• YAKHAK HOEJI
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• v.44 no.1
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• pp.29-35
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• 2000

The methanol extracts were prepared from 46 oriental medicines currently used for stroke treatment, and the effects were assessed on the excitotoxic neuronal cell death induced by L-glutamate(Glu) in primary cultured rat cortical neurons. The extracts from Angelicae gigantis Radix, Manitis Squama, Acori graminei Rhizoma, Uncariae Ramulus et Uncus, Alpiniae Fructus, Paeoniae Radix, and Cnidii Rhizoma inhibited the Glu-induced neurotoxicity with the IC$_50$ values of 95.2, 218.6, 263.3, 295.1, 297.9, 310.1, and 446.7 $\mu$g/ m$\ell$, respectively. The extracts from Arisaematis Rhizoma, Loranthi Ramulus, Anemarrhenae Rhizoma, Carthami Flos, Clematidis Radix, Bambusae Concretio Silicea, and Angelicae koreanae Radix also exhibited significant inhibition of the toxicity. In contrast, the extracts from Aconiti Tuber Araliae cordatae Radix, Curcumae Rhizoma, Leonuri Herba, Polygalae Radix, Salviae Radix, and Siegesbeckiae Herba increased the Glu-induced toxicity at the concentrations of 500 and 1000 $\mu$g/m$\ell$. Rest of the extracts evaluated in the present study showed minor or negligible inhibition. liken together the oriental medicines including Angelicae gigantis Radix, Muitis Squama, Acori graminei Rhizoma, Uncariae Ramulus et Uncus, and Alpiniae Fructus appear to exert pharmacological effects through the inhibition of excitotoxic neuronal cell death. Further studies are in progress to characterize active principles in these extracts.

• YAKHAK HOEJI
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• v.45 no.4
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• pp.419-425
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• 2001

The neuronal cell death induced by excess glutamate (Glu) has been implicated in many acute and chronic neurodegenerative diseases including cerebral ischemia. Glu-induced elevation of intra-cellular $Ca^{2+}$ plays a critical role in the excitotoxicity, partly through the activation of a variety of $Ca^{2+}$ dependent enzymes. In the present study, we investigated the Glu-induced modulation of $Ca^{2+}$/calmodulin-dependent protein kinase IV (CaMK IV), a multifunctional enzyme abundantly present in the nuclei of neurons. The exposure of cultured rat cortical neurons to $100{\mu}$M Glu for 3 min dramatically increased CaMK IV activity up to 4.5-fold of the control-treated enzyme activity. The activation was very rapid, reaching peak at 3 min and then declined gradually. Under the same experimental conditions, time-dependent acute and delayed neuronal cell death was observed. Immunoblot analyses using specific antibodies showed that the expressions of CaMK IV and $CaMKK_{\alpha}$ were time-dependently modulated by Glu. Taken together, these results imply that the modulation of CaMK IV activity by Glu may be involved in the cascade of events resulting in neuronal cell death in cortical cultures.

• Proceedings of the PSK Conference
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• 2000.04a
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• pp.172.1-172.1
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• 2000

• The Korean Journal of Physiology and Pharmacology
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• v.23 no.6
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• pp.483-491
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• 2019

Cordycepin exerts neuroprotective effects against excitotoxic neuronal death. However, its direct electrophysiological evidence in Alzheimer's disease (AD) remains unclear. This study aimed to explore the electrophysiological mechanisms underlying the protective effect of cordycepin against the excitotoxic neuronal insult in AD using whole-cell patch clamp techniques. ${\beta}$-Amyloid ($A{\beta}$) and ibotenic acid (IBO)-induced injury model in cultured hippocampal neurons was used for the purpose. The results revealed that cordycepin significantly delayed $A{\beta}$ + IBO-induced excessive neuronal membrane depolarization. It increased the onset time/latency, extended the duration, and reduced the slope in both slow and rapid depolarization. Additionally, cordycepin reversed the neuronal hyperactivity in $A{\beta}$ + IBO-induced evoked action potential (AP) firing, including increase in repetitive firing frequency, shortening of evoked AP latency, decrease in the amplitude of fast afterhyperpolarization, and increase in membrane depolarization. Further, the suppressive effect of cordycepin against $A{\beta}$ + IBO-induced excessive neuronal membrane depolarization and neuronal hyperactivity was blocked by DPCPX (8-cyclopentyl-1,3-dipropylxanthine, an adenosine $A_1$ receptor-specific blocker). Collectively, these results revealed the suppressive effect of cordycepin against the $A{\beta}$ + IBO-induced excitotoxic neuronal insult by attenuating excessive neuronal activity and membrane depolarization, and the mechanism through the activation of $A_1R$ is strongly recommended, thus highlighting the therapeutic potential of cordycepin in AD.

