'"''Alterations of tyrosine hydroxylase activity in various regions of brain from rats postnatally exposed to lead were tested. Three groups of animals were prepared; (1) Rats exposed te lead at a lew dose (O.05 % lead acetate, PbAc) ; (2) Rats exposed to lead at a high dose (O.2 % PbAc) ; (3) Age-matched normal control rats. At 2, 4, 6, and g weeks of age, weight of brain and body, and concentrations of iead in whole brain of animals in each group were measured, Activities of tyrosine hydroxylase and Na+-K+ ATPase were also measured at the same ages in 4 brain regions of each animal. Body weight gain was decreased after 6 weeks of age in rats exposed to lead at a high dose. Concentrations of lead ln whele brain were increased from O.37 to O,83 (ng!mg wet tissue) in these animals. Exposure of rats to lead generally increased tyrosine hydroxylase activity and decreased Na+-K+ ATPase activity. However, changes of tyrosine hydroxylase activiry were detected without concomitant changes of Na+-K+ ATPase activity in pons-medulla at 2 weeks of age and tetencephalon at 6 weeks of age in rats exposed to lead at a low dose, and in midbrain at 4 and 6 weeks of age in rats exposed to lead at a high dose. These data imply that catecholarninergic nervous system in the brain regions described above could be selectiyely affected by iead,

Marine actinomycetes have been drawing attention as prolific sources of new bioactive secondary metabolites [1]. In our research for new bioactive secondary metabolites form marine microorganisms, we selectively isolated actinomycete strains from marine sediment samples from the western area of Jeju Island in Republic of Korea and screened their secondary metabolites profiles by LC/MS. During the chemical profile analysis, we observed that one of the actinomycete strains named TH11 (Streptomyces sp.) produces previously reported antifungal butyrolactols A and B, which bear a seven consecutive 1,2-diol moiety [2], and their new derivatives, butyrolactols C and D. Because the absolute configuration of butyrolactol A has not previously determined, we established the stereochemistry of butyrolactol A by J-based configuration analysis using ROESY and HETLOC data and Mosher's method. Biosynthetically interestingly, four oxygen-bearing carbons are missing between the 5-membered lactone ring and the chain with a tertiary butyl group in butyrolactol C compared to butyrolactol A.

J. Neurochem. (2012) 123, 226-238. ABSTRACT:Fragile X syndrome (FXS), the most common single genetic cause of mental retardation and autistic spectrum disease, occurs when FMR1 gene is mutated. FMR1 encodes fragile X mental retardation protein (FMRP) which regulates translation of mRNAs playing important roles in the development of neurons as well as formation and maintenance of synapses. To examine whether FMRP regulates cell viability, we induced apoptosis in rat primary cortical neurons with glutamate in vitro and with middle cerebral artery occlusion (MCAO) in striatal neurons in vivo. Both conditions elicited a rapid, but transient FMRP expression in neurons. This up-regulated FMRP expression was abolished by pre-treatment with PI3K and Protein Kinase B (Akt) inhibitors: LY294002, Akt inhibitor IV, and VIII. Reduced FMRP expression in vitro or in vivo using small hairpin Fmr1 virus exacerbated cell death by glutamate or MCAO, presumably via hypophosphorylation of Akt and reduced expression of B-cell lymphoma-extra large (Bcl-xL). However, over-expression of FMRP using enhanced green fluorescent protein (eGFP)-FMRP constructs alleviated cell death, increased Akt activity, and enhanced Bcl-xL production. The pro-survival role of Akt-dependent up-regulation of FMRP in glutamate-stimulated cultured neuron as well as in ischemic brain may have a clinical importance in FXS as well as in neurodegenerative disorders and traumatic brain injury.

