Discrete Regions Of Mouse Brain Research Articles

Image-guided phenotyping of ovariectomized mice: altered functional connectivity, cognition, myelination, and dopaminergic functionality.

A large proportion of the population suffers from endocrine disruption, e.g., menopausal women, which might result in accelerated aging and a higher risk for developing cognitive disorders. Therefore, it is crucial to fully understand the impact of such disruptions on the brain to identify potential therapeutic strategies. Here, we show using resting-state functional magnetic resonance imaging that ovariectomy and consequent hypothalamus-pituitary-gonadal disruption result in the selective dysconnectivity of 2 discrete brain regions in mice. This effect coincided with cognitive deficits and an underlying pathological molecular phenotype involving an imbalance of neurodevelopmental/neurodegenerative signaling. Furthermore, this quantitative mass spectrometry proteomics-based analysis of molecular signaling patterns further identified a strong involvement of altered dopaminergic functionality (e.g., DAT and predicted upstream regulators DRD3, NR4A2), reproductive signaling (e.g., Srd5a2), rotatin expression (rttn), cellular aging (e.g., Rxfp3, Git2), myelination, and axogenesis (e.g., Nefl, Mag). With this, we have provided an improved understanding of the impact of hypothalamus-pituitary-gonadal dysfunction and highlighted the potential of using a highly translational magnetic resonance imaging technique for monitoring these effects on the brain.

The use of microwave irradiation for quantitative analysis of neurotransmitters in the mouse brain

BackgroundAssessing neurotransmitter metabolism in the brain is essential in studying the effects of drugs, dietary modification and characterizing transgenic mouse models of human neurodegenerative diseases. Regional brain concentrations of parent neurotransmitters and related metabolites are informative and provide a snap shot of the steady-state levels. The choice in method of sacrificing mice may differ from one laboratory to another, and the technique in removal of brain may have limitations depending on speed in which tissue can be dissected and frozen to prevent post-mortem changes. New methodsIn order to better assess neurotransmitter metabolism in an effective and standardized manner we evaluated microwave irradiation as a method of sacrificing mice. Mice were sacrificed by CO2 asphyxiation followed by cervical dislocation or microwave irradiation at 4 Kw for 1.1 s. Brain tissue was harvested into five regions and stored at −80 °C until analysis by either LC–MS/MS for acetylcholine, choline and GABA, or HPLC-EC for dopamine, serotonin and norepinephrine and related metabolites. ResultsThe results of our study showed considerable differences in the levels of neurotransmitters between the two methods of sacrifice. Overall, the concentrations of neurotransmitters were higher in mice sacrificed by microwave irradiation, except for GABA, which was lower. Comparison with existing method(s)Previous microwave irradiation studies employed presently outdated equipment and neurotransmitter analysis methods, and were not as comprehensive. ConclusionsThe combination of microwave irradiation with LC–MS/MS and HPLC-EC detection allows accurate and sensitive measurement of several neurotransmitter systems in discrete mouse brain regions.

A simple and sensitive high-performance liquid chromatography-electrochemical detection assay for the quantitative determination of monoamines and respective metabolites in six discrete brain regions of mice.

A rapid, sensitive, and reproducible assay is described for the quantitative determination of the monoamine neurotransmitters dopamine, norepinephrine and serotonin, their metabolites, and the internal standard 3,4-dihydroxybenzlyamine hydro-bromide in mouse brain homogenate using high-performance liquid chromatography with electrochemical detection. The method was validated in the following brain areas: frontal cortex, striatum, nucleus accumbens, hippocampus, substantia nigra pars compacta and ventral tegmental area. Biogenic amines and relevant metabolites were extracted from discrete brain regions using a simple protein precipitation procedure, and the chromatography was achieved using a C18 column. The method was accurate over the linear range of 0.300-30 ng/mL (r = 0.999) for dopamine and 0.300-15 ng/mL (r = 0.999) for norepinephrine, 3,4-dihydroxybenzlyamine hydro-bromide, homovanillic acid and 5-hydroxyindolacetic acid, with detection limits of ~0.125 ng/mL (5 pg on column) for each of these analytes. Accuracy and linearity for serotonin were observed throughout the concentration range of 0.625-30 ng/mL (r = 0.998) with an analytical detection limit of ~0.300 ng/mL (12 pg on column). Relative recoveries for all analytes were approximately ≥90% and the analytical run time was <10 min. The described method utilized minimal sample preparation procedures and was optimized to provide the sensitivity limits required for simultaneous monoamine and metabolite analysis in small, discrete brain tissue samples.

