14 acetylation or histone-3 lysine 4 (tri)methylation is associated
Figure 2. DNMT-mediated promoter hypermethylation in adult neurons.
DNMT: DNA methytransferase; SAM: S-adenosyl methionine.
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Expert Rev. Neurother. 9(1), (2009)
Environmental signals
Signaling pathways
Methylated promoter
CpG islands
Unmethylated promoter
CpG islands
Expert Rev. Neurother. © Future Science Group (2009)
Costa, Chen, Dong et al.
with a downregulation of DNA promoter methylation and con-
tributes in changing the rate of gene expression in neurons by two
Reducing DNA affinity for histones and making the DNA •?
accessible to DNA demethylases and transcription factors;
Reducing the recruitment of chromatin repressor complexes
including histone deacetylases (HDACs), methyl-binding
protein (MeCP)-2, DNMTs and other corepressors [32–34].
Based on the original observation that there is a recruitment of
methyl DNA-binding proteins to methylated gene promoters, it
was inferred that, in neurons, DNA methylation regulates chro-
matin conformation. However, it is becoming clear that the rela-
tionship between chromatin remodeling and DNA methylation
is bidirectional. For example, the targeting of DNA methylating/
demethylating enzymes to gene promoters (Figure 3) may be regu-
lated by chromatin-modifying enzymes. Hence, signaling pathways
that activate chromatin-modifying enzymes could potentially result
in alterations of DNA methylation patterns. As we will discuss later,
this concept has significant important functional implications for
the use of HDAC inhibitors (i.e., valproate [VPA]) or histone methyl
transferase activators (one recently discovered activator is clozapine
[CLZ]) [9] as drugs that may affect methylation of GABAergic
neuron promoters and cognate protein expression [9,35–37].
Aberrant epigenetic regulation of GABAergic neurons
in SZ & BP disorder patients
In the adult human brain, the expression of DNMTs is
restricted to GABAergic neurons
Central to the general hypothesis that a pathology of epigenetic
mechanisms is operative in the transcriptional downregulation
of reelin and GAD67 expression in SZ and BP patients is the
evidence that DNMTs (DNMT1 and DNMT3a) in the cor-
tex and basal ganglia are highly expressed in GABAergic neu-
rons (Figure 1 & Table 1) [11,12]. DNMT1 is the most frequently
expressed protein and it functions as a maintenance methylating
enzyme, while DNMT3a, which is also abundant, is a protein
with de novo methylating activity. In contrast to DNMT1 and
DNMT3a, DNMT3b is expressed at very low levels in adult
human PFC (Veldic M, Pers. Comm.).
Since the expression and the activity of DNMTs in somatic cells
generally decreases after cell division and differentiation, it is sur-
prising that, in the adult human brain, DNMT1 and DNMT3a
are highly expressed in a subset of terminally differentiated
GABAergic neurons. In a recent histochemical and biochemical
laser-assisted microdissection study, we have provided evidence
that DNMT1 and DNMT3a are highly expressed in cortical
layer I, II and IV GABAergic interneurons. The expression of
both proteins is also found in the caudate–putamen GABAergic
medium spiny neurons but is virtually
absent in pyramidal neurons [11,12]. Since
cortical GABAergic neurons are terminally
differentiated cells, it is likely that, in these
cells, DNMTs play a role in important
functions beyond the well-known DNA
methylation function at replication foci in
dividing cells.
In SZ & BP disorder patients, DNMT
overexpression is restricted to a
subset of GABAergic neurons
DNA methyltransferase has been
shown to be increased in GABAergic
caudate–putamen medium spiny neurons,
and in GABAergic interneurons of the
PFC layers I, II and IV in SZ patients with
psychosis (Figure 1). This upregulation (an
increase of two-to-threefold over control)
occurs in the same neurons expressing a
decrease of reelin and GAD67 (Table 1). In
addition, in the PFC of the same patients,
the increase of DNMTs is associated with
an elevated (60–80%) level of SAM [30].
Such an increase was not detected in the
PFC of depressed patients. Since our
studies show that the concentrations (low
µmol range) of SAM in the PFC of con-
trol subjects are below the concentrations
required to saturate DNMT catalytic sites,
an increase in SAM levels as observed in
Figure 3. Schematic representation of GABAergic promoter methylation or
demethylation reactions triggered by a variety of environmental signals.
(A) Environmental signals may activate promoter methylation and repress gene
transcription. (B) Environmental signals may activate promoter demethylation and
activate gene transcription.
Page 5
GABAergic promoter hypermethylation
SZ and BP patients may reflect an increase of DNMT1 activ-
ity. Hence, the available data indicate that the brains of SZ
or BP patients may express reelin and GAD67 promoter hyper-
methylation and downregulation of the cognate proteins for
two reasons:
The increased expression of DNMTs that selectively occurs in •?
GABAergic neurons in the upper layer of the cortex that are
contiguous to apical dendrites
The increased availability of SAM that is not present in saturating
concentrations in proximity to DNMTs
Methionine regulation of epigenetic mechanisms
In addition to environmental factors including stress [38] and
maternal behavior [32,39], the administration of methyl donors
(Figure 3) has also been identified as an important epigenetic factor
contributing to the aberrant regulation of reelin, GAD67 and also
to other promoters. For example, folic acid intake affects agouti
gene expression in Avy/a mice and unbalance global methylation
levels of the genome [40]. Maternal methyl supplementation in
mice induces epigenetic variations, including DNA methylation
in offspring [41].
The concept that reelin and GAD67 expression deficits in SZ
are mediated via epigenetic promoter hypermethylation can be
related to clinical observations in the late 1960s, which demon-
strated that daily administration of high methionine doses could
exacerbate or even trigger a psychotic episode in 40–50% of SZ
patients [42] but not in nonpsychiatric patients. In the mammalian
brain, methionine functions as the precursor of SAM that in turn
serves as the principal methyl donor participating with DNMTs
to maintain the methylation patterns of GABAergic promot-
ers [35]. This hypothesis was tested in a methionine-induced
murine endophenotype model of SZ.
Mice treated for a protracted period with repeated doses of
methionine (5.2 mmol/kg subcutaneously twice a day for 15 days)
model some of the behavioral patterns (prepulse inhibition [PPI]
and social interaction deficits) and molecular neuropathologies
that are similar to SZ symptomatology [34–36], including:
Elevated PFC levels of SAM; •?