Autism is a spectrum of neurodevelopmental disorders characterized by social isolation and lack of interaction. Anatomically, autism patients often show macrocephaly and high neuronal density. To investigate the mechanism underlying the higher neuronal populations seen in ASD, we subcutaneously injected VPA (400 mg/kg) into pregnant Sprague-Dawley rats on E12, an animal model often used in ASD study. Alternatively, cultured rat neural progenitor cells were treated with VPA. Until E18, VPA induced NPC proliferation and delayed neurogenesis in fetal brain, but the subsequent differentiation of NPCs to neurons increased brain neuronal density afterward. Similar findings were observed with NPCs treated with VPA in vitro. At a molecular level, VPA enhanced Wnt1 expression and activated the GSK-3β/β-catenin pathway. Furthermore, inhibition of this pathway attenuated the effects of VPA. The findings of this study suggest that an altered developmental process underlies the macrocephaly and abnormal brain structure observed in the autistic brain.

Oroxylin A is a flavone isolated from a medicinal herb reported to be effective in reducing the inflammatory and oxidative stresses. It also modulates the production of brain derived neurotrophic factor (BDNF) in cortical neurons by the transactivation of cAMP response element-binding protein (CREB). As a neurotrophin, BDNF plays roles in neuronal development, differentiation, synaptogenesis, and neural protection from the harmful stimuli. Adenosine A2A receptor colocalized with BDNF in brain and the functional interaction between A2A receptor stimulation and BDNF action has been suggested. In this study, we investigated the possibility that oroxylin A modulates BDNF production in cortical neuron through the regulation of A2A receptor system. As ex-pected, CGS21680 (A2A receptor agonist) induced BDNF expression and release, however, an antagonist, ZM241385, prevented oroxylin A-induced increase in BDNF production. Oroxylin A activated the PI3K-Akt-GSK-3β signaling pathway, which is inhibited by ZM241385 and the blockade of the signaling pathway abolished the increase in BDNF production. The physiological roles of oroxylin A-induced BDNF production were demonstrated by the increased neurite extension as well as synapse formation from neurons. Overall, oroxylin A might regulate BDNF production in cortical neuron through A2A receptor stimulation, which promotes cellular survival, synapse formation and neurite extension.

Alzheimer's disease (AD) is a neurodegenerative disorder associated with progressive cognitive and memory loss and neuronal cell death. Current therapeutic strategies for AD are very limited; thus, traditional herbal medicines or their active constituents receive much attention. The aim of this study was to investigate the cognitive enhancing effects of salvianolic acid B (SalB) isolated from Salvia miltiorrhiza and its ameliorating effects on various drug-induced amnesic models using the passive avoidance, Y-maze, and Morris water maze tasks. Drug-induced amnesia was induced by administering scopolamine, diazepam, muscimol, or amyloid-beta (A beta)(25-35) peptide. SalB (10 mg/kg, p.o.) was found to significantly reverse the cognitive impairments induced by scopolamine (1 mg/kg, i.p.) or A beta(25-35) (10 nmol/5 mu l, i.c.v.) injection. This ameliorating effect of SalB was antagonized by the GABA(A) receptor agonists, muscimol or diazepam, respectively. In addition, SalB alone was capable of improving cognitive performances. Furthermore, SalB (100 mu M) was found to inhibit GABA-induced outward Cl- currents in single hippocampal CA1 neuron. These results suggest that the observed ameliorations of cholinergic dysfunction- or A beta(25-35)-induced memory impairment by SalB were mediated, in part, via the GABAergic neurotransmitter system after a single administration. (C) 2011 Elsevier Ltd. All rights reserved.

As a member of neurotrophin family, brain derived neurotrophic factor (BDNF) plays critical roles in neuronal development, differentiation, synaptogenesis, and neural protection from the harmful stimuli. There have been reported that adenosine A2(A) receptor subtype is widely distributed in the brain regions, such as hippocampus, striatum, and cortex. Adenosine A2(A) receptor is colocalized with BDNF in brain regions and the functional interaction between A2(A) receptor stimulation and BDNF action has been suggested. In this study, we investigated the possibility that the activation of A2(A) receptor modulates BDNF production in rat primary cortical neuron. CGS21680, an adenosine A2(A) receptor agonist, induced BDNF expression and release. An antagonist against A2(A) receptor, ZM241385, prevented CGS21680-induced increase in BDNF production. A2(A) receptor stimulation induced the activation of Akt-GSK-3β signaling pathway and the blockade of the signaling pathway with specific inhibitors abolished the increase in BDNF production, possibly via modulation of ERK1/2-CREB pathway. The physiological roles of A2(A) receptor-induced BDNF production was demonstrated by the protection of neurons from the excitotoxicity and increased neurite extension as well as synapse formation from immature and mature neurons. Taken together, activation of A2(A) receptor regulates BDNF production in rat cortical neuron, which provides neuro-protective action.

