• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1996년도 춘계학술대회
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• pp.175-175
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• 1996

The spinal dorsal horn is the area where primary afferent fibers terminate and cutaneous sensory information is Processed. A number of putative neurotransmitter substances, including excitatory and inhibitory amino acids and peptides, are present in this region and sites and cellular mechanisms of their actions have been a target of numerous studies. In this study, single neurons were acutely isolated and the properties of whole cell current and responses to excitatory and inhibitory neurotransmitters were studied by the patch clamp method. Young rats (7-14 days) were anesthetized with diethyl-ether, and the lumbar spinal cord was excised and cut transversely at a thickness of 30$\mu\textrm{m}$ by Vibroslicer. The treatment of spinal slices with low concentration of proteases (pronase and thermolysin 0.75 mg/$m\ell$) and mechanical dissociation yielded isolated neurons with near intact morphology. Multipolar, ellipsoidal and bipolar, and pyramidal cells were shown. By applying step voltage pulses to neurons held at -70 mV, two types of inward currents and one outward currents observed. The fast activating and inactivating inward current was the Na$\^$+/ current because of its fast kinetics and blocking by 0.5${\mu}$M TTX, a specific blocker of Na$\^$+/ channel. The second type of inward currents were sustained. Based on their kinetics and current-voltage relations, it was likely that the second type of inward current was the voltage-dependent Ca$\^$2+/ current. In the presence of TTX, the steady-state currents mainly represented outward K$\^$+/ current which looked like the delayed rectifier K$\^$+/ current. In addition, the membrane currents produced by agonist of excitatory amino acid (EAA) receptor and the endogenous transmitter candidate L-glutamate were recorded in isolated whole-cell voltage clamped neurons as well as responses to inhibitory amino acids (${\gamma}$-amino butyric acid, glycine). Drugs were applied by a method that allows complete exchange of the solution within 1 sec; an infinite number of solutions can be applied to a single cell.

• Archives of Pharmacal Research
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• 제24권2호
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• pp.164-170
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• 2001

Glutamate receptors-mediated excitoxicity is believed to play a role in the pathophysiology of neurodegenerative diseases. The present study was performed to evaluate the inhibitory effect of fanschinoline, a bis-benzylisoquinoline alkaloid, which has a characteristic as a $Ca^{2+}$channel blockers on excitatory amino acids (EAAS)-induced neurotoxicity in cultured rat cerebellar granule neuron. Fangchinoline (1 and 5$\mu\textrm{m}$) inhibited glutamate (1 ${m}M$), N-methyl-D-aspartate (NMDA; 1 ${m}M$) and kainate (100$\mu\textrm{m}$)-induced neuronal cell death which was measured by trypan blue exclusion test. Fangchinoline (1 and 5$\mu\textrm{m}$) inhibited glutamate release into medium induced by NMDA (1 ${m}M$) and kainate (100$\mu\textrm{m}$), which was measured by HPLC. And fangchinoline (5$\mu\textrm{m}$) inhibited glutamate (1 ${m}M$)-induced elevation of intracellular calcium concentration. These results suggest that inhibition of $Ca^{2+}$influx by fangchinoline may contribute to the beneficial effects on neurodegenerative effect of glutamate in pathophysiological conditions.

• 대한약리학회지
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• 제32권2호
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• pp.283-292
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• 1996

