• 고려인삼학회:학술대회논문집
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• 고려인삼학회 2002년도 학술대회지
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• pp.401-415
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• 2002

Lim and his coworkers (1987; 1988; 1989) have also found that all of total Ginseng saponin, panaxadiol-and panaxatriol-type saponins cause the increased secretion of catecholamines (CA) in a $Ca^{2+}$ -dependent fashion from the isolated perfused rabbit adrenal glands through the activation of cholinergic (both nicotinic and muscarinic) receptors. These CA secretory effects are partly due to the direct action on the rabbit adrenomedullary chromaffin cells. However, the present study was designed to examine the effect of total ginseng saponin on CA secretion evoked by activation of cholinergic nicotinic receptors in the isolated perfused model of the rat adrenal gland. Total ginseng saponin given (100 ${\mu}g$/20 min) into an adrenal vein did fail to produce alteration of spontaneous CA release from the rat adrenal medulla. Acetylcholine(5.32 mM)- and DMPP(100 ${\mu}M$, a selective nicotinic receptor agonist)-evoked CA secretory responses were reduced markedly after the pretreatment with the total ginseng saponin at a rate of 100 ${\mu}g$/6.2 ml/20 min, respectively. Pretreatment with total ginseng saponin also depressed greatly high potassium (56 mM, a membrane depolarizing agent)- and Bay-K-8644 (10 ${\mu}M$, a calcium channel activator)-induced CA secretions. Taken together, it is thought that total ginseng saponin can inhibit the releasing effect of CA evoked by nicotinic receptor stimulation from the isolated perfused rat adrenal medulla, which seems to be associated to the direct inhibition of influx through L-type calcium channel into the rat adrenomedullary chromaffin cells. It seems that there is species differences in the adrenomedullary catecholamine secretion between the rabbit and rat.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 2002년도 학술대회지
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• pp.392-400
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• 2002

We have previously found that the saponins but not other components in the ginseng reduce the secretion of catecholamines (CAs) from bovine adrenal chromaffin cells, a model of sympathetic nerves, evoked by acetylcholine (ACh) due to the blockade of $Na^+$ influx through nicotinic ACh receptor-operated cation channels, and it has been concluded that the inhibitory effect may be associated with the anti-stress action of ginseng. However, the saponins, which showed the great reduction of the CA secretion, were mainly the protopanaxiatriols. The protopanaxadiol and oleanolic acid saponins had a little or little such effect. Recent studies demonstrated that the oligosaccharides connected to the hydroxyl groups of the aglycones of the saponins are in turn hydrolyzed by gastric acid and enzymes in the intestinal bacteria when the ginseng is orally administrated. In this study, the effects of their major 6 kinds of metabolites on the secretion of CAs were investigated. All metabolites (M1, 2, 3 and 5 derived from the protopanaxadiols, and M4 and 11 from the protopanaxiatriols) reduced the ACh-evoked secretion from the cells. In the metabolites, the M4 inhibition was the most potent ($IC_{50}({\mu}M):M4(9)$ < M2 (18) < M3 (19) < M1l (22) < M5 (36) < MI (38)). Although M4 also reduced the CA secretion induced by high $K^+$, a stimulation activating voltage-sensitive $Ca^{2+}$ channels, the inhibitory effect was much less than that on the ACh-evoked secretion. M4 inhibited the ACh-induced $Na^+$ influx into the cells in a concentration-dependent manner similar to that of the inhibition of the ACh-evoked secretion. When the cells were washed by the incubation buffer after the preincubation of the cells with M4 and then incubated without M4 in the presence of ACh, the M4 inhibition was not completely abolished. On the other hand, its inhibition was maintained even by increasing the external ACh concentration. These results indicate that the saponins are metabolized to the more active substances in the digestive tract and the metabolites attenuate the secretion of CAs from bovine adrenal chromaffin cells stimulated by ACh due to the noncompetitive blockade of the ACh-induced $Na^+$ influx into the cells. These findings may further explain the anti-stress action of ginseng.