Reelin and GAD •?
67 promoter hypermethylation;
Downregulation of reelin and GAD •?
67 expression;
Recruitment of methyl-CpG-binding domain proteins (MeCP2
and MDB2) on hypermethylated reelin and GAD67 promoters,
which suggests that this recruitment may be required to elicit
reelin and GAD67 downregulation [34–36].
In these mice, the HDAC inhibitor VPA, which increases brain
levels of acetylated histones in cortical GABAergic neurons, also
prevents methionine-induced reelin and GAD67 promoter methyla-
tion, and downregulates the increase of MeCP2 binding to reelin
and GAD67 promoters [34–36]. VPA also either prevents or reverses
the decrease of reelin and GAD67 expression elicited by methionine
[35,36]. These data suggest that treatment with methionine may pro-
duce a mouse model of SZ because of the casual relationship among
elevated brain SAM levels, increased DNMT1 activity, reelin and
GAD67 promoter hypermethylation, and induction of behavioral
responses reminiscent of SZ symptomatology. Furthermore, these
findings suggest that DNA hypermethylation and the associated
chromatin remodeling may be critically important in mediating the
epigenetic downregulation of reelin and GAD67 expression detected
in cortical and other GABAergic interneurons of SZ patients.
Collectively, these data challenge the classical concept that DNA
(cytosine-5) methylation patterns remain stable in postmitotic
cells. Our findings with promoter hypermethylation of reelin and
GAD67 in GABAergic neurons of adult mice receiving methio-
nine [34–36], and the recent report by Weaver et al. on the epige-
netic reprogramming of glucocorticoid receptors in hippocampal
pyramidal neurons of methionine-treated rodents, suggest that
promoter methylation is a dynamic process mediated by DNMT1
or DNMT3a [43].
Evidence that the methionine-induced reelin, GAD67 and exon17
glucocorticoid receptor promoter hypermethylation can be pre-
vented or effectively reversed by VPA and other HDAC inhibitors
[33,39,43], perhaps inducing a DNA demethylation process [33,37,44],
supports the concept that, in addition to DNMT1 and DNMT3a, a
putative DNA (cytosine-5) demethylase may also play a pivotal role
in regulating the appropriate dynamic balance of DNA cytosine-5
methylation patterns in animals receiving methionine.
Pharmacological strategies to reduce reelin & GAD67
promoter hypermethylation in SZ patients
Based on the evidence that DNMT1 and DNMT3a expression is
increased in cortical GABAergic neurons of SZ patients, the logi-
cal and theoretical considerations depicted in Figure 3 suggest that
Table 2. Epigenetic drugs.
DNA methyltransferase inhibitors
HydralazineERK inhibition
NicotinenAChR stimulation
DNA demethylase activators
ValproateHDAC inhibition
MS-275HDAC inhibition
Trichostatin AHDAC inhibition
SulpirideHDAC inhibition
ClozapineHDAC inhibition
*Incoporated in DNA in the place of cytosines.
‡Intercalates into DNA.
HDAC: Histone deacetylase; nAChR: Nicotinic acetylcholine receptor.
Page 6
Expert Rev. Neurother. 9(1), (2009)
Costa, Chen, Dong et al.
a new approach for the treatment of SZ morbidity should address
the hypermethylation of reelin, GAD67 and other gene promoters
expressed in GABAergic neurons. This approach should include
the study of:
Inhibitors of DNMT1 catalytic activity;•?
Drugs that downregulate the expression of DNMT1 and
Drugs (VPA or sulpiride [SULP], alone or associated with
antipsychotics) that induce DNA demethylation (Table 2).
DNMT inhibitors
Most of the inhibitors of DNMT1 catalytic activity available today
are molecules that can be incorporated into the DNA of proliferat-
ing cells (Table 2) [45]. The most-studied compounds are the cytosine
analogs 5-azacytidine and zebularine. Both nucleoside analogs are
converted in vivo into deoxynucleotide triphosphate derivatives
and are incorporated into replicating DNA. DNMTs bind to the
nucleoside analogs and remain irreversibly trapped in the DNA,
leading to depletion of DNMT stores [45]. These two inhibitors
have been used with some success to downregulate cell division in
cancer cells in vitro and in vivo, but it is expected that they would
fail to act on postmitotic cells and therefore are expected to be
devoid of action in differentiated neurons. However, recent studies
in rats show that zebularin and 5-aza-cytidine injected locally into
the CA1 region of the hippocampus prevent the DNMT-induced
promoter hypermethylation of the 'memory suppressor gene' PPI
[38]. Unfortunately, 5-azacytidine and zebularine do not readily
cross the BBB and, at the high doses needed to reach the brain,
they elicit significant toxicity.
Another potent inhibitor of DNMT activity is doxorubicin.
This antibiotic derivative of antracycline intercalates into the
DNA, thus preventing the action of DNMT and reducing its
expression [46]. Doxorubicin has been tested in the treatment of
peripheral tumors, but it causes considerable toxicity because it
reduces mitochondrial oxidative phosphorylation and causes severe
cardiotoxicity [46]. In addition, this compound does not cross the
BBB readily and therefore cannot be considered a candidate to
pharmacologically reduce DNMT activity in the brain.
A prospective DNA methylation inhibitor active in differ-
entiated neurons is procainamide, a drug used to treat cardiac
arrhythmias that also inhibits DNA methylation. This com-
pound is a non-nucleoside competitive inhibitor of DNA meth-
ylation [47] and is expected to be an effective inhibitor of DNA
hypermethylation in neurons. In initial studies, we demonstrated
that, given to mice in doses of 0.29 mmol/kg subcutaneously
four-times a day, procainamide prevents the hypermethylation of
reelin and GAD67 promoters induced by protracted methionine
treatment (5.2 mmol/kg subcutaneously twice daily) [33].
Drugs that downregulate DNMT1 expression
The expression of several transcription factor binding sites on
DNMT promoters [48,49] suggests that their expression in neurons
can be regulated by both environmental stimuli and neuroactive
drugs. Therefore, to downregulate DNMT expression upregula-
tion with drugs, one should also consider non-nucleoside com-
pounds that either decrease DNMT1 expression or reduce its cata-
lytic activity. As reported in Table 2, a non-nucleoside drug proven
to be an effective indirect DNA demethylase inhibitor in cancer
chemotherapy is hydralazine. This drug is used to treat hyper-
tension and, in nontoxic doses, downregulates DNA methylation
in T cells by inhibiting the ERK pathway and decreasing DNMT
expression [50]. This drug should be tested in psychopathologies
once its ability to cross the BBB has been established.
