• Toxicological Research
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• v.12 no.2
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• pp.203-211
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• 1996

Methanol has been widely used as an industrial solvent and environmental exposure to methanol would be expected to be increasing. In humans, methanol causes metabolic acidosis and damage to ocular system, and can lead to death in severe and untreated case. Clinical symptoms are attributed to accumulation of forrnic acid which is a metabolic product of methanol. In humans and primates, formic acid is accumulated after methanol intake but not in rodents due to the rapid metabolism of methanol. Neverthless, the developmental and reproductive toxicity were reported in rodents. Previous reports showed that perinatal exposure to ethanol produces a variety of damage in human central nervous system by direct neurotoxicity. This suggests that the mechanism of toxic symptoms by methanol in rodents might mimic that of ethanol in human. In the present study I hypothesized that methanol can also induce toxicity in neuronal cells. For the study, primary culture of rat hippocampal neurons and glias were empolyed. Hippocampal cells were prepared from the embryonic day-17 fetuses and maintained up to 7 days. Effect of methanol (10, 100, 500 and 1000 mM) on neurite outgrowth and cell viability was investigated at 0, 18 and 24 hours following methanol treatment. To study the changes in proliferation of glial cells, protein content was measured at 7 days. Neuronal cell viability in culture was not altered during 0-24 hours after methanol treatment. 10 and 100 mM methanol treatment significantly enhanced neurite outgrowth between 18-24 hours. 7-day exposure to 10 or 100 mM methanol significantly increased protein contents but that to 1000 mM methanol decreased in culture. In conclusion, methanol may have a variety of effects on growing and differentiation of neurons and glial cells in hippocampus. Treatment with low concentration of methanol caused that neurite outgrowth was enhanced during 18-24 hours and the numbers of glial cell were increased for 7 days. High concentration of methanol brought about decreased protein contents. At present, the mechanism responsible for the methanol- induced enhancement of neurite outgrowth is not clear. Further studies are required to delineate the mechanism possibly by employing molecular biological techniques.

• Korean Journal of Acupuncture
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• v.38 no.1
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• pp.32-42
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• 2021

Objectives : In this study, we investigated the neuroprotective effects of ethanol extract of Lonicera japonica flower buds (EELJ) on glutamate-induced neurotoxicity in mouse hippocampus-derived neuronal HT22 cells. Methods : After analyzing the cytoprotective effect of EELJ on glutamate in HT22 cells, the inhibitory effect of apoptosis was studied using flow cytometry. In order to analyze the antioxidant efficacy of EELJ, the levels of reactive oxygen species (ROS) and glutathione (GSH) were investigated, and the effects on the activities of superoxide dismutase (SOD) and catalase (CAT) were also analyzed. Furthermore, the effect of EELJ on the expression of apoptosis regulators such as Bax and Bcl-2 in glutamate-treated HT22 cells was investigated. Results : According the current results, pretreatment with EELJ significantly reduced glutamate-induced loss of cell viability and release of lactate dehydrogenase. EELJ also markedly attenuated glutamate-induced generation of intracellular ROS, which was associated with increased levels of GSH, and activity of SOD and CAT in glutamate-stimulated HT22 cells. In addition, EELJ was strikingly inhibited glutamate-induced apoptosis in HT22 cells. Furthermore, the expression of pro-apoptotic Bax was increased and the expression of anti-apoptotic Bcl-2 was decreased in glutamate-treated HT22 cells, while in the presence of EELJ, their expressions were maintained at the control levels. Conclusions : These findings indicate that EELJ protects glutamate-induced cytotoxicity in HT22 hippocampal neurons through antioxidant activity. Therefore, although identification of biologically active substances of EELJ and re-evaluation through animal experiments is necessary, this natural substance is a promising candidate for further research in preventing and treating oxidative stress-mediated neurodegenerative diseases.

• The Journal of Korean Medicine
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• v.29 no.2
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• pp.21-31
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• 2008

Backgrounds: Singihwan is used "to strengthen inborn energy" and we suspected a protective effect on brain neuron cells. Objectives: The aim of this study was to evaluate the effects of Singihwan (SGH) on traumatic brain injury-induced delayed apoptosis in rat hippocampal dentate gyrus. Methods: For a surgical induction of traumatic brain injury (TBI), a 5 mm diameter stainless rod was used to make traumatic attack from the surface of the brain used by an impactor. The protective effect of the aqueous extract of SGH against TBI in the rat hippocampal dentate gyrus was investigated by using step-down avoidance task, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, Bax immunohistochemistry, and 5-bromo-2'-deoxyuridine (BrdU) immunohistochemistry. Results: The aqueous extract of SGH suppressed the TBI-induced increase in apoptosis and cell proliferation in the hippocampal dentate gyrus. Conclusions: It is possible that the aqueous extract of SGH has a neuroprotective effect on TBI-induced neuronal cell death.