BACKGROUND:At the beginning of neurogenesis, massive brain cell death occurs and more than 50% of cells are eliminated by apoptosis along with neuronal differentiation. However, few studies were conducted so far regarding the regulation of neural progenitor cells (NPCs) death during development. Because of the physiological role of cell death during development, aberration of normal apoptotic cell death is detrimental to normal organogenesis.Apoptosis occurs in not only neuron but also in NPCs and neuroblast. When growth and survival signals such as EGF or LIF are removed, apoptosis is activated as well as the induction of differentiation. To investigate the regulation of cell death during developmental stage, it is essential to investigate the regulation of apoptosis of NPCs.METHODS:Neural progenitor cells were cultured from E14 embryonic brains of Sprague-Dawley rats. For in vivo VPA animal model, pregnant rats were treated with VPA (400 mg/kg S.C.) diluted with normal saline at E12. To analyze the cell death, we performed PI staining and PARP and caspase-3 cleavage assay. Expression level of proteins was investigated by Western blot and immunocytochemical assays. The level of mRNA expression was investigated by RT-PCR. Interaction of Bcl-XL gene promoter and NF-κB p65 was investigated by ChIP assay.RESULTS:In this study, FACS analysis, PI staining and PARP and caspase-3 cleavage assay showed that VPA protects cultured NPCs from cell death after growth factor withdrawal both in basal and staurosporine- or hydrogen peroxide-stimulated conditions. The protective effect of prenatally injected VPA was also observed in E16 embryonic brain. Treatment of VPA decreased the level of IκBα and increased the nuclear translocation of NF-κB, which subsequently enhanced expression of anti-apoptotic protein Bcl-XL.CONCLUSION:To the best of our knowledge, this is the first report to indicate the reduced death of NPCs by VPA at developmentally critical periods through the degradation of IκBα and the activation of NF-κB signaling. The reduced NPCs death might underlie the neurodevelopmental defects collectively called fetal valproate syndrome, which shows symptoms such as mental retardation and autism-like behavior.

BACKGROUND:Fragile X syndrome (FXS), the most commonly inherited mental retardation and single gene cause of autistic spectrum disorder, occurs when the Fmr1 gene is mutated. The product of Fmr1, fragile X linked mental retardation protein (FMRP) is widely expressed in HeLa cells, however the roles of FMRP within HeLa cells were not elucidated, yet. Interacting with a diverse range of mRNAs related to cellular survival regulatory signals, understanding the functions of FMRP in cellular context would provide better insights into the role of this interesting protein in FXS. Using HeLa cells treated with etoposide as a model, we tried to determine whether FMRP could play a role in cell survival.METHODS:Apoptotic cell death was induced by etoposide treatment on Hela cells. After we transiently modulated FMRP expression (silencing or enhancing) by using molecular biotechnological methods such as small hairpin RNA virus-induced knock down and overexpression using transfection with FMRP expression vectors, cellular viability was measured using propidium iodide staining, TUNEL staining, and FACS analysis along with the level of activation of PI3K-Akt pathway by Western blot. Expression level of FMRP and apoptotic regulator BcL-xL was analyzed by Western blot, RT-PCR and immunocytochemistry.RESULTS:An increased FMRP expression was measured in etoposide-treated HeLa cells, which was induced by PI3K-Akt activation. Without FMRP expression, cellular defence mechanism via PI3K-Akt-Bcl-xL was weakened and resulted in an augmented cell death by etoposide. In addition, FMRP over-expression lead to the activation of PI3K-Akt signalling pathway as well as increased FMRP and BcL-xL expression, which culminates with the increased cell survival in etoposide-treated HeLa cells.CONCLUSIONS:Taken together, these results suggest that FMRP expression is an essential part of cellular survival mechanisms through the modulation of PI3K, Akt, and Bcl-xL signal pathways.

