Rosalba Satta

DNA-methyltransferase 1 mRNA is selectively overexpressed in telencephalic GABAergic interneurons of schizophrenia brains

A down-regulation of reelin and glutamic acid decarboxylase (GAD) 67 mRNAs was detected in γ-aminobutyric acid (GABA)ergic cortical interneurons of schizophrenia (SZ) postmortem brains (10), suggesting that the availability of GABA and reelin may be decreased in SZ cortex. In situ hybridization of the mRNA encoding for DNA-methyltransferase 1, which catalyzes the methylation of promoter CpG islands, shows that the expression of this mRNA is increased in cortical GABAergic interneurons but not in pyramidal neurons of SZ brains. Counts of reelin mRNA-positive neurons in Brodmanns area 10 of either nonpsychiatric subjects or SZ patients show that the expression of reelin mRNA is decreased in layer-I, -II, and -IV GABAergic interneurons of SZ patients. These findings are consistent with the hypothesis that the increase of DNA-methyltransferase 1 expression in telencephalic GABAergic interneurons of SZ patients causes a promoter hypermethylation of reelin and GAD67 and perhaps of other genes expressed in these interneurons. It is difficult to decide whether this dysfunction of GABAergic neurons detected in SZ is responsible for this disease or is a consequence of this disorder. Although at present we cannot differentiate between these two alternatives, it is important to consider that so far a molecular pathology of cortical GABAergic neurons appears to be the most consistent finding associated with SZ morbidity.

Nicotine decreases DNA methyltransferase 1 expression and glutamic acid decarboxylase 67 promoter methylation in GABAergic interneurons.

Tobacco smoking is frequently abused by schizophrenia patients (SZP). The major synaptically active component inhaled from cigarettes is nicotine, hence the smoking habit of SZP may represent an attempt to use nicotine self-medication to correct (i) a central nervous system nicotinic acetylcholine receptor (nAChR) dysfunction, (ii) DNA-methyltransferase 1 (DMT1) overexpression in GABAergic neurons, and (iii) the down-regulation of reelin and GAD67 expression caused by the increase of DNMT1-mediated hypermethylation of promoters in GABAergic interneurons of the telencephalon. Nicotine (4.5–22 μmol/kg s.c., 4 injections during the 12-h light cycle for 4 days) decreases DNMT1 mRNA and protein and increases GAD67 expression in the mouse frontal cortex (FC). This nicotine-induced decrease of DNMT1 mRNA expression is greater (80%) in laser microdissected FC layer I GABAergic neurons than in the whole FC (40%), suggesting selectivity differences for the specific nicotinic receptor populations expressed in GABAergic neurons of different cortical layers. The down-regulation of DNMT1 expression induced by nicotine in the FC is also observed in the hippocampus but not in striatal GABAergic neurons. Furthermore, these data show that in the FC, the same doses of nicotine that decrease DNMT1 expression also (i) diminished the level of cytosine-5-methylation in the GAD67 promoter and (ii) prevented the methionine-induced hypermethylation of the same promoter. Pretreatment with mecamylamine (6 μmol/kg s.c.), an nAChR blocker that penetrates the blood–brain barrier, prevents the nicotine-induced decrease of FC DNMT1 expression. Taken together, these results suggest that nicotine, by activating nAChRs located on cortical or hippocampal GABAergic interneurons, can up-regulate GAD67 expression via an epigenetic mechanism. Nicotine is not effective in striatal medium spiny GABAergic neurons that primarily express muscarinic receptors.

Characterization of the action of antipsychotic subtypes on valproate-induced chromatin remodeling

Recent advances in schizophrenia (SZ) research indicate that the telencephalic gamma-aminobutyric acid (GABA)ergic neurotransmission deficit associated with this psychiatric disorder probably is mediated by the hypermethylation of the glutamic acid decarboxylase 67 (GAD(67)), reelin and other GABAergic promoters. A pharmacological strategy to reduce the hypermethylation of GABAergic promoters is to induce a DNA-cytosine demethylation by altering the chromatin remodeling with valproate (VPA). When co-administered with VPA, the clinical efficacy of atypical antipsychotics is enhanced. This prompted us to investigate whether this increase in drug efficacy is related to a modification of GABAergic-promoter methylation via chromatin remodeling. Our previous and present results strongly indicate that VPA facilitates chromatin remodeling when it is associated with clozapine or sulpiride but not with haloperidol or olanzapine. This remodeling might contribute to reelin- and GAD(67)-promoter demethylation and might reverse the GABAergic-gene-expression downregulation associated with SZ morbidity.

GABAergic promoter hypermethylation as a model to study the neurochemistry of schizophrenia vulnerability.

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.

Selective α4β2 nicotinic acetylcholine receptor agonists target epigenetic mechanisms in cortical GABAergic neurons.