The spinal dorsal horn is the area where primary afferent fibers terminate and cutaneous sensory information is processed. A number of putative neurotransmitter substances, including excitatory and inhibitory amino acids and peptides, are present in this region. In this study, single neurons of the spinal dorsal horn were acutely isolated and the properties of whole cell current and responses to excitatory and inhibitory neurotransmitters were studied by patch clamp technique. Transverse slice ($(300{\mu}m$) of lumbar spinal cords from young rats$(7{\sim}14\;days)$ were sequentially treated with two pretenses(pronase 0.75 mg/ml and thermolysin 0.75 mg/ml), then single neurons were mechanically dissociated. These neurons showed near-intact morphology such as multipolar, ellipsoidal and bipolar, and pyramidal cells and we recorded the typical whole cell currents of $K^+$, $Ca^{2+}$ and ligand-operated channels from these neurons. Glutamate $(30{\mu}M)$ and N-methyl-D-aspartate(NMDA, $30{\mu}M)$ induced inward currents of $117{\pm}12.4$ pA(n=5) and $49{\pm}6.9$ pA(n=3), respectively. Glycine $(1{\mu}M)$ potentiated glutamate-induced currents $4{\sim}5$ times and NMDA-induced currents $8{\sim}10$ times. In addition, glycine $(30{\mu}M)$ induced Inward current ($31{\pm}6.1$ nA, n=2), which was rapidly desensitized after the peak to a new steady-state level. However, the inward currents induced by ${\gamma}-amino$ butyric acid(GABA, $1{\mu}M$) decreased continuously after the peak($226{\pm}41.6$ pA, n=3) under the similar experimental condition. The ionic currents and pharmacological responses of isolated neurons in this work were similar to those observed in vivo or in vitro spinal cord slice, indicating that acutely isolated neurons could be effectively used for further pharmacological studies.

• 생명과학회지
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• 제21권1호
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• pp.102-109
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• 2011

턱끝혀근은 호흡을 위해 상부기도를 확보하는 중요한 근육이지만 정상적으로 수축하지 못하면 상부기도를 폐쇄함으로써 호흡장애가 생길 수있다. 이것은 턱끝혀근을 지배하는 운동신경원의 연접입력 이상으로 생각하고 있으나 이러한 연접입력에 대해서는 잘 알려져 있지 않다. 본 연구에서는 흰쥐의 턱끝혀근을 지배하는 운동신경원과 연접하는 억제성 및 흥분성 신경종말의 분포 양식에 대해 분석하고자 하였으며, 이를 위해 신경추적자인 HRP를 주입하여 턱끝혀근 지배 운동신경원을 표식하고, 운동신경원을 세포체와 가지돌기의 근위부, 중간부 및 원위부로 구분한 후, 전자현미경용 연속절편을 제작하여 GABA, glycine 및 glutamate 항체를 사용한 postembedding immunogold histochemistry를 시행하였다. 정량적 분석은 3개의 턱끝혀근 운동신경원에 연접한 622개의 신경종말 중 157개는 세포체에서, 188개는 14개의 가지돌기 근위부에서, 181개는 35개의 가지돌기 중간부에서, 96개는 28개의 가지돌기 원위부에서 각각 연접 양상을 분석하였다. 관찰한 신경종말의 71.9%에서 사용된 3종류의 아미노산에 대한 면역양성반응이 나타났는데 이 중 32.8%는 GABA 또는 glycine에 대한 면역양성반응을 보였고, 39.1%는 glutamate에 대한 면역양성반응을 보였다. GABA 또는 glycine에 대한 면역양성 신경종말 중 14.2%는 glycine에만 면역양성반응을 보였고, 13.3%는 glycine과 GABA에 동시에 면역양성반응을 보였으며, 5.3%만이 GABA에만 면역양성반응을 보였다. 억제성 아미노산에 면역양성반응을 보인 신경종말에는 납작하거나, 타원형 또는 둥근 형태의 연접소포가 함유된 반면, 흥분성 아미노산에 면역양성반응을 보인 신경종말에는 구형의 소포와 약간의 큰 치밀연접소포가 함유되어 있었다. 억제성 신경종말과 흥분성 신경종말의 분포 비율은 가지돌기 원위부에서 각각 23.9% 대 43.8%로 가장 높았지만, 세포체(35.7% 대 38.2%), 가지돌기의 근위부(34.6% 대 37.8%), 및 중간부(33.1% 대 38.7%)에서는 큰 차이를 보이지 않았다. 억제성 신경종말의 synaptic covering (%)은 세포체에서 가지돌기 원위부로 갈수록 감소되었지만, 흥분성 신경종말의 synaptic covering (%)은 각 부위에서 큰 차이를 나타내지 않았다. 본 연구를 통하여 턱끝혀근 지배 운동신경원은 세포체와 가지돌기의 부위에 따라 GABA, glycine 및 glutamate를 함유하는 전운동신경원들에 의해 서로 다른 양식의 제어를 받고 있다는 것을 알 수 있었으며, 운동신경원의 부위에 따른 이러한 억제성 및 흥분성 연접입력 양식의 차이는 유연한 혀운동의 제어기전과 밀접한 관련이 있을 것으로 추정된다.