• 한국의학물리학회지:의학물리
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• 제25권3호
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• pp.157-166
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• 2014

뉴론에서 ATP는 분비 과립내에 신경전달물질과 함께 다량 저장되어 있다가 신경전달물질과 함께 분비되는 것으로 알려져 있으므로 신경전달물질의 자극-분비(stimulus-secretion) coupling 과정에 있어 중요한 조절작용을 할 것으로 기대된다. 그러므로 본 연구에서는 뉴론과 그 발생학적 기원이 동일한 부신수질 세포(adrenal chromaffin cell)를 대상으로 하여 세포막 칼슘통로를 통한 세포막 전류에 미치는 ATP의 영향을 측정함으로써 신경전달물질이 자극-분비 coupling 과정에 작용하는 ATP의 조절 작용을 알아보고자 하였다. 부신수질 세포의 칼슘통로를 통한 세포막 전류는 패치클램프 테크닉으로 기록하였다. 10 mM $Ba^{2+}$을 포함한 세포 외 용액에서, $Ba^{2+}$ current는 0.1 mM ATP를 세포외부에 처치했을 때, 평균 $36{\pm}6%$ (n=6) 감소되어 나타났고 ATP를 씻어준 후 전류는 다시 회복되는 가역적 반응을 보였다. ATP의 전류 억제 기전을 알아보고자 칼슘통로에서 관찰되는 현상 중의 하나인 소통(facilitation)현상을 기록하였다. +80 mV의 큰 prepulse를 준 후 바로 테스트 펄스를 주며 측정한 전류는 큰 prepulse에 의해 억제효과가 풀리는(disinhibition) 현상을 나타내었다. ATP 처치 후 큰 자극을 주어 $37{\pm}5%$ (n=11)의 $Ba^{2+}$ 전류 증가가 있었고 이는 ATP가 없는 상태에서 순수하게 큰 자극에 의해 소통되는 $25{\pm}3%$ (n=12)과 유의한 차이를 보였다(p<0.05). ATP의 억제 기전이 G-protein을 매개로 한 것인지를 알아보고자 가수분해 되지 않는 GTP 유도체인 $GTP{\gamma}S$를 세포 내에 준 후 $Ba^{2+}$ 전류를 기록하였다. $GTP{\gamma}S$에 의해 55%의 전류 크기의 감소가 있었고 이 환경에서 큰 prepulse를 인가하였을 때 $34{\pm}4%$ (n=19)의 소통현상을 보였다. 이는 $GTP{\gamma}S$가 없는 환경에서의 $25{\pm}3%$ (n=12)의 소통현상을 보인 것과 유의한 차이를 보였다(p<0.05). $Ba^{2+}$ current trace의 활성화 과정(activation)을 curve-fitting한 결과, control은 single exponential curve로 fitting된 반면, ATP 또는 $GTP{\gamma}S$를 처치한 경우, 그리고 ATP와 $GTP{\gamma}S$ 모두 처치한 경우에서는 double-exponential curve로 가장 잘 fitting이 되었다. 즉, ATP나 $GTP{\gamma}S$를 처치했을 때 모두 전류가 더 느리게 활성화되는 모양을 나타내었고, 이상의 결과로 미루어 ATP와 $GTP{\gamma}S$는 같은 방식으로 칼슘통로를 억제하고, 이러한 억제효과는 세포막에 아주 큰 전압을 걸어주면 칼슘 통로에 결합했던 G-protein이 막전압 의존적으로 떨어짐으로써 소실(disinhibition)된다고 해석된다. 본 연구에서 확인한 ATP의 칼슘통로 억제효과는 자체 크로마핀 세포 또는 주변 세포에서 아드레날린이 적게 분비되게 하는 autocrine 또는 paracrine inhibition 과정의 중요한 기전으로 작용할 것이다.