Analysis of gene expression in mouse brain regions after exposure to 1.9 GHz radiofrequency fields

Purpose: To assess 1.9 GHz radiofrequency (RF) field exposure on gene expression within a variety of discrete mouse brain regions using whole genome microarray analysis. Materials and methods: Adult male C57BL/6 mice were exposed to 1.9 GHz pulse-modulated or continuous-wave RF fields for 4 h/day for 5 consecutive days at whole body average (WBA) specific absorption rates of 0 (sham), ∼0.2 W/kg and ∼1.4 W/kg. Total RNA was isolated from the auditory cortex, amygdala, caudate, cerebellum, hippocampus, hypothalamus, and medial prefrontal cortex and differential gene expression was assessed using Illumina MouseWG-6 (v2) BeadChip arrays. Validation of potentially responding genes was conducted by RT-PCR. Results: When analysis of gene expression was conducted within individual brain regions when controlling the false discovery rate (FDR), no differentially expressed genes were identified relative to the sham control. However, it must be noted that most fold changes among groups were observed to be less than 1.5-fold and this study had limited ability to detect such small changes. While some genes were differentially expressed without correction for multiple-comparisons testing, no consistent pattern of response was observed among different RF-exposure levels or among different RF-modulations. Conclusions: The current study provides the most comprehensive analysis of potential gene expression changes in the rodent brain in response to RF field exposure conducted to date. Within the exposure conditions and limitations of this study, no convincing evidence of consistent changes in gene expression was found in response to 1.9 GHz RF field exposure.

Suppression of mitochondrial oxidative phosphorylation and TCA enzymes in discrete brain regions of mice exposed to high fluoride: amelioration by Panax ginseng (Ginseng) and Lagerstroemia speciosa (Banaba) extracts.

Chronic fluoride intoxication results in pathophysiological complications pertaining to soft tissues, called non-skeletal fluorosis. This study examined whether fluoride-induced alterations in selected parameters that are indicative of mitochondrial dysfunction accompany the toxic effects of fluoride in discrete brain regions in vivo and also explored the possibility of treatment with Ginseng (GE) and Banaba (BLE) either alone or with their co-exposure which is capable of reversing parameters indicative of fluoride-induced impairments in mitochondrial function. Swiss mice, Mus musculus, were given 270 ppm fluoride (600 ppm NaF) in their drinking water for 30 days, while continuing the fluoride exposure, toxicated animals were given differential doses (50-250 mg/kg body wt) of phytoextracts through oral gavage for 2 weeks. Discrete brain regions separated from dissected animals to perform biochemical assessments. Disturbances in mitochondrial enzyme complexes (I-IV) and decrements in TCA enzymes (ICDH, SDH, and aconitase) were noted in discrete brain regions upon F exposure, suggesting mitochondrial dysfunction. In addition, a significant reduction in oxidative stress indices with increased MDA content as well as decrease in reduced glutathione content and increases in catalase and SOD enzyme activity suggests the involvement of severe oxidative stress affecting the mitochondrial function(s). Treatment with either GE or BLE reversed F-induced alterations in augmenting the suppressed complex enzymes followed by TCA enzymes and oxidative stress indices in a dose independent manner. However, the co-exposure of GE and BLE at a dose of 150 mg/kgbw appeared to restore mitochondrial functioning. These results provide in vivo evidence supporting the hypothesis that fluoride induces impairments in mitochondrial function, which can be reversed by treatment with GE and BLE as well their co-exposure at 150 mg/kgbw.