• International Journal of Oral Biology
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• v.34 no.2
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• pp.97-103
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• 2009

L-trans-pyrrolidine-2,4-dicarboxylate (PDC) is a potent inhibitor of glutamate transporters. In our current study, we investigated whether the neuronal death induced by PDC involves mechanisms other than excitotoxicity in mixed mouse cortical cultures. Cortical cultures at 13-14 days in vitro were used and cell death was assessed by measuring the lactate dehydrogenase efflux into bathing media. Glutamate and PDC both induced neuronal death in a concentration-dependent manner but the neurotoxic effects of glutamate were found to be more potent than those of PDC. Treatment with 10, 100 and 200 ${\mu}$M PDC equally potentiated 50 ${\mu}$M glutamate-induced neuronal death. The neuronal death induced by 75 ${\mu}$M glutamate was almost abolished by treatment with the NMDA antagonists, MK-801 and AP-5, but was unaffected by NBQX (an AMPA antagonist), trolox (antioxidant), BDNF or ZVAD-FMK (a pan-caspase inhibitor). However, the neuronal death induced by 200 ${\mu}$M PDC was partially but significantly attenuated by single treatments with MK-801, AP-5, trolox, BDNF or ZVAD-FMK but not NBQX. Combined treatments with MK-801 plus trolox, MK-801 plus ZVAD-FMK or MK-801 plus BDNF almost abolished neuronal death, whereas combined treatments with trolox plus ZVADFMK, trolox plus BDNF or ZVAD-FMK plus BDNF did not enhance the inhibitory action of any single treatment with these drugs. These results demonstrate that the neuronal death induced by PDC involves not only in the excitotoxicity induced by the accumulation of glutamate but also the oxidative stress induced by free radical generation. This suggests that apoptotic neuronal death plays a role in PDCinduced oxidative neuronal injury.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.6
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• pp.699-705
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• 1997

Although it is known that neuronal cell death during development occurs by apoptosis, the mechanisms underlying excitatory amino acid-induced neuronal cell death remain poorly understood. In this study we have examined the mechanism by which L-glutamate, an excitatory amino acid neurotransmitter, induces cell death in PC12 cell lines. To characterize cell death, we employed sandwich enzyme-linked immunosorbent assay(ELISA) method for cellular DNA fragmentation, DNA agarose gel electrophoresis and chromatin staining by acridine orange and ethidium bromide after treating the PC12 cells with L-glutamate. L-Glutamate caused dose-dependent cell death with a maximum at 24 hrs after the treatment. These cellular fragmentation was blocked by pretreatment of MK-801, a noncompetitive N-methyl-D-aspartic acid(NMDA) receptor antagonist, and nerve growth factor(NGF). Analysis of DNA integrity from L-glutamate-treated cells revealed cleavage of DNA into regular sized fragments, a biochemical hallmark of apoptosis. The PC12 cells that were induced to die by L-glutamate treatment exhibited classical chromatin condensation under the light microscopy after acridine orange and ethidium bromide staining. These results suggest that apoptosis is one of the key features that are involved in L-glutamate-induced excitotoxic cell death in PC12 cells, and these cell death are mediated by NMDA receptor and depend on NGF.

• Journal of Ginseng Research
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• v.47 no.3
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• pp.390-399
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• 2023

Background: Gintonin (GT), a Panax ginseng-derived lysophosphatidic acid receptor (LPAR) ligand, has positive effects in cultured or animal models for Parkinson's disease, Huntington's disease, and so on. However, the potential therapeutic value of GT in treating epilepsy has not yet been reported. Methods: Effects of GT on epileptic seizure (seizure) in kainic acid [KA, 55mg/kg, intraperitoneal (i.p.)]-induced model of mice, excitotoxic (hippocampal) cell death in KA [0.2 ㎍, intracerebroventricular (i.c.v.)]-induced model of mice, and levels of proinflammatory mediators in lipopolysaccharide (LPS)-induced BV2 cells were investigated. Results: An i.p. injection of KA into mice produced typical seizure. However, it was significantly alleviated by oral administration of GT in a dose-dependent manner. An i.c.v. injection of KA produced typical hippocampal cell death, whereas it was significantly ameliorated by administration of GT, which was related to reduced levels of neuroglial (microglia and astrocyte) activation and proinflammatory cytokines/enzymes expression as well as increased level of the Nrf2-antioxidant response via the upregulation of LPAR 1/3 in the hippocampus. However, these positive effects of GT were neutralized by an i.p. injection of Ki16425, an antagonist of LPA1-3. GT also reduced protein expression level of inducible nitric-oxide synthase, a representative proinflammatory enzyme, in LPS-induced BV2 cells. Treatment with conditioned medium clearly reduced cultured HT-22 cell death. Conclusion: Taken together, these results suggest that GT may suppress KA-induced seizures and excitotoxic events in the hippocampus through its anti-inflammatory and antioxidant activities by activating LPA signaling. Thus, GT has a therapeutic potential to treat epilepsy.