Another small non-nucleoside molecule that should be consid-
ered is nicotine. We have recently found that nicotine downregu-
lates cortical DNMT1 mRNA and protein expression, suggest-
ing that nicotine abuse, which is very frequent in SZ patients,
may be motivated by an attempt to reduce the SZ symptoms
via a downregulation of the increase of brain DNMT1. In fact,
our results suggest that nicotine and selective agonists of α4/β2
nicotinic cholinergic receptors downregulate DNMT1 expres-
sion and increase GAD67 expression in cortical and hippocampal
GABAergic neurons [51].
HDAC inhibitors
A possible strategy to elicit a pharmacological normalization of the
reduced reelin or GAD67 expression in cortical GABAergic neurons
of SZ patients is to administer HDAC inhibitors to reduce DNMT1
accessibility to cytosine embedded in the promoter CpG islands of
these genes. The possible success of such a strategy is supported by
a report showing VPA administered in therapeutic doses to human
subjects increases acetylation of chromatin histones [52] and, at least
in mice and rats, this drug prevents the reelin and GAD67 promoter
hypermethylation induced by methionine [33–37].
Four major classes of HDAC inhibitors have been described
(Table 2): hydroxamic acids, short-chain fatty acids, cyclic
tetrapeptides and benzamides.
The hydroxamic acid derivative trichostatin A is the most
potent HDAC inhibitor available and acts in vitro in the
nanomolar concentration range. However, this compound
does not readily cross the BBB and, if given parentally, can-
not inhibit CNS HDACs. Suberoylanilide hydroxamic acid is
another hydroxamic acid derivative active in micromolar con-
centrations in vitro but it appears to be a weak inhibitor of brain
HDACs in vivo [53], presumably because of its short half-life
and its low BBB penetrability. Better results were obtained with
the benzamide derivatives MS-275 [33,54] and SULP [37]. When
injected in mice, both compounds are at least ten-times more
potent than VPA in increasing brain acetyl histone-3 content
and they are well tolerated when administered repeatedly during
the day for several days.
Interestingly, SULP is a dopamine D2 and D3 receptor antago-
nist with a low extrapyramidal side-effect liability. Sulpiride is
used in Europe as an effective antipsychotic to treat psychotic
exacerbation episodes in chronic SZ patients [55]. Because of its
HDAC inhibitory activity, its efficacy as an antipsychotic cannot
be ascribed exclusively to its D2, D3 receptor function inhibition
but can also involve the inhibition of HDACs.
Page 7
GABAergic promoter hypermethylation
It was previously assumed that the HDACs expressed in the
brain were approximately equally sensitive to different inhibitors
of these enzymes. While this may be the case for trichostatin A and
its derivatives, recent indirect evidence suggests that this might
not be the case for VPA and MS-275. For example, it has been
reported that class II HDACs are five-times less susceptible to
VPA inhibition than class I HDACs [53]. Furthermore, it has been
reported that, among the class I HDACs, HDAC1 is most sensitive
to MS-275, whereas HDAC3 and HDAC8 have a significantly
lower sensitivity [53,54]. This finding suggests that the development
of new HDAC inhibitors targeted to specific HDACs combined
with pertinent information on selective tissue expression of the
various molecular forms of HDACs may allow the preparation of
tailored HDAC inhibitor cocktails to be tested as pharmacological
interventions to treat psychotic episodes in SZ and BP disorders.
HDAC inhibitors & DNA demethylation
Recent studies suggest that GABAergic neurons, in addition to
DNMTs and SAM, also express an active DNA-demethylating
enzyme that may play an important role in determining the level
of gene-specific cytosine methylation [33,37,38]. Thus, to study
whether hypermethylated reelin and GAD67 promoters in the
brain can be demethylated by the action of DNA demethylase
expressed in the mouse brain, we tested whether reelin and GAD67
promoter hypermethylation induced by protracted methionine
administration is reversed after methionine withdrawal, and
whether the demethylation characteristics can be modified by
the association of VPA or MS-275 with antipsychotics.
In mice receiving methionine (5.2 mmol/kg subcutanously
twice daily for 7 days), reelin and GAD67 promoters isolated
from the frontal cortex were hypermethylated compared with
promoters extracted from glycine-treated controls [33,34]. This
hypermethylation declined by 50% after 6 days of methio-
nine withdrawal [33]. By contrast, when VPA (2 mmol/kg) was
given after termination of methionine treatment, it dramati-
cally accelerated the reelin and GAD67 promoter demethylation
that occurred in the following 48–72 h [33]. At the doses used
(0.25–2.2 mmol), VPA increased the content of nuclear acetyl-
histone 3 bound to reelin and GAD67 promoters by 40–50% [34],
suggesting that a histone code involving covalent histone tail
modification may facilitate the efficacy of DNA demethylation
by this drug.
The HDAC inhibitors (i.e., VPA and MS-275) ability to
demethylate reelin and GAD67 promoters could be facilitated
by an inhibition of DNMT activity, or the activation of a DNA
(cytosine-5) demethylase.
There are two sets of data that suggest that the accelerated
reelin and GAD67 promoter demethylation elicited by VPA or
MS-275 treatment could be directly or indirectly related to a
DNA demethylase induction but not to reduced DNMT1
Reduction of DNMT1 activity with procainamide fails to acti-
vate demethylation of the hypermethylated reelin and GAD67
promoters [33];
VPA fails to inhibit SAM biosynthesis and in fact may even
facilitate its biosynthesis [33], suggesting that an indirect reduc-
tion of DNMT activity due to a reduced availability of SAM
cannot be related to VPA-induced promoter demethylation.
Hence, we believe that, in mice, the mechanism by which the
recovery of reelin and GAD67 downregulation is accelerated by
VPA or MS-275 after methionine withdrawal could be related to
the induction of a putative DNA demethylase activity. These data
agree with a previous report by Detich et al. demonstrating that
VPA triggers a replication-independent DNA demethylation [44].
