Glutamatergic and GABAergic Metabolism in Mouse Brain under Chronic Nicotine Exposure: Implications for Addiction

BACKGROUND AND PURPOSE: The effects of nicotine on cerebral metabolism and its influence on smoking behavior is poorly understood. An understanding of the chronic effects of nicotine on excitatory and inhibitory metabolic demand, and corresponding neurotransmission may provide clues for designing strategies for the optimal smoking cessation intervention. The objective of the current study was to investigate neuronal and astroglial metabolism in mice exposed to nicotine (0.5 and 2.0 mg/kg, sc) three times in a day for 4 weeks. EXPERIMENTAL APPROACH/PRINCIPAL FINDINGS: Metabolic measurements were carried out by co-infusing [U-(13)C(6)]glucose and [2-(13)C]acetate, and monitoring (13)C labeling of amino acids in brain tissue extract using (1)H-[(13)C] and (13)C-[(1)H]-NMR spectroscopy. Concentration of (13)C-labeled glutamate-C4 was increased significantly from glucose and acetate with chronic nicotine treatment indicating an increase in glucose oxidation by glutamatergic neurons in all brain regions and glutamate-glutamine neurotransmitter cycle in cortical and subcortical regions. However, chronic nicotine treatment led to increased labeling of GABA-C2 from glucose only in the cortical region. Further, increased labeling of glutamine-C4 from [2-(13)C]acetate is suggestive of increased astroglial activity in subcortical and cerebellum regions of brain with chronic nicotine treatment. CONCLUSIONS AND SIGNIFICANCE: Chronic nicotine exposure enhanced excitatory activity in the majority of brain regions while inhibitory and astroglial functions were enhanced only in selected brain regions.     

Genotypic Association of the DAOA Gene with Resting-State Brain Activity in Major Depression

Compelling evidence suggests that the glutamatergic system may contribute to the pathophysiology of major depression (MDD). While the D-amino acid oxidase activator (DAOA) gene can affect glutamatergic function, its genetic associations with MDD and abnormal resting-state brain activity have yet to be elucidated. A total of 488 patients with MDD and 480 controls were recruited to examine MDD association for the DAOA gene in a Chinese population, of whom 53 medication-free patients and 46 well-matched controls underwent resting-state functional magnetic resonance imaging for regional homogeneity (ReHo) analysis. The differences in ReHo between genotypes of interest were initially tested by the Student??s t test, and the 2?×?2 (genotypes?×?disease status) ANOVA was then performed to identify the main effects of genotypes, disease status, and their interactions in MDD. Allelic association of the DAOA gene with MDD was observed for rs2391191, rs3918341, and rs778294 and haplotypic association for 2- and 3-SNP haplotypes. Six clusters in the cerebellum, right middle frontal gyrus and left middle temporal gyrus showed genotypic association between altered ReHo and rs2391191. The main effects of rs2391191 genotypes were found in the right culmen and right middle frontal gyrus. The left uvula and left middle temporal gyrus showed a genotypes?×?disease status interaction. Our results suggest that the DAOA gene may confer genetic risk of MDD. Genotypic effect of rs2391191 and its interaction with disease status may contribute to the altered ReHo in patients with MDD. Glutamatergic modulation may play an important role in alteration of the resting-state brain activities.     

Statistical Analysis of Multiplex Brain Gene Expression Images

Analysis of variance (ANOVA) was employed to investigate 9,000 gene expression patterns from brains of both normal mice and mice with a pharmacological model of Parkinson's disease (PD). The data set was obtained using voxelation, a method that allows high-throughput acquisition of 3D gene expression patterns through analysis of spatially registered voxels (cubes). This method produces multiple volumetric maps of gene expression analogous to the images reconstructed in biomedical imaging systems. The ANOVA model was compared to the results from singular value decomposition (SVD) by using the first 42 singular vectors of the data matrix, a number equal to the rank of the ANOVA model. The ANOVA was also compared to the results from non-parametric statistics. Lastly, images were obtained for a subset of genes that emerged from the ANOVA as significant. The results suggest that ANOVA will be a valuable framework for insights into the large number of gene expression patterns obtained from voxelation.     

Glutamatergic and GABAergic inputs to the RVL mediate cardiovascular adjustments to noxious stimulation

Stimulation of cutaneous and muscle afferents induces several cardiovascular adjustments such as hypertension, tachycardia, and muscle vasodilation. Although previous studies have demonstrated that the rostral ventrolateral medulla (RVL) mediates sympathoexcitation and pressor responses to sciatic nerve stimulation (SNS), whether it also mediates blood flow adjustments remains unclear. Therefore, in the present study, we examined the role of the RVL in the vasodilation induced by SNS and the possible neurotransmitters involved. In Urethane-anesthetized, paralyzed, and artificially ventilated rats, SNS (square pulses, 1 ms, 20 Hz, 800--1200 microA, 10 s) produced increases in blood pressure, heart rate, blood flow, and vascular conductance of the stimulated limb. Unilateral microinjection of kainic acid (2 nmol/100 nl) into the RVL contralateral to the stimulated limb abolished cardiovascular adjustments to SNS. Unilateral microinjections of kynurenic acid (2 nmol/100 nl) selectively abolished the pressor response to SNS, whereas bicuculline (400 pmol/100 nl) abolished the increases in blood flow without changing the pressor response. These results suggest that glutamatergic synapses within the RVL mediate pressor responses, whereas GABAergic synapses may mediate the vasodilation to SNS.     