• Applied Microscopy
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• 제22권1호
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• pp.33-41
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• 1992

To clarify the exocytotic features in adrenal medullary aminergic cells, the authors observed rat adrenal medulla prepared by the TAGO method with transmission electron microscope. Rat adrenal medulla contains two types of aminergic cells, adrenergic and noradrenergic, as described. They were present as a group. In a single group both adrenergic and noradrenergic cells were present, but the same kind of cells showed the tendency forming small groups. Adrenergic cells were characterized with the granules having relatively electroluscent cores. These granules were relatively uniform in size, and the cores filled the granules with only thin halos. Noradrenergic cells were characterized with the granules of various size and forms. Most of the cores of these granules were generally more electron-dense than those of the adrenergic cells and only partly filled the granules without forming the halos. But, some granules were very similar in the shape and electron density as those of the adrenergic cells. Even empty-looking granules were present. Exocytotic figures with the classical omega figures were observed in both types of aminergic cells, but they were more frequent in adrenergic cells. These figures were mainly present along the plasma membranes toward the capillary. The excreted materials could be identified in the cleft of the omega figures. Apocrine-like secretory patterns but without cytoplasmic rims were identified in noradrenergic cells. Some vesicles, possibly formed from the cytoplsmic tubular systems were released. Some irregular lamellar structures of varying sizes were also observed. They looked like membranous structures sneaking through the plasma membranes. We could not, however, found any evidences of their involvement in exocytotic processes. These were present toward the capillaries and found only in the adrenergic cells. The authors conclude that the secretory processes in adrenal chromaffin cells may include not only the classical exocytotic processes but also the unusual direct secretions of granules or parts of cellular organelles. The membranous lamellar structures may indicate the remnants of excreted granules or functionally inactive excess membranes of the organelles removed from the cytoplasm.

• 생명과학회지
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• 제19권10호
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• pp.1346-1351
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• 2009

Neural crest는 신경계의 발생과정에서 생긴 특정화된 외배엽으로서 말초신경계(peripheral nervous system)의 모든 sensory cells과 peripheral cells, unipolar spinal ganglion cell, cranial sensory ganglia, peripheral nerve의 neurolemmal sheath cells, ganglia의 capsule cells, sympathetic ganglia, chromaffin cells, pigment cell 등의 자율신 경계의 대부분의 세포로 분화 한다. 최근pluripotetic neural crest cells의 운명이 이미 제한되어 있으며, 이러한 fate-restricted crest cells이 neural tube에서 emigration된다고 보고된바 있다. 또한 본 연구자는 Wnt와 Wnt의 antagonist가 neural crest cell의 specification이 일어나는 시기에 발현하여, neural crest cell의 segregation과 differentiation에 직접적으로 관여함을 밝혔다. 이를 보다 명확히 규명하기 위해, 본 연구에서는 neural tube에 Wnt-3a expressing cell의 grafting 혹은 dominant negative GSK construct의 electroporation을 통해 Wnt signaling을 modulation 하여 downstream mediator를 조사하였다. Wnt signaling의 stimulation은 neural crest cell의 melanoblast 로의 commitment를 유도하였으며, 이와 더불어 cadherin 7과 slug의 발현을 조절함을 확인하였다.

• IMMUNE NETWORK
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• 제1권1호
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• pp.53-60
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• 2001

Background: The human mic2 gene is a pseudoautosomal gene that encodes a cell surface antigen, CD99. High levels of CD99 constitute a tumor marker in Ewing s sarcoma (ES). We have recently demonstrated that CD99-induced apoptosis occurs only in undifferentiated ES cells, not in differentiated ES cells, raising the possibility of the involvement of CD99 in neural ontogeny. Methods: To elucidate the relations between the expression of CD99 and the differentiation of neural cells and the mechanism by which the expression of CD99 is regulated, we analyzed the differential patterns of CD99 expression in SH-SY5Y by treatment of 12-O-tetradecanoyl-13-phorbol acetate (TPA) and retinoic acid. In addition, to explore the transcriptional activity of CD 99 during neural cell differentiation, SH-SY5Y cells were transiently transfected with a CD99 promoter-driven luciferase construct, and treated with the inducers. Results: In immunoblotting and flow cytometry, the expression level of CD99 was increased on differentiated SH-SY5Y cells induced by TPA and retinoic acid. The luciferase activity was elevated by the treatment with TPA, known to mature SH-SY5Y cells toward a sympathetic neuronal lineage, whereas retinoic acid inducing a sympathetic chromaffin lineage displayed little effect. Conclusion: The result indicates that CD99 might be expressed only on cells maturing toward a neuronal lineage among differentiating primitive neuronal cells. In addition, the expression of CD99 seems to be regulated at the transcriptional level during the differentiation.