Non-Laser Capture Microscopy Approach for the Microdissection of Discrete Mouse Brain Regions for Total RNA Isolation and Downstream Next-Generation Sequencing and Gene Expression Profiling

As technological platforms, approaches such as next-generation sequencing, microarray, and qRT-PCR have great promise for expanding our understanding of the breadth of molecular regulation. Newer approaches such as high-resolution RNA sequencing (RNA-Seq)(1) provides new and expansive information about tissue- or state-specific expression such as relative transcript levels, alternative splicing, and micro RNAs(2-4). Prospects for employing the RNA-Seq method in comparative whole transcriptome profiling(5) within discrete tissues or between phenotypically distinct groups of individuals affords new avenues for elucidating molecular mechanisms involved in both normal and abnormal physiological states. Recently, whole transcriptome profiling has been performed on human brain tissue, identifying gene expression differences associated with disease progression(6). However, the use of next-generation sequencing has yet to be more widely integrated into mammalian studies. Gene expression studies in mouse models have reported distinct profiles within various brain nuclei using laser capture microscopy (LCM) for sample excision(7,8). While LCM affords sample collection with single-cell and discrete brain region precision, the relatively low total RNA yields from the LCM approach can be prohibitive to RNA-Seq and other profiling approaches in mouse brain tissues and may require sub-optimal sample amplification steps. Here, a protocol is presented for microdissection and total RNA extraction from discrete mouse brain regions. Set-diameter tissue corers are used to isolate 13 tissues from 750-μm serial coronal sections of an individual mouse brain. Tissue micropunch samples are immediately frozen and archived. Total RNA is obtained from the samples using magnetic bead-enabled total RNA isolation technology. Resulting RNA samples have adequate yield and quality for use in downstream expression profiling. This microdissection strategy provides a viable option to existing sample collection strategies for obtaining total RNA from discrete brain regions, opening possibilities for new gene expression discoveries.

Non-Laser Capture Microscopy Approach for the Microdissection of Discrete Mouse Brain Regions for Total RNA Isolation and Downstream Next-Generation Sequencing and Gene Expression Profiling

Non-Laser Capture Microscopy Approach for the Microdissection of Discrete Mouse Brain Regions for Total RNA Isolation and Downstream Next-Generation Sequencing and Gene Expression Profiling

Effects of Atomoxetine on Levels of Monoamines and Related Substances in Discrete Brain Regions in Mice Intermittently Deprived of Rapid Eye Movement Sleep

An imbalance between noradrenergic and dopaminergic systems is implicated in hyperactivity disorders, such as attention deficit/hyperactivity disorder (ADHD). We previously showed that the explosive jumping behavior elicited by intermittent rapid eye movement sleep deprivation (REMSD) may serve as a useful model of ADHD (see [Biogenic Amines, 20, 99-111]). Here, we investigated whether intermittent REMSD might cause changes in monoamine turnover in the mouse forebrain. Our main findings were as follows. Intermittent REMSD led to a significant decrease in dopamine turnover and a significant increase in noradrenaline turnover in the frontal cortex. The latter effect, but not the former, was attenuated by atomoxetine, which is used clinically to treat ADHD symptoms. These results suggest (a) that intermittent REMSD induces hypernoradrenergic and hypodopaminergic states within the frontal cortex, and (b) that the therapeutic effects of atomoxetine may include an inhibition of this hypernoradrenergic state.

A protein–protein interaction of stress‐responsive myosin VI endowed to inhibit neural progenitor self‐replication with RNA binding protein, TLS, in murine hippocampus

We have shown preferential expression of both mRNA and corresponding protein for myosin VI (Myo6) in the murine hippocampus within 24 h after the extreme traumatic experience, water-immersion restraint stress (WIRS), prior to a drastic decrease in neural progenitor proliferation in the dentate gyrus. Myosin (Myo6) protein levels were significantly increased in hippocampus within 24 h after flashback experience in mice previously exposed to WIRS. Myo6 protein was ubiquitously distributed in discrete mouse brain regions with exceptionally high expression in olfactory bulb, whereas Myo6 protein was expressed in cultured rat astroglia and neurons, in addition to Myo6 mRNA expression by cultured neural progenitors. In mouse embryonal carcinoma P19 cells endowed to proliferate and differentiate, Myo6 protein was expressed in line with astroglial marker protein expression. Transient over-expression of Myo6 induced a significant decrease in the size of clustered aggregates as an index of self-replication in P19 cells. Immunoprecipitation analysis revealed the interaction between Myo6 and the RNA-binding protein, translocated in liposarcoma (TLS), while TLS was predominantly expressed by neurons in the cortex, striatum, cerebellum, and hippocampus. These results suggest that Myo6 may play a pivotal role in the mechanism underlying the suppressed adult neurogenesis after traumatic stress in association with TLS.