Since the biochemical identification of a putative DNA demethy-
lase in mammalian cells is still unclear [32], one cannot infer whether
the accelerated reelin and GAD67 promoter demethylation elicited by
HDAC inhibitors in the mouse frontal cortex is the result of DNA
demethylase activation or an induction of DNA demethylase biosyn-
thesis. The characterization of an inducible demethylation activity
in the mammalian brain is an attractive possibility for discovering a
new epigenetic mechanism that includes a putative DNA demethyla-
tion. Obviously, the identification of a DNA demethylase would be
essential to develop a new line of pharmacological intervention to
treat psychosis in SZ or BP disorder patients.
GABAergic gene promoter demethylation as a new
epigenetic target for antipsychotics
Published clinical studies infer that drugs such as VPA are effica-
cious in the treatment of SZ and BP disorder when used in combi-
nation with typical or atypical antipsychotics [56–58]. However, no
study has suggested the mechanisms underlying this interaction.
The symptomatic benefits elicited by a combination of VPA
and antipsychotics in the treatment of SZ prompted us to study
whether the targets of this drug combination could lead to changes
of specific chromatin remodeling events mediated by the activa-
tion of a DNA demethylase operative at selected hypermethylated
GABAergic promoters (i.e., reelin and GAD67).
Our experiments were carried out with various typical and
atypical antipsychotics, including haloperidol (HAL), CLZ,
SULP and olanzapine (OLZ). HAL (a butyrophenone deriva-
tive) is a typical antipsychotic that relieves the positive but not
the negative symptoms of SZ. In addition, it is a potent D2 recep-
tor blocker causing a high liability for extrapyramidal side effect
induction. CLZ (pyperazinyl-dibenzo-[1-4]-diazepine) and OLZ
(pyperazinyl-[1-5]-benzodiazepine) are two atypical antipsychot-
ics with low efficacy on negative symptoms. At the doses used as
anti psychotics, they have weak antagonistic action on D2 recep-
tors and therefore express low extrapyramidal side-effect liability.
SULP (aminosulfonyl-methoxybenzamide) is an atypical antip-
sychotic that also ameliorates the severity of negative symptoms
and, only at high doses, may act as a D3/D2 receptor antagonist
that may include extrapyramidal side effects. In a recent study, we
report that frontocortical and striatal reelin and GAD67 promoter
hypermethylation induced in mice by a 7-day methionine admin-
istration can be reversed by atypical antipsychotics (i.e., CLZ or
SULP) given either alone or coadministered with VPA, but not
by HAL or OLZ given either alone or with VPA [37].
Page 8
Expert Rev. Neurother. 9(1), (2009)
Clozapine (C)
Frontal cortex
Haloperidol (H)
Reelin promoter demethylation (%)
Costa, Chen, Dong et al.
As shown in Figure 4, CLZ (3.8–15 µmol/kg subcutaneously)
administered twice a day for 3 days elicits a dose-related increase
of frontocortical reelin promoter demethylation. Furthermore, at
every dose studied, CLZ or SULP (12.5–50 µmol/kg subcutane-
ously) synergistically enhance reelin promoter demethylation
elicited by a dose of VPA that, per se, only partially (30–35%)
increases promoter demethylation (Figure 4).
As opposed to CLZ and SULP, HAL (1.5–4 µmol/kg sub-
cutaneously) (Figure 4) and OLZ (4–16 µmol/kg subcutane-
ously) [37] failed to induce statistically significant changes even
when administered in combination with VPA.
To elicit a marked reelin promoter demethylation, CLZ or
the combination of CLZ and VPA must be given repeatedly
(multiple injections for 3 days); in fact, reelin promoter demeth-
ylation was not increased 2 h after a single injection of CLZ
(15 µmol/kg subcutaneously) or a single coadministration of
VPA (0.5 mmol/kg subcutaneously) plus CLZ (15 µmol/kg sub-
cutaneously). For example, after a single injection, the percentage
of reelin promoter methylation is 62 ± 5% in vehicle, 59 ± 8%
in CLZ alone and 47 ± 13% in VPA plus CLZ-treated mice
(n = 3) [37].
The association of VPA (0.5 mmol/kg subcutaneously) with
CLZ (15 µmol/kg subcutaneously twice daily) at a dose schedule
that induces reelin and GAD67 promoter demethylation in the
frontal cortex also exerts a similar effect in the striatum [37].
We used this brain area because its GABAergic neurons are a
possible target for the beneficial action of typical neuroleptics
in psychosis, and striatum expresses an almost homogeneous
population (∼90%) of GABAergic medium spiny neurons that
include reelin [11].
Promoter demethylation induced by coadministration of VPA
and CLZ is brain selective. In the liver, the reelin promoter is
constitutively hypermethylated (more than 80% of the cytosine
sites are methylated) and in fact its degree of methylation is
not significantly increased by methionine administration [37].
Moreover, in the liver, the methylation of reelin promoter
cytosines failed to be lowered by the coadministration of VPA
(0.5 mmol/kg twice a day for 3 days) or CLZ (15 µmol/kg
twice a day for 3 days) at doses that dramatically decrease reelin
promoter methylation in the frontal cortex and striatum of the
same mice [37].
Taken together, these data suggest that coadministration to
mice of well-tolerated, clinically relevant doses of VPA and CLZ
or VPA and SULP induces brain reelin and GAD67 promoter
demethylation, presumably by activating a DNA demethylase
that must be expressed in selected populations of cortical or stri-
atal GABAergic neurons. Nonetheless, such an activity cannot
be expected when VPA is associated with HAL or OLZ.
The similarities between the frontal cortex and striatum and
the difference of both tissues from the liver further suggest that
DNA demethylation induced by coadministration of clinically
relevant doses of VPA and CLZ, or VPA or SULP, is a process
restricted to selected populations of brain GABAergic neurons
and cannot be generalized to other tissues or cell types.
Expert commentary
Typical and atypical antipsychotics tar-
geted to dopamine, norepinephrine or
serotonin neurotransmission have rep-
resented a fundamental breakthrough in
the treatment of positive symptoms in SZ
and BP disorder patients. However, these
drugs are ineffective for treatment of nega-
tive symptoms and cognitive dysfunction.
Indeed, the advances made during the last
50 years, since the introduction of chlo-
rpromazine in the treatment of negative
symptoms or cognitive deficits in SZ and
BP disorder, have been minimal [1]. Thus,
there is a need to identify novel molecu-
lar targets with a well-defined role in the
pathophysiology of these illnesses.