• 대한약리학회지
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• 제31권1호
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• pp.63-74
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• 1995

저산소 상태에서는 부신수질로부터 카테콜아민 (CA) 유리작용이 활성화되지만 반면에 소의 배양 chromaffin cell에서는 고통도의 $K^+$에 의한 CA 분비작용이 억제된다고 알려져 있다. 본 연구에서는 적출 흰쥐 관류부신에서 콜린성 자극과 막탈분극에 의한 CA 분비작용에 대한 저산소증의 영향을 검색하고 그 작용기전을 규명코자 하였다. 본 연구목적을 위하여, 적출 흰쥐 관류부신을 이용, 저산소증이 니코틴($N_1$), 무스카린($M_1$) 수용체 흥분약, 막탈분극 약물, 칼슘채널 활성화 약물, 세포내 칼슘유리 약물 및 ACh에 의한 CA 분비에 미치는 영향을 연구하였으며, 저산소증은 95% 질소 및 5% 이산화탄소 혼합가스를 Krebs액에 주입하여 유발시켰으며, $3{\sim}4$시간동안 유지하였다. 저산소증 유발시, DMPP ($100{\mu}M$), McN-A-343 ($100{\mu}M$), ACh (5.32 mM), Bay-K-8644 ($10{\mu}M$) 및 high $K^+$ (56 mM)에 의한 CA 분비작용을 시간의존적으로 점차 유의성인 감소를 나타내었다. 그러나, cyclopiazonic acid ($10{\mu}M$)에 의한 CA 분비반응에는 하등의 영향을 일으키지 못하였다. 또한 저산소증 자체가 CA의 기초분비 작용에는 영향을 미치지 않았다. 이와같은 실험결과로 보아, 저산소증시 콜린성 자극 및 막탈분극에 의한 CA 분비 작용이 억제되며, 이러한 억제작용은 chromaffin cell내로 $Ca^{++}$ 유입을 직접적으로 억제시키는 결과에 기인되며, 세포내 칼슘저장고로부터 칼슘유리작용과는 관계없는 것으로 사료된다.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 1998년도 Advances in Ginseng Research - Proceedings of the 7th International Symposium on Ginseng -
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• pp.40-46
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• 1998

We investigated the influence of the root of Panax ginseng C. A. Meyer on the secretion of catecholamines from bovine adrenal chromaffin cells, which are used as a model of nervous systems. In two major parts extracted from the ginseng root, the crude saponin fraction, but not the non-saponin fraction, reduced the secretion from the cells, stimulated by acetylcholine (ACh). Ginseng saponins (ginsenosides) are classified into three groups, the panaxadiol, the panaxatriol and the oleanolic acid groups, on the basis of the chemical structures of their saponins. Both the panaxadiol and the panaxatriol saponins, excluding only one oleanolic acid saponin ginsenoside-Ro, generally reduced the ACh-evoked secretion. The inhibitory effects of the panaxatriol were much stronger than those of the panaxadiol. However, ginsenoside-Rg, and -Rh3 in the panaxadiol saponins were the potent inhibitors comparable to the panaxatriol saponins. Ginsenoside-Rg2 in the panaxatriol was the most effective. It is probable that the ginsenoside inhibition of the catecholamine secretion is due to the suppression of the function of the nicotinic ACh receptor-cation channels. On the other hand, ginsenoside-Rg2 did not affect the angiotensin II-, the bradykinin-, the histamine- and the neurotensin- induced catecholamine secretions from the chromaffin cells and the muscarine- and the histamine- induced contraction of the ileum in guinea-pigs. Ginsenoside-Rbl, a panaxadiol saponin, and ginsenoside-Ro had no or only a slight effect on them. On the contrary, ginsenoside-Rg3 not only competitively inhibited the muscarine-induced ileum contraction but also reduced the angiotensin R -, the bradykinin-, the histamine- and the neurotensin-induced catecholamine secretions. Thus, the ginseng root contains active ingredients, namely some ginsensides, which suppress the responses induced by receptor stimulation. The inhibitory effects of ginseng saponins may be one of the action mechanisms for the pharmacological effects of the Panax ginseng root.