LPS‐INDUCED STAT3 AND CYTOKINE PROTEIN IN MOUSE BRAIN

The peripheral immune system and the brain communicate through signaling mechanisms. Immune activation alters central nervous system (CNS) and the CNS modulates immune function. During infection there are changes in complex behavior, including sleep. What is not well known is by what mechanisms, and where, in brain immune signals act to alter sleep. Cytokines, such as IL‐1, TNF and IL‐6 mediate infection‐induced alterations in sleep. Receptor signaling for these cytokines include Signal Transducer and Activator of Transcription 3 (STAT3). To determine a potential role in altered sleep during immune challenge, we injected LPS into several mouse strains and determined changes in STAT3 and cytokine in discrete brain regions.Adult male mice (C57BL/6J; Balb/c; TLR4 mutant; n = 3‐4 / strain / condition) were injected IP at light onset with either 0.2ml vehicle or 10μg LPS (E. coli serotype O111:B4). These animals were sacrificed 2‐ or 4 h post injection. STAT3 from 2 h treatments and cytokine from 4 h treatments were analyzed in brain and spleen using Bioplex assays.STAT3 protein increased from 4 ‐ 10 fold 2 h after LPS in all samples assayed from C57BL/6J mice and Balb/c mice. In these two mouse strains, IL‐1, TNF and IL‐6 increased from 2.5 ‐ 11 fold 4 h after LPS. In contrast, due to the defect in TLR4 a signaling transducer for LPS, there were no LPS‐induced increases in STAT3 or in cytokine protein in the TLR4 mutant mice.These results demonstrate that the STAT3 signaling pathway for proinflammatory cytokines may be activated in discrete mouse brain regions and suggests that these pathways play a role in transducing peripheral immune signals into alterations in CNS‐regulated sleep/behavior.

Lipopolysaccharide-induced increases in cytokines in discrete mouse brain regions are detectable using Luminex xMAP ® technology

Methods to determine cytokine protein content in samples of interest, such as enzyme-linked immunosorbent assay (ELISA), are often labor-intensive and costly. Furthermore, because ELISA requires relatively large sample volumes and protein concentrations, it is difficult using this technique to determine protein content for multiple cytokines from individual samples. Recently, Luminex has developed an open source hardware platform combining flow cytometry- and bead-based antibody capture that is capable of detecting multiple analytes from a single sample. In the present study we employed the Luminex 200 ® platform to determine the cytokine protein content in discrete brain regions of C57BL/6J mice. In spike-and-recovery experiments, known concentrations of murine recombinant interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)α were added either singly or as a mixture of all three to whole brain homogenates containing known quantities of total protein. Spiked samples were assayed for either a single cytokine or for multiple cytokines using 1-plex or 3-plex assay kits, respectively. In whole mouse brain homogenate we recovered between 81% and 103% of the recombinant cytokines. We then injected C57BL/6J mice intraperitoneally with bacterial lipopolysaccharide (LPS) and sacrificed them 4 h later. We detected in samples taken from LPS-stimulated mice 4- to 870-fold increases in serum or spleen cytokine protein, and 1.5- to 16-fold increases in cytokine protein in discrete brain regions, relative to protein content in samples obtained from vehicle-treated animals. These results indicate that multiple cytokines may be reliably assayed from discrete regions of mouse brain using a single sample.

Atropine, a muscarinic cholinergic receptor antagonist increases serotonin, but not dopamine levels in discrete brain regions of mice

We investigated the effects of atropine, a muscarinic acetylcholine (ACh) receptor antagonist, on the level of serotonin in discrete brain regions, the nucleus raphe dorsalis (NRD), nucleus caudatus putamen (NCP), cerebral cortex and the cerebellum. Biogenic amines were assayed employing HPLC electrochemistry in these regions 30 min following different doses of atropine (5, 10, 25mg/kg; i.p.), and at various time points (15, 30, 60, 120 min) after 25mg/kg of the drug. The cholinergic receptor antagonist caused a dose-dependent alteration in the level of serotonin in NRD, but the increase was not dose-dependent for other regions studied. The metabolite of serotonin, 5-hydroxyindoleacetic acid was unaffected. Atropine did not affect the levels of dopamine or its metabolites dihydroxyphenyl acetic acid and homovanillic acid. The present study suggests significant effect of this antimuscarinic agent on the synthesis of serotonin in the central serotoninergic pathways, which may have clinical relevance.