• Biomedical Science Letters
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• v.26 no.2
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• pp.57-65
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• 2020

Excessive drinking of alcohol is known to be one of the main causes of various neurological diseases, such as Alzheimer's disease. Scopoletin is known to have anti-inflammatory and antioxidative properties, and to protect nerve cells. This study examined whether scopoletin inhibits the alcohol-induced apoptosis of primary hippocampal neurons, and how scopoletin regulates several factors associated with the caspase-mediated pathway. To achieve this, the cell viability and apoptosis rate of primary hippocampal neurons were measured by Cell Counting Kit-8 and flow cytometry, respectively. Apoptosis-related protein expressions (Bax, Bid, caspase-3, caspase-9, and Poly (ADP-ribose) polymerase (PARP)) were analyzed by Western blotting, and the ANOVA method was used to confirm the significance of the measured results. As a result, scopoletin inhibited the expressions of alcohol-induced apoptosis and apoptosis-related proteins in primary hippocampal neurons. These results suggest that down-regulation of Bid, Bax, and cleaved caspase-9 expression induced by scopoletin down-regulates the expression of cleaved caspase-3, inhibits the expression of cleaved PARP, and finally, inhibits mitochondrial apoptotic pathways. The study suggests that scopoletin is worth developing as a candidate for neuroprotective agent.

• Journal of Oriental Neuropsychiatry
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• v.20 no.2
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• pp.31-46
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• 2009

Objectives : This experiment was designed to investigate the effect of Gojineumja(Guzhenyinzi, GJEJ) on damaged neural tissue in cultured glial cells and in the mouse brain tissue. Methods : The effects of the GJEJ on activation of astrocytes and caspase 3-positive cell counts in cultured glial cells administered with ${\beta}$-amyloid peptide were investigated. The effects of the GJEJ on levels of glial fibrillary acidic protein(GFAP)-positive reactive astrocyets and caspase 3-positive cells in the hippocampal subfields in the rats administered with scopolamine were investigated. Results : 1. GJEJ reduced levels of activated astrocytes and caspase 3-positive cell counts in cultured glial cells administered with ${\beta}$-amyloid peptide. 2. GJEJ reduced levels of GFAP-positive reactive astrocyets and caspase 3-positive cells in the hippocampal subfields in the rats administered with scopolamine. Conclusions : The present data. suggest that GJEJ may have a protective function of neuronal and non-neuronal cells in damaged neural tissue caused by AD-like stimulations. Further studies on identification of effective molecular components of GJEJ and their interactions with damaged neural cells would be important for understanding molecular mechanism and may be further applicable for the development of therapeutic strategies.

• The Korea Journal of Herbology
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• v.26 no.3
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• pp.75-82
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• 2011

Objectives : In this study, we evaluated the effect of Chungganhaeju-hwan(CGHJH) on hydrogen peroxide($H_2O_2$)-induced and ethanol(EtOH)-induced neuronal damage in vitro and in vivo, respectively. Methods:We carried out the anti-oxidant effects of CGHJH against hydrogen peroxide($H_2O_2$)-induced toxicity in HT22 and PC12 cells using thiazolyl blue tetrazolium bromide. Then, to investigate the protective effect on CGHJH against EtOH-induced memory impairment and hippocampal cell damage in male ICR mice, we performed novel object recognition test(NORT), and analysed the brain tissues after immunohistochemistry and western blotting. Results:CGHJH showed protective effect from $H_2O_2$-induced cell toxicity at doses of $1\sim100{\mu}g$/mL in both HT22 and PC12 cells. CGHJH had also recovery effect from EtOH-induced memory impairment in ICR mice from NORT and it protected hippocampal cells against EtOH toxicity in the result of cresyl violet and NeuN immunoreactivity. Conclusion : These results demonstrate that CGHJH has protective effect in neuronal cells against $H_2O_2$ and EtOH toxicities and this effect could be a main role of recovery effect on EtOH-induced memory loss.

• Journal of Integrative Natural Science
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• v.11 no.3
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• pp.139-147
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• 2018

Hippocampal atrophy is a well-established imaging biomarker of Alzheimer disease (AD). However, hippocampus is a non-homogenous structure with cytoarchitecturally and functionally distinct sub-regions or subfield, with each region performing distinct functions. Certain regions of the subfield have shown selective vulnerability to AD. Here, we are interested in studying the effects of normal aging and mild cognitive impairment on these sub-regional volumes. With a reliable automated segmentation technique, we segmented these subregions of the hippocampus in 101 cognitively normal (CN), 135 early mild cognitive impairment (EMCI), 67 late mild cognitive impairment (LMCI) and 48 AD subjects. Thereby, dividing the hippocampus into hippocampal tail (tail), subiculum (SUB), cornu ammonis 1 (CA1), hippocampal fissure (fissure), presubiculum (PSUB), parasubiculum (ParaSUB), molecular layer (ML), granule cells/molecular layer/dentate gyrus (GCMLDG), cornu ammonis 3(CA3), cornu ammonis 4(CA4), fimbria and hippocampal-amygdala transition area (HATA). In this cross sectional study of 351 ADNI subjects, no differences in terms of age, gender, and years of education were observed among the groups. Though, the groups had statistically significant differences (p < 0.05 after the multiple comparison correction) in the Mini-Mental State Examination (MMSE) scores. There was asymmetrical volume loss in the early stages of AD with the left hemisphere showing volume loss in regions that were unaffected in the right hemisphere. Bilateral parasubiculum, right cornu ammonis 1, 3 and 4, right fimbria and right HATA regions did not show any volume loss till the late MCI stages. Our findings suggest that the hippocampal subfield regions are selectively vulnerable to AD and also that these vulnerabilities are asymmetrical especially during the early stages of AD.