Recent breakthroughs in the study of
aberrant epigenetic mechanisms opera-
tive in SZ and BP disorder, including an
increased expression of DNMT1, suggest
that promoter hypermethylation in CNS
GABAergic neurons could be the pathoge-
netic mechanism underlying specific symp-
toms, such as lack of motivation, anhedo-
nia and working memory and executive
function deficits.
Figure 4. Clozapine, alone or in combination with VPA, but not haloperidol
accelerates reelin promoter demethylation. Reelin promoter demethylation was
measured after 3 days treatment with C or C + VPA, or H and H + VPA following
methionine treatment termination.
Mice were pretreated for 7 days with methionine (5 mmol/kg subcutaneously, twice
daily). Data represent the mean ± standard error of three animals per group.
*p < 0.05; **p < 0.01 compared with vehicle.
C: Clozapine; H: Haloperidol; V: Vehicle; VPA: Valproate.
For methods see Dong et al. [33].
Page 9
GABAergic promoter hypermethylation
It is clear that presently available treatments for SZ and BP
disorder are not directly targeted to the molecular abnormalities
in the GABAergic dysfunction operative in these disorders. To
correct GABAergic neuron deficits, we propose the following two
principal strategies:
Enhancement of defective GABAergic transmission by drugs •?
acting as selective positive allosteric modulators of GABA action
at pertinent GABAA receptor subtypes [59];
Drugs acting to correct chromatin remodeling abnormalities •?
due to epigenetic dysregulation mechanisms (as described in
this review).
Elaborating on the first strategy, one may consider that benzo-
diazepines, which are devoid of intrinsic activity at GABAA
receptors, including α1 subunits, and act exclusively at GABAA
receptors expressing α2, α3, α5 subunit combinations, should
counteract the GABAergic signal-transduction deficit without
eliciting sedation, amnesia, tolerance or dependence liabilities.
Benzodiazepines acting at α1-expressing GABAA receptor sub-
types are prescribed to psychotic patients but one has to deal with
problems related to their sedative action. One benzodiazepine
devoid of intrinsic activity at GABAA receptors, including α1
subunits, but acting as a full allosteric modulator at α5, and per-
haps α2 and α 3 subunits, is imidazenil. This drug is anxiolytic
and anticonvulsant and, moreover, it fails to produce sedation or
amnesia [59]. Hence, we suggest that a combination of imidazenil
with antipsychotics should be considered and eventually tested
in the treatment of the GABAergic dysfunction operative in SZ
and BP disorders.
An alternative strategy to correct the GABAergic neuron
deficit in SZ and BP disorder patients may be to use drugs that
diminish the DNMT1 overexpression typical of these illnesses.
One can speculate that a protocol to treat SZ and BP disorders
may include procainamide (a competitive inhibitor of DNMTs)
or nicotine acetylcholine receptor agonists such as A-85380 or
varenicline to downregulate the expression of DNMT1 [51].
These agonists may be administered with VPA used to inhibit
HDACs and to activate chromatin transcriptional activity of
selective GABAergic genes including, for example, GAD67, which
is downregulated in SZ and BP disorders [51]. This is probably
the reason why VPA is presently prescribed with antipsychotics
as an augmentation strategy to treat multiple symptoms of the
SZ syndrome.
The site of action of antipsychotics is not clear. However,
recent studies suggest that they may act on nuclear chromatin
remodeling in GABAergic neurons [37]. Hence, antipsychotic
drugs and their coadjuvants (imidazenil and HDAC inhibitors)
should be studied in animal experiments to evaluate their puta-
tive action on chromatin remodeling, and their ability to correct
the GABAergic downregulation typical in SZ and BP disorder
with psychosis.
Five-year view
For over a decade, VPA has been associated with atypical anti-
psychotics in the treatment of the cognitive deficits and anxiety,
which are two symptoms commonly expressed by SZ and BP
disorder patients [56–58]. It has been suggested that when VPA is
used in combination with atypical antipsychotics, this cotreat-
ment is not only well tolerated, but in fact leads to an earlier and
more pronounced improvement of psychotic symptoms com-
pared with that following antipsychotic monotherapy [56–58].
The data presented here suggest that the coadministration of
VPA and atypical antipsychotics may correct the altered nuclear
epigenetic function operative in selected populations of cortical
GABAergic neurons of SZ and BP disorder patients. In fact,
both VPA and CLZ, by covalently modifying histone tail lysines
(via acetylation or methylation), activate DNA demethylation
and increase GAD67 mRNA and protein expression in the frontal
cortex of rodents [9,34–36]. To prove the concept that 'histone
code' remodeling is at the base of VPA augmenting antipsychotic
action, a reasonable working hypothesis is to associate antipsy-
chotics with other more-potent, more-specific and chemically
unrelated HDAC inhibitors [33].
Although HDAC inhibitors, such as benzamide MS-275, elicit
brain histone code modifications and activate DNA demethyla-
tion in a manner similar to that caused by VPA [33,34], there are
no clinical studies as yet reporting on the action of MS-275
administered alone or in association with anti psychotics in the
treatment of psychosis. However, we can predict that, in the
future, increasing our knowledge of the cellular brain distri-
bution of various HDAC subtypes or of DNA demethylases
will be possible and would help to develop specific nontoxic
epi genetically active drugs to target neuronal populations dys-
functional in SZ or BP disorder brains. However, as a general
rule, we must avoid the use of drugs that produce a gener-
alized and indiscriminate increase of DNA demethylation,
because such an activation may induce serious neuropathologies
(i.e., loss of myelin stability in the brains of multiple sclerosis
patients [60]).
Our working hypothesis is that the epigenetic processes opera-
tive in the pathophysiology of SZ and BP disorder may include a
dysfunction of DNA demethylase activity presumably expressed
in brain GABAergic neurons of these patients. However, the his-
tochemical localization and the biochemical properties of such
an enzyme require further investigation. Moreover, the induc-
tion of a DNA demethylase by a specific class of antipsychot-
ics may mediate the promoter hypomethylation of vulnerable
GABAergic genes that are downregulated in psychosis by pro-
moter hypermethylation. A drug that upregulates DNA demeth-
ylation appears of interest in order to relieve the characteristic
GABAergic pathologies operative in SZ and BP disorder.