We screened the major bioactive flavones isolated from Scutellaria baicalensis (baicalin, baicalein and oroxylin A) for their convulsion related activities. In electrogenic response score system and the pentylenetetrazole seizure model, baicalein but not oroxylin A and baicalin exhibited anticonvulsant effects. In vitro studies also revealed that baicalein induced intracellular Cl(-) influx, whereas oroxylin A blocked muscimol- and baicalein-induced intracellular Cl(-) influx. The anticonvulsant effect of baicalein was inhibited by flumazenil, a benzodiazepine(BZD) receptor antagonist. Therefore, anticonvulsive effect of baicalein was mediated by the BZD binding site of GABA(A) receptor. The 5, 7-dihydroxyl group is present in the structure of the three flavones. It is postulated that this group played a key role in inducing convulsion-related activities.

Forsythiaside, a phenylethanoside, has been reported to have anti-oxidative activity and memory ameliorating effects against a scopolamine-induced memory deficit model. The aim of this study was to determine whether forsythiaside has neuroprotective activity on transient cerebral global ischemia in gerbil. Transient cerebral ischemia was induced by bilateral common carotid artery occlusion for 5 min and followed by reperfusion for 7 days. Oral administration of forsythiaside was conducted immediately after reperfusion and once a day over the next 7 days. The forsythiaside administration significantly increased the number of viable neurons detected by neuronal nuclei immunostaining and decreased degenerating neuronal cells detected by Fluoro-Jade B staining in the hippocampal CA1 region, at the 7th day post-ischemia (P<0.05). Forsythiaside also significantly decreased the number of ionized calcium-binding adaptor molecule-1-detected activated microglia and glial fibrillary acidic protein-detected astrocytes, both of which were increased after ischemic insults, and decreased interleukin-1β and tumor necrosis factor-α expression levels, which were also increased after the insults (P<0.05). In addition, forsythiaside significantly improved ischemia-induced cognitive impairments in the Y-maze task (P<0.05). These results suggest that forsythiaside exhibits neuroprotective properties, which are, in part, mediated by its anti-inflammatory activities supported by forsythiaside-induced reductions of activated glial cells and expression levels of interleukin-1β and tumor necrosis factor-α.

Prenatal exposure to valproic acid (VPA) induces neural tube defects and impairment in social behaviors related to autistic spectrum disorder in newborns, which make it a useful animal model of autism. In this study, we compared the effects of different time window of prenatal valproic acid exposure for inducing the altered social behaviors relevant to autism from embryonic day 7 to embryonic day 15 in Sprague–Dawley rats to determine the critical periods for the impairment. Compared to E7, E9.5 and E15 exposure, VPA exposure at E12 showed most significant changes in behaviors over control animals with reduced sociability and social preference. E9.5 exposure to valproic acid showed strong reproductive toxicity including decrease in the number of live birth. In general, exposure at E15 showed only marginal effects on reproduction and social behaviors. Finally, VPA-exposed rats at E12 were more sensitive to electric shock than VPA-exposed rats at any other periods. These results suggested that E12 is the critical period in rats when valproate exposure has prominent effects for inducing the altered social behavior similar to human autistic behavior.

Astrocytes are the most abundant cells in the brain, playing vital roles in neuronal survival, growth, and function. Understanding the mechanism(s) regulating astrocyte proliferation will have important implications in brain development, response to injury, and tumorigenesis. Cyclin B1 is well known to be a critical regulator of mitotic entry via its interaction with cyclin-dependent kinase 1. In rat astrocytes, we now show that the mRNA binding protein cytoplasmic polyadenylation element binding protein 1 (CPEB1) is associated with cyclin B1 mRNA and that this interaction is enriched at the centrosome. In addition, if growth-arrested astrocytes are stimulated to divide, CPEB1 is phosphorylated and cyclin B1 mRNA is polyadenylated, both hallmarks of CPEB1 activation, resulting in an increase in cyclin B1 protein. CPEB1 binding to mRNA initially inhibits translation; therefore, removing CPEB1 from mRNA should result in an increase in translation due to derepression. Indeed, when we either knocked down CPEB1 protein with siRNA or sequestered it from endogenous mRNA by expressing RNA containing multiple CPEB1 binding sites, cyclin B1 protein was increased and cell proliferation was stimulated. Our data suggest a mechanism wherein CPEB1 is bound and represses cyclin B1 mRNA translation until a signal to proliferate phosphorylates CPEB1, resulting in an increase in cyclin B1 protein and progression into mitosis. Our results demonstrate for the first time a role for CPEB1 in regulating cell proliferation in the brain.