• 대한약리학회지
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• 제29권2호
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• pp.237-244
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• 1993

소 부신수실 크롬친화성 세포(BAMC)에서 $[Met^5]-enkephalin$ (ME)의 분비와 Proenkephalin A (proENK) mRNA의 함량에 대한 nicotine의 영향과 이에 대한 indomethacin, nordihydroguaiaretic acid(NDGA) 및 captopril의 작용을 연구하였다. 10 uM Nicotine으로 BAMC 세포를 장기간 자극시 proENK mRNA의 함량과 배양액으로의 ME 분비가 유의하게 증가하였다. BAMC 세포를 NDGA(lipoxygenase 억제제, 10 uM), indomethacin (cyclooxygenase 억제제), captopril (angiotensin 변환효소 억제제)만으로 처리시에는 ME 분비와 proENK mRNA의 함량에 영향이 없었다. Nicotine에 의한 ME 분비와 proENK mRNA 함량의 증가는 NDGA에 의하여 억제되었다. 그러나 indomethacin과 captopril은 nicotine의 작용에 대하여 아무 영향이 없었다. 이들 결과는 nicotine에 의한 ME 분비와 proENK mRNA 함량의 증가가 부분적으로 lipoxygenase 경로에 의해 매개되며, cyclooxygenase 및 내재성 renin-angiotensin 경로는 관련되지 않음을 나타낸다.

• The Korean Journal of Physiology and Pharmacology
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• 제8권5호
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• pp.265-272
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• 2004

The present study was attempted to investigate the effect of cilnidipine (FRC-8635), which is a newly synthesised novel dihydropyridine (DHP) type of organic $Ca^{2+}$ channel blockers, on secretion of catecholamines (CA) evoked by acetylcholine (ACh), high $K^+$, DMPP and McN-A-343 from the isolated perfused rat adrenal gland. Cilnidipine $(1{\sim}10{\mu}M)$ perfused into an adrenal vein for 60 min produced relatively dose- and time-dependent inhibition in CA secretory responses evoked by ACh $(5.32{\times}10^{-3}M),\;DMPP\;(10^{-4}M\;for\;2\;min)$ and McN-A-343 $(10^{-4}M\;for\;2\;min)$. However, lower dose of cilnidipine did not affect CA secretion by high $K^+\;(5.6{\times}10^{-2}\;M)$, higher dose of it reduced greatly CA secretion of high $K^{+}$. Cilnidipine itself did fail to affect basal catecholamine output. In the presence of cilnidipine $(10{\mu}M)$, the CA secretory responses evoked by Bay-K-8644 $(10{\mu}M)$, an activator of L-type $Ca^{2+}$ channels and cyclopiazonic acid $(10{\mu}M)$, an inhibitor of cytoplasmic $Ca^{2+}$-ATPase were also inhibited. Moreover, ${\omega}-conotoxin\;GVIA\;(1{\mu}M)$, a selective blocker of the N-type $Ca^{2+}$ channels, given into the adrenal gland for 60 min, also inhibited time-dependently CA secretory responses evoked by Ach, high $K^+$, DMPP, McN-A-343, Bay-K-8644 and cyclopiazonic acid. Taken together, these results demostrate that cilnidipine inhibits CA secretion evoked by stimulation of cholinergic (both nicotinic and muscarinic) receptors from the isolated perfused rat adrenal gland without affecting the basal release. However, at lower dose, cilnidipine did not affect CA release by membrane depolarization while at larger dose inhibited that. It seems likely that this inhibitory effect of cilnidipine is exerted by blocking both L- and N-type voltage-dependent $Ca^{2+}$ channels (VDCCs) on the rat adrenomedullary chromaffin cells, which is relevant to inhibition of both the $Ca^{2+}$ influx into the adrenal chromaffin cells and intracellular $Ca^{2+}$ release from the cytoplasmic store. It is thought that N-type VDCCs may play an important role in regulation of CA release from the rat adrenal medulla.