Since there is strong evidence for a downregulation of reelin
and GAD67 expression mediated by the hypermethylation of CpG
islands embedded in the promoter regions of SZ patients, the pos-
sibility that a new generation of antipsychotics that may activate
DNA demethylase and thereby overcome the downregulation
of reelin and GAD67 expression will open new horizons to reach
important pharmacological targets (defined in this review) that
should be addressed in the treatment of the symptoms associated
with SZ and BP disorders.
Page 10
Expert Rev. Neurother. 9(1), (2009)
Costa, Chen, Dong et al.
Financial & competing interests disclosure
A Guidotti has funding support (NIMH4RO1MH70855). E Costa has
funding support (NIMH2RO1MH71667). The authors have no other
relevant affiliations or financial involvement with any organization or
entity with a financial interest in or financial conflict with the subject
matter or materials discussed in the manuscript apart from those
No writing assistance was utilized in the production of this manuscript.
Key issues
A deficit of GABAergic neurotransmission emerges as the pathophysiological core mechanism operative in schizophrenia (SZ) and
bipolar (BP) disorders.
In telencephalic GABAergic neurons of SZ and BP disorder patients, the expression of glutamic acid decarboxylase (GAD
that synthesizes the inhibitory neurotransmitter GABA, and reelin, an extracellular matrix protein with a trophic synaptic function, are
consistently downregulated.
The downregulation of GAD
of DNA methyltransferase 1 (DNMT1).
An approach to correct the defective GABAergic transmission of psychotic patients should be to administer drugs acting as positive
allosteric modulators of GABA action at selective GABAA receptor subtypes. Here, we propose that a benzodiazepine such as imidazenil
– which fails to induce sedation because it is devoid of action on α1-containing GABAA receptor subtypes – might be an ideal
compound to upregulate GABAergic transmission deficits in psychotic patients.
A pharmacological strategy to reverse the GAD
acting on chromatin remodeling.
We have established that clozapine, when coadministered with valproate (a histone deacetylase inhibitor), facilitates chromatin
remodeling and induces reelin and GAD67 promoter demethylation in a manner that may lead to normalization of the downregulation
of GABAergic gene expression in SZ and BP patients.
67), the enzyme
67 and reelin is most likely caused by their promoter hypermethylation catalyzed by an increased expression
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Erminio Costa, MD •?
Professor and Director, Psychiatric
Institute, Department of Psychiatry,
University of Illinois at Chicago,
1601 Taylor, Chicago, IL 60612, USA
Tel.: +1 312 413 4591
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Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
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Department of Psychiatry, University of
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Dennis R Grayson, PhD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
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Marija Kundakovic, PhD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
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Department of Psychiatry, University of
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William (Brad) Ruzicka, PhD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
Tel.: +1 312 413 4591
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Rosalba Satta, PhD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
Tel.: +1 312 413 4591
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Marin Veldic, MD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
Tel.: +1 312 413 4591
Fax: +1 312 413 4569
Adrian Zhubi, MD
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
Tel.: +1 312 413 4591
Fax: +1 312 413 4569
Alessandro Guidotti, MD
Scientific Director, Psychiatric Institute,
Department of Psychiatry, University of
Illinois at Chicago, 1601 Taylor, Chicago,
Tel.: +1 312 413 4594
Fax: +1 312 413 4569


text:
Day 4: NY Fashion Week
Blake Lively
Gossip Girl Blake Lively took in Diane von Furstenberg's bright, bohemian collection wearing a sparkling sequin jacket and skinny jeans. Fellow fashionistas Mariska Hargitay and Michelle Trachtenberg also joined the actress in the front row.
-with reporting by Joyann King and Lauren Tumas
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From setting a date to compiling the guest list, read up on engagement party planning basics.
Your top to-do after saying "yes" is to soak up that once-in-a-lifetime just-engaged glow, so put these 10 nonurgent details on the back burner for now.
No matter how long you and your fiancé dated or how many times you've been a bridesmaid in your friends' weddings, there are some truths you just don't find out about until you get engaged.
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-- Anja Winikka


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Well, The infection turned out to be staph(sp?), and particularly nasty. It's one of those antibiotic-resistant strains you hear about on the news. There's only one effective medicine for it at her age, and she has to have it via IV every six hours. So, she has to stay in the hospital. At least 2 more days, and maybe up to 10 days, the doc said.
On a positive note, she's back to her old self. The downside to this is,she's going stir crazy. She's been walking well for about 2 months now, and running for the last month. Now she's stuck in a 10x14 room. We've been walking her up and down the corridors, but it doesn't cut it. The staff has also been having trouble keeping her IVs in, since she's so active.
I'm really worried that the time in the hospital is going to slow her development. There just isn't a way to get her the level of mental stimulation she needs while she's in there.


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ROUSES POINT - Fourth of July weekend events have been taking place in Rouses Point for the last five decades. This year, however, a special ceremony will take place that has even warranted sending an invitation to President Barack Obama.
On Friday, July 1, a special flag measuring 30 feet by 57.5 feet will be brought to the Rouses Point Civic Center on Lake Street. The flag has been traveling the country since Aug. 10, 2010, beginning in San Diego, Calif., as a tribute to the Armed Forces, first and second responders, those who died during the Sept. 11, 2001 attacks on the United States and their families.
Geri Favreau, one of the organizers of the annual Fourth of July events, explained Rouses Point is one of only five stops that will be made with the flag in New York State. The village was added to the itinerary after Chazy native Tom Middleton, coordinator for the Vermont stops, asked Favreau if they'd be interested in having it brought there.
"I think it's very special to have this flag coming to Rouses Point. If it weren't for Tom Middleton, we wouldn't have it," Favreau said.
Favreau said the original plan was to have the flag hang off the Korean War Veterans Memorial Bridge between Rouses Point and Alburgh, Vt. However, concerns of wind blowing the flag off the bridge changed plans to be hang the flag from aerial trucks at the civic center.
The ceremony will be organized by members of American Legion Montgomery Post 912. Favreau explained the Legion Commander Robert St. John sent her a note explaining the ceremony will be done "to honor the memory of the civilians, fire fighters and police officers who died Sept. 11, 2001, and to honor the memory of the military personnel who have and continue to die in the global war on terror."
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Microsoft's current desktop OS is close to stealing second place from Windows XP in the desktop OS arena, according to new stats from Web tracker Net Applications.
Though Microsoft no longer supports the aged OS, a fair share of businesses haven't yet kicked the XP habit.