Oxidative stress resulting from excessive free-radical release is likely implicated in the initiation and progression of epilepsy. Therefore, antioxidant therapies aimed at reducing oxidative stress have received considerable attention in epilepsy treatment. However, much evidence suggests that oxidative stress does not always have the same pattern in all seizures models. Thus, this review provides an overview aimed at achieving a better understanding of this issue. We summarize work regarding seizure models (i.e., genetic rat models, kainic acid, pilocarpine, pentylenetetrazol, and trimethyltin), oxidative stress as an etiologic factor in epileptic seizures (i.e., impairment of antioxidant systems, mitochondrial dysfunction, involvement of redox-active metals, arachidonic acid pathway activation, and aging), and antioxidant strategies for seizure treatment. Combined, this review highlights pharmacological mechanisms associated with oxidative stress in epileptic seizures and the potential for neuroprotection in epilepsy that targets oxidative stress and is supported by effective antioxidant treatment.

In this study, the effect of daidzin or daidzein isolated from Pueraria lobata on the memory impairments induced by scopolamine was assessed in male mice using the passive avoidance and the Morris water maze tasks. Administration of daidzin (5 mg/kg) or daidzein (5 mg/kg) significantly reversed the scopolamine (1 mg/kg)-induced cognitive impairments in male mice as evidenced by the passive avoidance test (p < 0.05) and on the Morris water maze test (p < 0.05). Moreover, the ameliorating effects of daidzin or daidzein were antagonized by tamoxifen (1 mg/kg), the nonspecific estrogen receptor antagonist. These results indicate that daidzin or daidzein may be useful in cognitive impairment induced by cholinergic dysfunction, and this beneficial effect is mediated, in part, via estrogen receptor.

Neural progenitor cells (NPCs) differentiate into astrocytes, neurons and oligodendrocytes, which is controlled by various factors in brain. Recent evidences suggest that small molecules modulating the proliferation and differentiation of NPCs may have therapeutic value as well as the potential use as chemical probes. Phylligenin is a lignan with anti-inflammatory activity that is isolated from the fruits of Forsythia koreana. We investigated effects of phylligenin on proliferation and differentiation of NPCs. Treatment of phylligenin decreased the number of proliferating NPCs in culture without effects on the differentiation and survival of neural cells such as neurons and astrocytes. To examine the mechanism of the decreased NPCs number, we performed cell cycle analysis. Proliferation of NPCs was decreased via G1-S transition block by phylligenin treatment, and it was mediated by the increase of p21 level. However, phylligenin did not induce apoptosis of NPCs as determined by TUNEL assay and PARP cleavage. We also found that viability of glioma cell lines such as C6 and U87MG glioma cells, but not that of primary neuron and astrocyte, was inhibited by phylligenin. These results suggest that phylligenin selectively inhibits proliferation of rapidly growing cells such as neural stem cells and glioma cells. Given that the possible role of brain tumor stem cells in the pathology of brain cancers, the inhibitory effects of phylligenin might be useful in the development of new therapeutic agents against brain cancers.