• The Korean Journal of Physiology and Pharmacology
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• 제3권3호
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• pp.237-244
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• 1999

Magnesium ion is known to selectively block the N-methyl-D-aspartate (NMDA)-induced responses and to have anticonvulsive action, neuroprotective effect and antinociceptive action in the behavioral test. In this study, we investigated the effect of $Mg^{2+}$ on the responses of dorsal horn neurons to cutaneous thermal stimulation and graded electrical stimulation of afferent nerves as well as to excitatory amino acids and also elucidated whether the actions of $Ca^{2+}$ and $Mg^{2+}$ are additive or antagonistic. $Mg^{2+}$ suppressed the thermal and C-fiber responses of wide dynamic range (WDR) cell without any effect on the A-fiber responses. When $Mg^{2+}$ was directly applied onto the spinal cord, its inhibitory effect was dependent on the concentration of $Mg^{2+}$ and duration of application. The NMDA- and kainate-induced responses of WDR cell were suppressed by $Mg^{2+}$, the NMDA-induced responses being inhibited more strongly. $Ca^{2+}$ also inhibited the NMDA-induced responses current-dependently. Both inhibitory actions of $Mg^{2+}$ and $Ca^{2+}$ were additive, while $Mg^{2+}$ suppressed the EGTA-induced augmentation of WDR cell responses to NMDA and C-fiber stimulation. Magnesium had dual effects on the spontaneous activities of WDR cell. These experimental findings suggest that $Mg^{2+}$ is implicated in the modulation of pain in the rat spinal cord by inhibiting the responses of WDR cell to noxious stimuli more strongly than innocuous stimuli.

• The Korean Journal of Physiology and Pharmacology
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• 제4권6호
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• pp.455-461
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• 2000

Zinc contained in the neurons of central nervous system is activity-dependently released and then attenuates NMDA (N-methyl-D-aspartate)-induced neurotoxicity while augmenting non-NMDA-induced neurodegeneration. Zinc also has been reported to produce antinociceptive action on the inflammation- and nerve injury-induced hyperalgesia in the behavioral test. In this study, we investigated the effects of zinc on the responses of dorsal horn cells to NMDA, kainate and graded electrical stimulation of C-fibers. In the majority of WDR cells (70.6%), zinc current-dependently inhibited WDR cell responses to NMDA and in the remaining cells, produced biphasic responses; excitation followed by inhibition. Zinc augmented the responses of WDR cells to iontophoretical application of kainate. The dominant effect of $Zn^{2+}$ on the responses of WDR cells to C-fiber stimulation was excitatory, but inhibition, excitation-inhibition and no change of the responses to C-fiber stimulation were induced. $Ca^{2+}-EDTA$ antagonized the excitatory or inhibitory effects of $Zn^{2+}$ on the WDR cell responses. These experimental findings suggest that $Zn^{2+}$ modulates the transmission of sensory information in the rat spinal cord.

• Biomolecules & Therapeutics
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• 제3권1호
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• pp.91-96
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• 1995

The effects of toluene inhalation on the contents of amino acid neurotransmitters in rat brain were investigated and blood toluene concentrations inducing changes of behavior and amino acid neurotransmitter contents in rat brain were observed. Male wistar rats were exposed to toluene vapor (single dose : 1700, 5000 and 10000 ppm for 2 hrs, repeated dose : 1700 and 5000 ppm for 2 hrs/day$\times$6 days). Toluene concentrations in blood and the inhalation chamber were assayed by GC with headspace sampler. HPLC method following PITC derivatization was used to measure the amino acid contents in brain tissues such as frontal cortex, caudate, hippocampus, cerebellum and brain stem. Glutamic acid and aspartic acid levels were increased by single inhalation of toluene (5000 ppm) in all the brain areas assayed in this experiment. In caudate and cerebellum, taurine levels were decreased by single inhalation of low dose toluene (1700 ppm), but increased by repeated administration. At high blood toluene concentration, GABA levels were increased in all the brain areas assayed in this experiment and the increasing extents of inhibitory amino acid contents measured in caudate and hippocampus were greater than those of excitatory amino acids. These results suggest that the changes of amino acid neurotransmitter contents in brain by exposure to toluene may modulate toluene-induced behaviors.