Nicotine improves cognitive performance and attention in both experimental animals and in human subjects, including patients affected by neuropsychiatric disorders. However, the specific molecular mechanisms underlying nicotine-induced behavioral changes remain unclear. We have recently shown in mice that repeated injections of nicotine, which achieve plasma concentrations comparable to those reported in high cigarette smokers, result in an epigenetically induced increase of glutamic acid decarboxylase 67 (GAD67) expression. Here we explored the impact of synthetic α4β2 and α7 nAChR agonists on GABAergic epigenetic parameters. Varenicline (VAR), a high-affinity partial agonist at α4β2 and a lower affinity full agonist at α7 neuronal nAChR, injected in doses of 1–5 mg/kg/s.c. twice daily for 5 days, elicited a 30–40% decrease of cortical DNA methyltransferase (DNMT)1 mRNA and an increased expression of GAD67 mRNA and protein. This upregulation of GAD67 was abolished by the nAChR antagonist mecamylamine. Furthermore, the level of MeCP2 binding to GAD67 promoters was significantly reduced following VAR administration. This effect was abolished when VAR was administered with mecamylamine. Similar effects on cortical DNMT1 and GAD67 expression were obtained after administration of A–85380, an agonist that binds to α4β2 but has negligible affinity for α3β4 or α7 subtypes containing nAChR. In contrast, PNU–282987, an agonist of the homomeric α7 nAChR, failed to decrease cortical DNMT1 mRNA or to induce GAD67 expression. The present study suggests that the α4β2 nAChR agonists may be better suited to control the epigenetic alterations of GABAergic neurons in schizophrenia than the α7 nAChR agonists.

Drosophila metabolize 1,4-butanediol into γ-hydroxybutyric acid in vivo

Abstract A solvent, 1,4-butanediol, is also abused as a recreational drug. In mammals, 1,4-butanediol is metabolized into γ-hydroxybutyric acid (GHB), which stimulates metabotropic γ-aminobutyric acid (GABA) GABA B and putative GHB receptors. Here we show that in vivo injection of 1,4-butanediol into adult Drosophila leads to GHB synthesis (GHB was detectable 5 min after 1,4-butanediol injection and increased dramatically 1–2 h later). This synthesis of GHB was accompanied by an impairment of locomotor activity that was mimicked by a direct injection of GHB into flies. We propose Drosophila as a model to study the molecular actions of 1,4-butanediol and GHB.

Estradiol increases the sensitivity of ventral tegmental area dopamine neurons to dopamine and ethanol

Gender differences in psychiatric disorders such as addiction may be modulated by the steroid hormone estrogen. For instance, 17β-estradiol (E2), the predominant form of circulating estrogen in pre-menopausal females, increases ethanol consumption, suggesting that E2 may affect the rewarding properties of ethanol and thus the development of alcohol use disorder in females. The ventral tegmental area (VTA) is critically involved in the rewarding and reinforcing effects of ethanol. In order to determine the role of E2 in VTA physiology, gonadally intact female mice were sacrificed during diestrus II (high E2) or estrus (low E2) for electrophysiology recordings. We measured the excitation by ethanol and inhibition by dopamine (DA) of VTA DA neurons and found that both excitation by ethanol and inhibition by dopamine were greater in diestrus II compared with estrus. Treatment of VTA slices from mice in diestrus II with an estrogen receptor antagonist (ICI 182,780) reduced ethanol-stimulated neuronal firing, but had no effect on ethanol-stimulated firing of neurons in slices from mice in estrus. Surprisingly, ICI 182,780 did not affect the inhibition by DA, indicating different mechanisms of action of estrogen receptors in altering ethanol and DA responses. We also examined the responses of VTA DA neurons to ethanol and DA in ovariectomized mice treated with E2 and found that E2 treatment enhanced the responses to ethanol and DA in a manner similar to what we observed in mice in diestrus II. Our data indicate that E2 modulates VTA neuron physiology, which may contribute to both the enhanced reinforcing and rewarding effects of alcohol and the development of other psychiatric disorders in females that involve alterations in DA neurotransmission.

Estrogen Receptor β in the Nucleus Accumbens Regulates the Rewarding Properties of Cocaine in Female Mice

Abstract Background Females are more vulnerable to developing cocaine addiction compared with males, a phenomenon that may be regulated by the steroid hormone 17β-estradiol. 17β-Estradiol enhances cocaine reward as measured by the conditioned place preference test. It is currently not known which estrogen receptor is involved or the neuroanatomical locations in which estrogen receptors act to enhance cocaine responses. The purpose of this study was to determine if the estrogen receptors ERα and ERβ regulate cocaine conditioned place preference in mice and whether they act in the nucleus accumbens, a brain region critically involved in the development of cocaine abuse. Methods Ovariectomized mice were treated with 17β-estradiol or agonists selective for ERα or ERβ and tested for cocaine conditioned place preference and for c-fos expression in the nucleus accumbens. Female mice with intact ovaries were also tested for cocaine conditioned place preference after RNA interference-mediated knockdown of ERα or ERβ in the nucleus accumbens. Results We found that mice treated with 17β-estradiol or an ERβ agonist exhibited increased cocaine conditioned place preference, while knockdown of ERβ, but not ERα, in the nucleus accumbens of females with intact ovaries abrogated cocaine conditioned place preference. Acute treatment with 17β-estradiol or an ERβ agonist induced expression of the immediate-early gene c-fos in the nucleus accumbens, whereas the ERα agonist did not. Conclusions These data indicate that ERβ in the nucleus accumbens regulates the development of cocaine conditioned place preference in female mice. 17β-Estradiol may activate neurons in the nucleus accumbens via ERβ. We speculate that this might increase the saliency of cocaine cues that predict drug reward.