Businesses are finally getting around to upgrading the decade-plus-old XP operating system. This is boosting PC demand in the US and other mature markets, says Citi.
Microsoft no longer supports XP users, but a host of antivirus products tested by AV-Test can still defend you from viruses.
Though no longer supported by Microsoft, XP commanded a quarter of all desktop OS traffic seen by Net Applications last month.
Stronger demand for business PCs has delivered better-than-expected second-quarter sales for Intel.
The hack seemingly allows XP users to continue to install security updates, but Microsoft is advising people not to use it.
XP still accounted for 26 percent of all desktop OS traffic tracked last month by Net Applications.
The browser bug was so severe the US and UK issued warnings. Surprisingly, Microsoft's fix brings an update to its outmoded XP software.
From the good-sport department, Microsoft takes a shot at itself with a Web game that encourages players to send Windows XP back to the early naughts, permanently.


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Jack Thompson Weighs in on Oblivion 505
from the zonk-smash dept.
from the zonk-smash dept.
Robotron23 writes "Jack Thompson has commenced his attack on The Elder Scrolls IV: Oblivion by seeking appearances on several talk shows. A press release announcing his availability speaks of Take Two not having 'learned its lesson' over the Hot Coffee scandal, before continuing to detail the issues surrounding Oblivion's re-rating, in particular regarding nudity - concluding that the game content will spawn 'an even worse disaster' than occurred during Hot Coffee."


text:
Spring cleaning for the mind
Anti-U.S. socialism in Latin American led by Venezuela's president
Hugo Chavez, president of Venezuela, plans to expand his philosophy in Latin America and his rule in Venezuela.
Tuesday's voters called the school system's bluff
The county clerk threw us a curve ball
With the election results coming in sooner than expected, the Missourian staff was juggling three options for delivering the news to you.
Traumatic experiences should be left in the past
Which airline do you want to see move to Columbia?
Three airlines were proposed for Columbia Regional Airport. Hawaii Island Air was eliminated because the flight times were unacceptable. Great Lakes and Mesaba have their advantages and disadvantages.
McCain shows disregard for law, U.S. Constitution
Sen. John McCain is unfit to be president of the United States because he has confused moral values.
Making Missouri homes energy independent is good for the environment
The state of Missouri should partner with homeowners to make homes energy efficient. Although energy independent additions to home are expensive, they save money over time.
Technology doesn't save time; it just breeds dependence
Computers constantly break down, and people waste a lot of time trying to fix the problems. Most young people rely too heavily on technology.
Election looks like it will bring change to Zimbabwe
The apparent defeat of Robert Mugabe has brought cautious optimism.
Trust Columbia Public School administrators and support tax levy
Although the school board members are not their best advocates, they are dediated and hard-working. We need to elect more members that care and trust their judgment regarding the tax levy.
Letter: Accomplishments make Sturtz ideal candidate
Paul Sturtz's commitment makes him the ideal candidate to represent the First Ward.
Letter: Approving tax levy will help decrease ignorance and want
Supporting public education will help us deal with current, future problems.
Letter: Sturtz will meet our expectations of a City Council member
Paul Sturtz has the creativity and knowledge of government needed for Columbia.
Letter: Columbia Public School tax levy is all about the kids
Vote yes for the proposed tax levy on April 8.
Letter: Sturtz has shown he has vision
Through his advancement of the arts in Columbia, Paul Sturtz has shown he would make a good City Council member.
Letter: Tax levy will keep Columbia Public Schools competitive
The Columbia Public School District is asking voters to approve a 54-cent tax levy. Based on an assessment by financial consultants, this increase, as well as an internal budget reduction of $5 million, will result in financial stability for the next five years.
We built this city on sewers
Even sewage systems factor into decisions made on Election Day.
Baby falls asleep but doesn't stay asleep
Audrianna is just one month old, and she has been keeping her parents awake at night.
Discriminatory language still unbelievably prevalent
Hateful language flows too often and too easily, and sometimes comes from unlikely sources ... and it needs to be stopped.


text:
Built with a high-strength cast aluminum structure and a polycarbonate globe, the fixture, the winning result of a Central Park Conservancy design competition, utilizes a range of sources including HID, compact fluorescent, and induction. Measuring 18 inches in diameter, several decorative bases are available for coordination with ornamental posts.
Three aimable optical systems contained in a single in-ground luminaire, the fixture uses either LED, HID, or linear fluorescent lamps for evenly distributed spot, flood, or accent lighting. It can be installed in both landscape and poured concrete applications and contains parabolic reflectors that can rotate up to 360 degrees and tilt up to 15 degrees for precise aiming.
Industrial is an exterior-rated LED fixture that contains a 1W LED in each lamp head. Available as a dual-axis, adjustable, single-head, remotely fed fixture, a four-head aluminum integral wall sconce, or a four-head aluminum bollard, the line comes with LEDs in cool-white or warm-white color temperatures.
The Treviso LED line recreates classic, late 1800s outdoor area and site luminaires, poles, and bases with LED light source options. The collection features large, ribbed domes with top-mounted finials and three decorative supports. Each luminaire uses just 18W of power and can be specified in cast aluminum or steel.
A classic pillar silhouette housing an energy-saving LED source, Illume measures 3½ inches in diameter and is offered in lengths of 12, 24, and 36 inches for base mount and 12, 24, 36, 48, and 60 inches for direct burial. LEDs are available in cool- and warm-white light, as well as color, and each fixture uses only 2.5W per foot.
A series of LED-powered light columns in four heights (8, 10, 13, and 21 feet) with a companion bollard, the Sierra collection provides softly diffused colored illumination for accent, brand identification, or zonal definition. Both the column (shown) and bollard designs embody contemporary aesthetics, are made of extruded aluminum tubing, and come with LEDs in a choice of RGB, white, blue, green, red, or royal blue. The columns also are available in building- and wall-mounted versions.
One in a family of Eco Series fixtures, the Twin Pole Lite metal halide MR16 20W is a durable, high-efficiency outdoor luminaire that offers the latest low-energy lamp technology. A pole-mounted version of the company's Spike Spot Adjustable Metal Halide, it is easily inserted into the ground and has fully adjustable heads allowing 360-degree rotation and 0- to 90-degree elevation adjustment.
A line of stainless-steel LED walkover luminaires, Steel is offered in square and round profiles in single or double configurations. It is IP67 rated for temporary submersion in water and installs flush or with a decorative and functional Visa trim, which directs light outward in single or double patterns.