기본설계 단계에서 회전익 항공기의 가격을 보다 정확하게 예측하기 위해서 Harris & Scully에 의해 제시된 가격 예측 식을 최신의 추세식으로 수정하였다. 이를 위해 1997년 이후의 총 74개에 대한 회전익기 가격 및 제원 정보를 이용하여 가격 예측 식을 업데이트하였다. 업데이트 된 가격 예측식을 Tishchenko의 기본설계 기법과 통합하여 기본설계 단계에서 가격을 고려하도록 구성하였다. 본 기본설계 및 가격예측 프로그램을 현재 운용되고 있는 미국 Sikorsky사의 S-76C+를 비롯한 여러 기종의 헬리콥터에 적용하여 가격과 제원 결과를 도출하고 검증하여 본 연구의 유효성을 확인하였다. In this work, the helicopter price estimation formula from Harris & Scully is revised and updated for more accurate prediction of the current prices of helicopters which can be applied in the preliminary design stage. As many as 74 helicopter price and sizing data after the year 1997 are newly included to the out-dated coefficients in the cost estimation formula. In addition, the updated price estimation formula is added to Tishchenko's preliminary sizing method for price consideration in the preliminary design stage. Sikorsky's S-76C+ helicopter is used along with other various helicopters for comparison and validation of the sizing code as well as the new updated price estimation formula.

Brain-derived neurotrophic factor (BDNF) is a neurotrophic factor involved in neuronal differentiation, plasticity, survival and regeneration. BDNF draws massive attention mainly due to the potential as a therapeutic target in neurological diseases such as depression and Alzheimer's disease. In a primary screening for the natural compounds enhancing BDNF release from cultured rat primary cortical neuron, we found that compounds such as baicalein, tanshinone lla, cinnamic acid, epiberberine, genistein and wogonin among many others increased BDNF release. All the compounds at 0.1 mu M of concentration barely showed stimulatory effect on BDNF induction, however, their combination (mixture 1; baicalein, tanshinone lla and cinnamic acid, mixture 2; epiberberine, genistein and wogonin) showed synergistic increase in BDNF release as well as mRNA and protein expression. The level of BDNF expression was comparable to the maximum BDNF stimulation attainable by a positive control oroxylin A (20 mu M) without cell toxicity as determined by MTT analysis. Both mixtures synergistically increased the phosphorylation of extracellular signal-regulated kinase (ERK) as well as cAMP response element binding protein (CREB), an immediate and essential regulator of BDNF expression. Similar to these results, mixture of these compounds synergistically inhibited the up-regulation of inducible nitric oxide synthase (iNOS) induced by lipopolysaccharide treatments in rat primary astrocytes. These results suggest that the combinatorial treatment of natural compounds in lower concentration might be a useful strategy to obtain sufficient BDNF stimulation in neurological disease condition such as depression, while minimizing potential side effects and toxicity of higher concentration of a single compound.

Background Prenatal ethanol exposure during pregnancy induces a spectrum of mental and physical disorders called fetal alcohol spectrum disorder (FASD). The central nervous system is the main organ influenced by FASD, and neurological symptoms include mental retardation, learning abnormalities, hyperactivity and seizure susceptibility in childhood along with the microcephaly. In this study, we examined whether ethanol exposure adversely affects the proliferation of NPC and de-regulates the normal ratio between glutamatergic and GABAergic neuronal differentiation using primary neural progenitor culture (NPC) and in vivo FASD models. Methods Neural progenitor cells were cultured from E14 embryo brain of Sprague-Dawley rat. Pregnant mice and rats were treated with ethanol (2 or 4 g/kg/day) diluted with normal saline from E7 to E16 for in vivo FASD animal models. Expression level of proteins was investigated by western blot analysis and immunocytochemical assays. MTT was used for cell viability. Proliferative activity of NPCs was identified by BrdU incorporation, immunocytochemistry and FACS analysis. Results Reduced proliferation of NPCs by ethanol was demonstrated using BrdU incorporation, immunocytochemistry and FACS analysis. In addition, ethanol induced the imbalance between glutamatergic and GABAergic neuronal differentiation via transient increase in the expression of Pax6, Ngn2 and NeuroD with concomitant decrease in the expression of Mash1. Similar pattern of expression of those transcription factors was observed using an in vivo model of FASD as well as the increased expression of PSD-95 and decreased expression of GAD67. Conclusions These results suggest that ethanol induces hyper-differentiation of glutamatergic neuron through Pax6 pathway, which may underlie the hyper-excitability phenotype such as hyperactivity or seizure susceptibility in FASD patients.