Chlorpyrifos induces pro‐inflammatory environment in discrete regions of mouse brain

It has been proposed that exposure to environmental toxicants plays a role in the development of neurodegenerative diseases. Mechanisms of action underlying this process, however, have not yet been clearly established. The present study was designed to address potential mechanisms that contribute to brain injury after exposure to the organophosphorus insecticide chlorpyrifos. C57BL/6 mice were administered a single intraperitoneal injection of chlorpyrifos in increasing dosages (20, 70, and 140 mg/kg). At 4, 8, 16, and 24 h of exposure, four discrete regions of the brain (hippocampus, cerebral cortex, striatum, and cerebellum) were isolated, and expression levels of pro-inflammatory mediators were assessed by quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR). A significant increase in expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), MCP-1, and E-selectin was observed in all brain regions in a time- and dose-dependent manner. These data suggest that chlorpyrifos-induced brain injury may be mediated through pro-inflammatory pathways. It also offers the potential to contribute to the development of new therapeutic strategies specifically targeted against pro-inflammatory pathways of environmental agent-induced neurotoxicity (This work was supported by SBES Seed Grants).

LPS induced alterations in proinflammatory cytokine expression in mouse brain

Bacterial cell-wall lipopolysaccharide (LPS) induces peripheral release of interleukin (IL)1β, IL6 and tumor necrosis factor (TNF)α. These cytokines may influence CNS mechanisms regulating sleep by activating neuronal pathways in brain. We previously demonstrated that LPS-induced fever and increases in sleep are attenuated by inhibition of nuclear translocation of NFkB in mouse brain. To date, there are few studies of the impact of LPS on cytokine expression in discrete mouse brain regions involved in the regulation of sleep. In the present study, we determined cytokine mRNA and protein content in mouse brain after LPS challenge. Adult male mice (~25 g; C57BL/6J) were injected IP at light onset with 10μg LPS (E.coli serotype O111:B4) or vehicle (PFS) and sacrificed 2 or 4 h later. Discrete brain regions were dissected, and tissues frozen and stored at −80°C. RNA was extracted from the 2 h group ( LPS, n=8; PFS, n=8). QPCR (TaqMan; Applied Biosystems) and the comparative threshold cycle (ΔΔCT) method was used to determine cytokine mRNA. LPS increased IL1β, IL6 and TNFα mRNA in hypothalamus, hippocampus and brain stem by 2 to 7 fold. Protein was extracted from the 4 h group for Bio-Plex assay (BioRad; LPS, n=6, ;PFS, n=6). Cytokine protein in these brain regions increased by 3 to 5 fold. Present results, and our previous data, suggest that LPS-induced increases in cytokines in brain are involved in alterations in sleep after this challenge. Studies are underway to determine if inhibition of nuclear translocation of NFkB blocks LPS-induced increases in cytokine protein in mouse brain.

Cocaine-induced kindling is associated with elevated NMDA receptor binding in discrete mouse brain regions