• Journal of Life Science
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• v.26 no.4
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• pp.475-483
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• 2016

The current study was conducted to evaluate beneficial effects of a new formula (CWC-9) using four traditional Oriental medicinal herbs, Cynanchum wilfordii, Rehmannia glutinosa, Polygala tenuifolia, and Acorus gramineus, on hippocampal cells and memory function. To examine the neuroprotective effects of a new four-herb extract, cell viability, cytotoxicity, and reactive oxygen species (ROS) assays were performed in HT22 cells and behavioral tests (Morris water maze and passive avoidance retention), Western blot, and immunohistochemistry were performed in a mouse model of focal cerebral ischemia. In HT22 hippocampal cells, pretreatment with CWC-9 resulted in significantly reduced glutamate-induced cell death with suppression of ROS accumulation caused by glutamate. In a mouse model of focal cerebral ischemia, we observed significant improvement of spatial and short-term memory function by treatment with CWC-9. Phosphorylated p38 mitogen-activated protein kinases (MAPK) in hippocampus of ischemic mice were decreased by treatment with CWC-9, but phosphorylated phosphatidylinositol-3 kinase (PI3K) and cAMP response element binding protein (CREB) were significantly enhanced. By immunohistochemical analysis, we confirmed higher expression of phosphorylation of CREB in the hippocampal neurons of CWC-9 treated mice. These results suggest that new multi-herb formula CWC-9 mainly exerted beneficial effects on cognitive function through regulation of neuro-protective signaling pathways associated with CREB.

• The Korea Journal of Herbology
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• v.30 no.2
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• pp.43-48
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• 2015

Objectives : The aim of this study was to investigate the memory enhancing properties of extract of Hoelen Cum Radix (HCR) and its possible mechanism in mice of normal condition. Methods : We evaluated the effects of HCR on cognitive function and memory enhancement in normal mice. Male ICR mice were orally administrated with HCR 100 mg/kg for 7 days and equal volume of saline was administrated to the control group in the same condition. We conducted two behavioral tests which measure the spatial working memory (Y-maze test) and cognitive fear memory (passive avoidance test). We also investigated whether HCR affects the hippocampal neurogenesis in the brain. To assess the effects of HCR on neural progenitor cell differentiation and neurite outgrowth in the early stage of hippocampal neurogenesis, we performed doublecortin (DCX), a direct neurogenesis marker, immunohistochemical analysis in the dentate gyrus (DG) of the mouse hippocampus. Results : HCR significantly enhanced memory and cognitive function as determined by the Y-maze test (p<0.05) and passive avoidance test (p<0.001). Moreover, HCR increased DCX positive cells (p<0.01) and neurite length (p<0.01) compared to the control group. These results indicated that HCR stimulates differentiation of neural progenitor cells and promotes neurite outgrowth in hippocampal DG of the mice. Conclusion : We concluded that HCR shows memory enhancing effects through the stimulation of hippocampal neurogenesis as a consequence of accelerated neuronal differentiation and neurite outgrowth in the DG of the hippocampus after HCR treatment.

• Korean Journal of Exercise Nutrition
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• v.23 no.3
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• pp.13-21
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• 2019

[Purpose] Chronic stress is a precipitating factor for depression, whereas exercise is beneficial for both the mood and cognitive process. The current study demonstrates the anti-depressive effects of regular exercise and the mechanisms linked to hippocampal neurogenesis. [Methods] Mice were subjected to 14 consecutive days of restraint, followed by 3 weeks of treadmill running, and were then subjected to behavioral tests that included the forced swimming and Y-maze tests. Protein levels were assessed using western blot analysis and newborn cells were detected using 5-bromo-2'-deoxyuridine (BrdU). [Results] Three weeks of treadmill running ameliorated the behavioral depression caused by 14 days of continuous restraint stress. The exercise regimen enhanced BrdU-labeled cells and class III β-tubulin levels in the hippocampal dentate gyrus, as well as those of thioredoxin-1 (TRX-1) and synaptosomal β2-adrenergic receptors (β2-AR) under stress. In vitro experiments involving treatment with recombinant human TRX-1 (rhTRX-1) augmented the levels of phospho-extracellular signal-regulated kinases 1 and 2 (ERK1/2), nuclear β-catenin, and proliferating cell nuclear antigens, which were previously inhibited by U0216 and FH535 (inhibitors of ERK1/2 and β-catenin/T cell factor-mediated transcription, respectively). The hippocampal neurogenesis elicited by a 7-day exercise regimen was abolished by a selective inhibitor of β2-AR, butoxamine. [Conclusion] These results suggest that TRX-1-mediated hippocampal neurogenesis by β2-AR function is a potential mechanism underlying the psychotropic effect of exercise.