A General Mechanism for Viral Resistance to Suicide Gene Expression

Bacteriophage T7 was challenged with either of two toxic genes expressed from plasmids. Each plasmid contained a different gene downstream of a T7 promoter; cells harboring each plasmid caused an infection by wild-type T7 to abort. T7 evolved resistance to both inhibitors by avoidance of the plasmid expression system rather than by blocking or bypassing the effects of the specific toxic gene product. Resistance was due to a combination of mutations in the T7 RNA polymerase and other genes expressed at the same time as the polymerase. Mutations mapped to sites that are unlikely to alter polymerase specificity for its cognate promoter but the basis for discrimination between phage and plasmid promoters in vivo was not resolved. A reporter assay indicated that, relative to wild-type phage, gene expression from the plasmid was diminished several-fold in cells infected by the evolved phages. A recombinant phage, derived from the original mutant but lacking a mutation in the gene for RNA polymerase, exhibited intermediate activity in the reporter assay and intermediate resistance to the toxic gene cassettes. Alterations in both RNA polymerase and a second gene are thus responsible for resistance. These findings have broad evolutionary parallels to other systems in which viral inhibition is activated by viral regulatory signals such as defective-interfering particles, and they may have mechanistic parallels to the general phenomena of position effects and gene silencing. Received: 18 July 2000 / Accepted: 8 February 2001     

Subregional Expression of Hippocampal Glutamatergic and GABAergic Genes in F344 Rats with Social Isolation after Weaning

Many studies have shown that postweaning social isolation (pwSI) alters various behavioral phenotypes, including hippocampus-dependent tasks. Here, we report the comprehensive analysis of the expression of glutamatergic and GABAergic neurotransmission-related genes in the distinct hippocampal subregions of pwSI rats. Male F344 rats (age, 4 wk) experienced either pwSI or group housing (controls). At 7 wk of age, the hippocampus of each rat was removed and laser-microdissected into the CA1 and CA3 layers of pyramidal cells and the granule cell layer of the dentate gyrus. Subsequently, the expression of glutamatergic- and GABAergic-related genes was analyzed by quantitative RT-PCR. In the CA1 and CA3 pyramidal cell layers, 18 of 24 glutamate receptor subunit genes were at least 1.5-fold increased in expression after pwSI. In particular, the expression of several N-methyl-D-aspartate and kainate receptors (for example, Grin2a in CA1, Grik4 in CA3) was significantly increased after pwSI. In contrast, pwSI tended to decrease the expression of GABA_A receptor subunit genes, and Gabra1, Gabra2, Gabra4, Gabra5, Gabrb2, Gabrg1, and Gabrg2 were all significantly decreased in expression compared with the levels in the group-housed rats. These results indicate a subregion-specific increase of glutamate receptors and reduction of GABA_A receptors, suggesting that the hippocampal circuits of pwSI rats may be in more excitable states than those of group-housed rats.Abbreviations: AMPA, α-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid, LMD, laser microdissection, NMDA;N-methyl-D-aspartic acid, pwSI, postweaning social isolation, DG;dentate gyrusBehavioral phenotypes differ among strains of laboratory animals, including mice and rats. The acquisition of strain-specific behavioral traits is governed not only by the genetic background but also by the postnatal rearing environment. The level of maternal care is a representative environmental factor. For example, inbred Fischer 344 pups raised by Wistar dams showed Wistar-like behavioral traits in adulthood in socially interactive situations.³⁵ Furthermore, the effect of environmental factors is not limited to neonatal periods. In animals that form a society among conspecifics, such as mice and rats, postweaning social environments have a marked effect on behavioral traits. Many studies have reported that postweaning social isolation (pwSI) alters various strain-specific behavioral phenotypes, including aggressiveness,^15,32,34 novelty preference,²² locomotor activity,²⁹ anxiety-like behavior,²⁰ and learning and memory.^13,15,20,31 In other words, pwSI means the deprivation of several social interactions among conspecifics. The expression of social contact begins fundamentally in the postweaning periods, its frequency increases to a peak at 4 to 5 wk of age, and it declines thereafter until sexual maturity.^2,17,30 Therefore, rats were isolated from their conspecifics to deprive them of social contact during the current study.In the brain, the hippocampus is necessary for the acquisition of episodic and spatial memory,²⁸ but the influences of pwSI on hippocampal functions remain largely unclear. The hippocampus has a lamellar organization of neurons, and the intrinsic neuronal circuit of the hippocampus, termed the trisynaptic circuit, consists of 3 topographically and morphologically distinct neuronal layers: the pyramidal cell layer in subfields CA1 and CA3 and the granule cell layer in the dentate gyrus (DG). Sensory information is carried first to the DG by a perforant pathway that originates in the entorhinal cortex. DG granule cells project to the apical dendrites of the CA3 pyramidal cells through mossy fibers. In turn, CA3 pyramidal cells project to the CA1 layer through Schaffer collaterals.^1,18 Glutamate is a key excitatory neurotransmitter in the hippocampus and plays a central role in the activation of the trisynaptic circuit, whereas the inhibitory neurotransmitter GABA modulates the activated circuit. This excitatory–inhibitory balance is critical for the appropriate functioning of the hippocampal circuit. Here, we comprehensively investigated the effect of pwSI on the expression of glutamatergic and GABAergic neurotransmission-related genes in the 3 hippocampal subregions of inbred F344 rats.     

Global Gene Expression Analysis of Yeast Cells during Sake Brewing

During the brewing of Japanese sake, Saccharomyces cerevisiae cells produce a high concentration of ethanol compared with other ethanol fermentation methods. We analyzed the gene expression profiles of yeast cells during sake brewing using DNA microarray analysis. This analysis revealed some characteristics of yeast gene expression during sake brewing and provided a scaffold for a molecular level understanding of the sake brewing process.