• Development and Reproduction
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• v.12 no.1
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• pp.1-8
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• 2008

Specific protocols to increase the differentiation of neuronal cells from embryonic stem (ES) cells have been well established, such as retinoic acid induction and lineage selection of neuronal cells. For the neuropathological studies, ES-derived neurons (ES neurons) must show normal physiological characteristics related to cell death and survival and should be maintained in vitro for a sufficient time to show insults-specific cell death without spontaneous death. When mouse ES cells were plated onto astrocytes monolayer after retinoic acid induction, most ES cells differentiated into neuronal cells, which were confirmed by the presence of specific neuronal markers, and the cultures were viable for at least four weeks. When these cultures were examined for vulnerability to glutamate excitotoxicity, ES neurons were vulnerable to excitotoxic insults mediated by agonist-specific receptors. The vulnerability to excitotoxic death increased with developmental age of ES neurons in vitro. Specific receptors for Neurotrophin and GDNF family ligands were present in ES neurons. GDNF and NT-3 could modulate the survival and excitotoxic vulnerability of ES neurons. The vulnerability and resistance to toxic insults, which are essential requirements of model culture systems for neuropathological studies, make ES neurons to a useful model culture system. Especially ES cell are highly amenable to genetic modification unlikely to primary neuronal cells, which will give us a chance to answer more complicated neurophysiological questions. Recently there was an outstanding attempt to explore the cellular toxicity using human ES cells (Schrattenholz & Klemm, 2007) and it suggested that ES cells could be a new model system for neurophysiological studies soon and go further a large-scale screening system for pharmacological compounds in the future.

• The Korean Journal of Physiology and Pharmacology
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• v.7 no.6
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• pp.307-310
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• 2003

Kainic acid (KA) is a structural analogue of glutamate that interacts with specific presynaptic and postsynaptic receptors to potentiate the release and excitatory actions of glutamate. Systemic or intracerebroventricular (i.c.v.) administration of KA to experimental animals elicits multifocal seizures with a predominantly limbic localization, and results in neuronal death of cornu ammonia 1 (CA1), reactive gliosis and biochemical changes in the hippocampus and other limbic structures. Several lines of evidence suggest that reactive oxygen species (ROS) play a pivotal role in the pathogenesis of excitotoxic death by KA. Curcumin has been known to possess anti-oxidative and anti-inflammatory activities. In this study, the effects of curcumin on KA induced hippocampal cell death, reactive gliosis and biochemical changes in reactive glia were investigated by immunohistochemical methods. Our data demonstrated that curcumin attenuated KA-induced astroglial and microglial activation although it did not protect KA-induced hippocampal cell death.

• The Journal of Korean Medicine
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• v.26 no.3 s.63
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• pp.135-145
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• 2005

Objectives : This study investigated effect of Evodiae fructus water extract (EVOR) on apoptotic cell death induced by NaCN in SK-N-SH neuroblastoma cell lines. NaCN stimulates glutamate release which can activate glutamate receptors to initiate excitotoxic processes. This study examines the role of EVOR in mediating NaCN-induced cytotoxicity. Methods & Results : Cytotoxicity was assessed by measuring lactate dehydrogenase (LDH) in the culture media. NaCN(0.1mM) produced cytotoxicity following 12hrs of incubation. NaCN-induced cytotoxicity was partially blocked by EVOR. The treatment of EVOR in simultaneous exposure of cultures to NaCN provided complete protection against cytotoxicity. NaCN-induced cytotoxicity was found to inhibit DNA fragmentation, repaired by cell cycle and simultaneous exposure to NaCN, regenerated with neurite outgrowh by EVOR. These results indicate thaf damage by NaCN in neumnal cell cultures was repaired by EVOR, whereas NaCN-induced cytotoxicity is blocked Primarily by activation of anti-apoptosis. Conclusions : These results suggest that EVOR may be beneficial for the treatment of dementia and other degenerative problems of the central nervous system.