The Exterior 200 LED is a long-life programmable luminaire that offers full RGB plus amber and white color mixing. Weatherproofed for the illumination of outdoor architecture, it features a comprehensive color range from 44 high-power LEDs and a choice of optional spread angles. The lenses also are available as accessories, allowing the luminaire to be reconfigured on site if necessary.
A contemporary-styled, exterior-grade wall sconce, the mini eSconce was developed to covert an unwelcome but necessary object on a building's surface into an integral design element. Now available with LEDs in white, blue, green, red, and amber, the fixture consumes only 5W of power and is offered in numerous colors with optional fascia panels and colored lenses.
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Mike Casey's Blog
Sleep Number
by Mountain Geeks
posted Jan 17 2013 1:29PM
As you may or not know, Mike Casey has been rocking a Sleep Number bed and now he's checking in with Sleep Expert, Pete Bils. Find out what tips and tricks Pete Bils has to offer. Listen to the interview below.
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Sleep Number Blog
Wow, a whole radio interview (with no volume control) all about the sleep nnumber bed. Why is this a titled as a blog and not just another annoying commercial? Way to sell out. Hope they paid you well.
what is your # number
hi my name is ariana and I am 9 years old love your music by the way my name on facebook is love ariana lee rickey ,what is your number.
Do you love it?
I'm in the market for a new bed and am thinking about this. What's your final verdict?
Best bed= Sleep number
The interview was really useful and I think that sleep number bed is comfortable and gives me the best sleep I've had in years. airbedpros.com -Jannette F. Labrecque
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Twitter helps smokers get rid of the habit
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January 14, 2015
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Political will deficit could undermine Sustainable Development Goals
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Health Ministry proposes fine of Rs 1000 on smoking in public places, ban on sale of loose cigarettes
January 14, 2015
New Delhi: In a bid to curb tobacco consumption, the Health Ministry has now formally proposed changes in anti-smoking law with a ban on sales of loose or single sticks of cigarettes. The ministry sought to raise the fine on smoking in public places from Rs 200 to Rs 1000...
Declare polling stations in J-K as smoking free zones: JKVHA
December 3, 2014
Srinagar: Jammu and Kashmir Voluntary Health Association (JKVHA) has asked the state election authorities to declare all polling stations as 'smoking free zones' during the ongoing Assembly elections. "JKVHA in a letter to the Chief Election Officer (CEO), has demanded that keeping in view the general health of the people, the...
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October 18, 2014
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Smoking in public likely to earn you a fine of Rs 20,000
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West Bengal: Smoke in hospital, 22 patients go missing
August 31, 2014
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Is putting age limit for smoking good idea to curb tobacco use?
August 21, 2014
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Watch: It took a 13yearold to shame these Indian smokers to admitting they want to quit smoking
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A worrying new study reveals that smoking in front of children not only passes on the harmful effects of the smoke in the air it can also get them hooked on cigarettes. Most smokers are very aware of the damage they are doing to their own health and...
Chinese pet dog is a smoking addict
July 1, 2014
Beijing: A twoyearold dog owned by a smoker in China has picked up his master's habit and become so addicted to smoking that it must have a cigarette before going to sleep. The husky in Anda, Heilongjiang province began smoking last year, when it was adopted, according to its owner,...
TWTW: Cyrus Broacha discusses the 'advantages' of cigarette smoking
June 22, 2014
While Health Minister Harsh Vardhan asked Finance Minister Arun Jaitley, to increase the rates of cigarettes, Cyrus Broacha in this episode of 'The Week That Wasn't' discusses the 'advantages' of smoking. ...
Prisons made smokingfree zones in Kerala
June 21, 2014
Thiruvananthapuram: Prisons in Kerala have been made smokingfree zones in compliance of the Supreme Court order banning smoking in public places. A circular in this regard has been issued by DGP (Prison) T P Senkumar, directing jail officials to enforce the ban within the prison compounds. "Though smoking in public...
US teens are drinking less, texting more: Study
June 15, 2014
New York: American teens are smoking less, drinking less and fighting less. But they're texting behind the wheel and spending a lot of time on video games and computers, according to the government's latest study of worrisome behaviour. Generally speaking, the news is good. Most forms of drug use, weapons use...
Kolkata records highest cigarette consumption in India: Survey
June 12, 2014
Kolkata: With a person in Kolkata on an average smoking 910 cigarettes per day, the city has recorded the highest cigarette consumption in India, a survey report released on Thursday said. It said that the frequency of tobacco consumption is the highest in Kolkata among the cities. A Mumbaikar, for example,...
Complaint against Shakti Kapoor for smoking in public
June 4, 2014
A complaint has been filed with a local court against Bollywood actor Shakti Kapoor for allegedly smoking publicly at Sanganer airport in Jaipur. The complainant Nem Singh, a lawyer, has sought action against the actor and the airport authority under Section 5/11 of Rajasthan Prevention of Smoking Act. The complaint was...
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text:
A comparison of two methods for assessing drug sensitivity in Giardia duodenalis
Pearce, D.A., Reynoldson, J.A. and Thompson, R.C. (1996) A comparison of two methods for assessing drug sensitivity in Giardia duodenalis. Applied parasitology, 37 (2). pp. 111-116.
This study compared two published methods for assessing the in-vitro drug sensitivity of Giardia duodenalis: the 3H-thymidine incorporation assay (BOREHAM et al. 1984) which radiometrically measures nucleic acid synthesis; and inhibition of adherence (MELONI et al. 1990). Giardia trophozoites were exposed to a range of concentrations of metronidazole or albendazole for 24 hours and their viability measured by both assays in order to determine the degree of correlation between the two methods of measuring viability. Due to the different modes of action of metronidazole and albendazole on Giardia, measuring the inhibition of adherence appears to be a more accurate indicator of trophozoite viability than measuring 3H-thymidine incorporation.
Publication Type: | Journal Article |
Murdoch Affiliation: | School of Veterinary Studies |
Publisher: | Gustav Fischer Verlag |
URI: | http://researchrepository.murdoch.edu.au/id/eprint/10664 |
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text: Ramadan is a month of blessings and prayers. In this month, Muslims from all around the world fast in the daytime. All kinds of food and drinks are forbidden from dawn to dusk, or daytime. It is a month of kindness which ends with one of the biggest festivals in … Continue reading »