The present study was undertaken to investigate the involvement of N-methyl- d-aspartate (NMDA) type of glutamate receptors in the induction and maintenance of kindling generated by daily cocaine (35 mg/kg) injections to Swiss Webster mice. In addition, the regulation of NMDA receptor binding following the development of sensitization to horizontal locomotor activity produced by daily injections of a low dose of cocaine (15 mg/kg for 5 days) was investigated. Three days following the administration of the high dose of cocaine (35 mg/kg) a marked augmentation in cocaine-induced horizontal and vertical activities was observed (induction phase). Subsequently, after 10 days of cocaine administration, mice developed stage 5 seizures (Racine scale). Binding of [ 3H]CGP 39653 to the NMDA receptors revealed a marked increase in receptor densities in the striatum, amygdala and hippocampus associated with the induction phase. The elevation of NMDA receptor binding in the striatum and amygdala was sustained for 10 days following the induction phase. The pattern of altered NMDA receptor binding following the expression of cocaine kindled seizures was different. One day after the expression of kindled seizures NMDA receptor binding was elevated in striatum, amygdala, hippocampus and frontal cortex. However, only the elevation of NMDA receptor binding in the amygdala and hippocampus was sustained for 10 days following the expression of cocaine kindled seizures. In the brains of mice sensitized to the low dose of cocaine (15 mg/kg) no change in NMDA receptor binding was observed compared with control values. The present findings suggest the following: (a) The induction of cocaine kindling is associated with increased NMDA receptor binding activity in the striatum, amygdala and hippocampus; (b) the maintenance of cocaine kindling depends on increased NMDA receptor binding in the amygdala and hippocampus; (c) sensitization to cocaine-induced horizontal locomotor activity may be independent of elevation in NMDA receptor binding.

Effects of ketamine on dopamine metabolism during anesthesia in discrete brain regions in mice: comparison with the effects during the recovery and subanesthetic phases

The effects of ketamine on the levels of dopamine (DA), norepinephrine (NE), 5-hydroxytryptamine (5-HT, serotonin) and their metabolites were examined in discrete brain regions in mice. A high dose of ketamine (150 mg/kg, i.p.) did not change DA metabolism in the frontal cortex, nucleus accumbens, striatum and hippocampus, but did decrease it in the brainstem during anesthesia. In contrast, during recovery from the ketamine anesthesia, the high dose increased the level of homovanillic acid (HVA) in all brain regions. A low subanesthetic dose of ketamine (30 mg/kg, i.p.) increased the concentrations of both 3,4-dihydroxyphenylacetic acid (DOPAC) and HVA only in the nucleus accumbens. The DA level was not affected by any ketamine treatment. During ketamine anesthesia, the content of 3-methoxy-4-hydroxy-phenylglycol (MHPG) was decreased in the brainstem, whereas during recovery from anesthesia, the MHPG level was increased in the frontal cortex, nucleus accumbens and brainstem. The NE content was not altered in any region by ketamine treatment. The concentration of 5-hydroxyindoleacetic acid (5-HIAA) was reduced in the frontal cortex, striatum, hippocampus and brainstem during ketamine anesthesia. The 5-HT level was unaltered in all regions except the brainstem where it was reduced. In contrast, after anesthesia, the concentrations of both 5-HT and 5-HIAA were increased in the striatum. During the subanesthetic phase, however, the levels of NE, 5-HT and their metabolites were unchanged. These neurochemical results are consistent with the electrophysiological findings that a high dose of ketamine does not change the basal firing rates of nigrostriatal DA neurons during anesthesia, while low subanesthetic doses significantly increase those of ventral tegmental DA neurons.

Hyperlocomotion during recovery from isoflurane anesthesia is associated with increased dopamine turnover in the nucleus accumbens and striatum in mice.

It was recently reported that isoflurane increases dopamine release in the striatum in rats both in vivo and in vitro, and that isoflurane inhibits uptake of dopamine in the rat brain synaptosomes. However, the functional role of these effects of isoflurane on dopamine neurons is uncertain. Dopaminergic mechanisms within the nucleus accumbens and striatum play an important role in the control of locomotor activity, and a change in dopamine turnover depends essentially on a change in impulse flow in the dopamine neurons. In this study, the effects of isoflurane on locomotor activity and on dopamine turnover were investigated in discrete brain regions in mice. Mice were placed in individual airtight clear plastic chambers and spontaneously breathed isoflurane in 25% oxygen and 75% nitrogen (fresh gas flow, 4 l/min). Locomotor activity was measured with an Animex activity meter. Animals were decapitated after treatments with or without isoflurane, and the concentrations of monoamines and their metabolites in different brain areas were measured by high-performance liquid chromatography. During the 10 min after the cessation of the 20-min exposure to isoflurane, there was a significant increase in locomotor activity in animals breathing 1.5% isoflurane but not 0.7% isoflurane. This increase in locomotor activity produced by 1.5% isoflurane was abolished by a low dose of haloperidol (0.1 mg/kg), a dopamine receptor antagonist. Regional brain monoamine assays revealed that 1.5% isoflurane significantly increased the 3,4-dihydroxyphenylacetic acid:dopamine ratio (one indicator of transmitter turnover) in the nucleus accumbens and striatum, but a concentration of 0.7% did not. This significant increase in dopamine turnover in these regions continued during 20 min after the cessation of the administration of 1.5% isoflurane. These results suggest that isoflurane-induced hyperlocomotion during emergence may be associated with increased dopamine turnover in the nucleus accumbens and striatum.