• 대한한방내과학회지
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• 제21권1호
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• pp.116-125
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• 2000

Objectives : The aim of this study is to investigate that Sunghyangjeongki-San which has been frequently medicated in the early stage of stroke can protect against ischemic damage in rat brain Methods : Extracellular levels of amino acids(glutamate, aspartate, GABA, glycine, taurine, tyrosine, alanine), organic acids(pyruvate, lactate), and cerebral infarction volume were measured at the striatum of rats subjected to permanant focal cerebral ischemia induced by 2 hours of middle cerebral artery occiusion(MCAO). Rats were orally administered with Sunghyangjeongki-San at 30mins before MCAO and the microdialysate was collected by intracerebral microdialysis three times before MCAO and six times after MCAO at 20mins interval and analyzed by HPLC. After a microdialysis study, the brain was sliced and stained with cresyl violet buffer for the measurement of cerebral infarcted area and volume by image analyzer system Results : The concentrations of glutamate, aspartate, and tyrosine known as excitatory neurotransmitters were significantly decreased in Sunghyangjeongki-San group compared with control group, The concentrations of GABA, glycine, taurine and alanine known as inhibitory neurotransmitters were significantly increased in Sunghyangjeongki-San group compared with control group. The concentrations of pyruvate and lactate showed little significant change in Sunghyangjeongki-San group compared with control group. The measurement of cerebral infarcted area and volume by image analyzer system were significantly decreased in Sunghyangjeongki-San group compared with control group. Conclusions : Sunghyangjeongki-San can affect on protecting against cerebral ischemia by regulating extracellular levels of excitatory and inhibitory amino acid neurotransmitters and improve the conditions of the patients in the early stage of stroke.

• Clinical and Experimental Pediatrics
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• 제49권7호
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• pp.718-725
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• 2006

The pubertal activation of gonadotrophin releasing hormone(GnRH) requires coordinated changes in excitatory or inhibitory amino acids, growth factors, and a group of transcriptional regulators. The age of onset of puberty is progressing to younger age. Factors affecting early puberty include genetic traits, nutrition(body fat) and exposure to endocrine disrupting chemicals. In rapidly progressing central precocious puberty, gonadotrophin releasing hormone(GnRH) agonists(GnRHa) appear to increase final height if treated early stage. Further large scaled long-term follow-up study of the effects of GnRHa on final height is needed.

• 한국버섯학회지
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• 제17권4호
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• pp.275-283
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• 2019

Sparassis latifolia is a fungus abundant in β-glucan and amino acids and is highly valued as a medicinal mushroom. Among amino acids, γ-aminobutyric acid (GABA) is a free amino acid and has biological effects, such as increase/decrease of hypertension, improvement of cerebral blood flow, and prevention of dementia. In this study, biological elicitors were used to increase bioactive substances as a biofortification method. Sodium alginate extracted from seaweed (Sargassum horneri, Sargassum fulvellum, Sargassum fusiforme) were used as the elicitor. The levels of β-glucan and GABA in the mycelium and fruiting body grown by adding the elicitor to the medium were investigated. Addition of sodium alginate positively affected GABA production and negatively affected the β-glucan production in these fungi. Sodium alginates extracted from S. fulvellum induced the highest increase in GABA in the mycelium and fruiting bodies. Moreover, we investigated the effects of the extracts from mycelium and fruiting bodies on dendrite development in primary cortical neurons. We found that the extract from the fruiting bodies of sodium alginate treated fungi with increased levels of GABA inhibited the dendrite outgrowth of excitatory neurons, but not inhibitory neurons.