Effects of chaihu-guizhi-ganjiang-tang on the levels of monoamines and their related substances, and acetylcholine in discrete brain regions of mice.

The effects of the extract powder (CggT) from Chaihu-Guizhi-Ganjiang-Tang (Saiko-keishi-kankyo-to, in Japanese) on the monoamines and their related substances and the acetylcholine in mouse brain were examined. 1) A single administration of CggT significantly increased the levels of HVA and 5-HIAA in the cerebral cortex, hypothalamus, corpus striatum and hippocampus at 75 mg/kg, and those in the hypothalamus, corpus striatum and hippocampus at 750 mg/kg. 2) The repeated administration of CggT significantly increased the level of 5-HT in the hippocampus at 75 mg/kg, and the levels of 5-HT in the corpus striatum and of NE and 5-HT in the hippocampus at 750 mg/kg. 3) The level of ACh was significantly increased in the hypothalamus alone after single administration of CggT. These findings suggest that CggT stimulates function of the dopaminergic and serotonergic nervous systems in mice, but not most of the NEnergic and cholinergic nervous systems.

Effects of GBR 12909 on locomotor activity and dopamine turnover in mice: comparison with apomorphine

The effects of GBR 12909 1-[2-[bis(4-fluorophenyl)methoxy]-ethyl]-4-[3-phenylpropyl]piperazine, a very potent and selective dopamine uptake inhibitor, and apomorphine, a dopamine receptor agonist, alone and in combination were investigated on locomotor activity and dopamine turnover in discrete brain regions of mice. The levels of dopamine and its metabolites were examined 40 min after the administration of GBR 12909 and/or apomorphine, when the effects of the drugs on locomotor activity were approximately at a peak. GBR 12909 (10 mg/kg i.p.) reversed a low dose of apomorphine (0.05 mg/kg s.c.)-induced suppression in locomotor activity and significantly increased this activity. Despite the dramatic change in the behavior, GBR 12909 did not influence the decrease in 3,4-dihydroxyphenylacetic acid (DOPAC)/dopamine ratio (which is one of the indications of transmitter turnover) induced by a low dose of apomorphine in the nucleus accumbens and striatum. In contrast, GBR 12909 did not enhance the high-dose apomorphine (2 mg/kg s.c.)-induced hyperlocomotion, and did not modify the larger decrease in dopamine turnover produced by the high dose of apomorphine in the frontal cortex, nucleus accumbens and striatum. This suggests that postsynaptic dopamine receptors may reach maximum stimulation at a high dose of apomorphine. These results indicate that a behavioral change induced via stimulation of postsynaptic dopamine receptors does not necessarily lead to an alteration in dopamine turnover.

A rapid assay for neurotransmitter amino acids, aspartate, glutamate, glycine, taurine and gamma-aminobutyric acid in the brain by high-performance liquid chromatography with electrochemical detection.

For simultaneous assay of the five neurotransmitter amino acids, Asp, Glu, Gly, Tau, and GABA in brain tissues, a very rapid and simple chromatographic method using high-performance liquid chromatography with electrochemical detection in combination with o-phthalaldehyde derivatization is described. Because the present method permits the determination of these five amino acids within less than five minutes in one chromatographic run, up to 100 samples a working day can be analyzed using an autosampler. Within-run coefficients of variation for these five amino acids were less than 2% (n = 20). The quantitative detection limit was 2.5 pmol for the 5 amino acids. The present method has been applied to the measurement of the five amino acid neurotransmitter levels in several discrete brain regions of mice treated with and without electroconvulsive shock.