• 한국의학물리학회지:의학물리
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• 제8권1호
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• pp.3-15
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• 1997

Adrenal chromaffin cells secrete catecholamine in response to acetylcholine. The secretory response has absolute requirement for extracellular calcium, indication that $Ca^{2+}$ influx through voltage dependent $Ca^{2+}$ channel (VDCC) is the primary trigger of the secretion cascade. Although the existence of various types of $Ca^{2+}$ channels has been explored using patch clamp technique in adrenal chromaffin cells, the contribution of different types of $Ca^{2+}$ channels to catecholamine secretion remains to be established. To investigate the quantative contribution of different types of $Ca^{2+}$ channels to cate-cholamine secretion, $Ca^{2+}$ current($I_{Ca}$) and the resultant membrane capacitance increment($\Delta{C}_{m}$) were simultaneoulsy measured. Software based phasor detector technique was used to monitor $\Delta{C}_{m}$. After blockade of L type VDCC with nicardipine (1$\mu$M), $I_{ca}$ was blocked to 43.85$\pm$6.72%(mean$\pm$SEM) of control and the resultant ㅿC$_{m}$ was reduced ot 30.10$\pm$16.44% of control. In the presence of nicardipine and $\omega$-conotoxin in GVIA(l$\mu$M), an N type VDCC antagonist, $I_{ca}$ was blocked to 11.62$\pm$2.96% of control and the resultant $\Delta{C}_{m}$ was reduced to 26.13$\pm$8.25% of control. Finally, in the presence of L, N, and P type $Ca^{2\pm}$ channel antagonists(nicardipine, $\omega$-Conotoxin GVIA, and $\omega$-agatoxin IVA, respectively), $I_{ca}$ and resultant $\Delta{C}_{m}$ were almost completely blocked. From the observation of parallel effects of $Ca^{2+}$ channel antagonists on $I_{ca}$ and $\Delta{C}_{m}$, it was concluded that L, N, and also P type $Ca^{2+}$ channels served and $Ca^{2+}$ source for exocytosis and no difference was observed in their efficiency to evoke exocytosis amost L, N, and P type $Ca^{2+}$ channels.

• The Korean Journal of Physiology and Pharmacology
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• 제6권2호
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• pp.113-119
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• 2002

Long-term potentiation (LTP) at hippocampal CA3-CA1 synapses is often associated with increases in quantal size, traditionally attributed to enhanced availability or efficacy of postsynaptic glutamate receptors. However, augmented quantal size might also reflect increases in neurotransmitter concentration within the synaptic cleft since AMPA-type glutamate receptors are not generally saturated during basal transmission. Here we report evidence that peak cleft glutamate concentration $([glu]_{cleft})$ increases during LTP, as indicated by a lessening of the blocking effects of rapidly unbinding antagonists of AMPA. The efficacy of slowly equilibrating antagonists remained unchanged. The elevated $[glu]_{cleft}$ helps support the increased quantal amplitude of AMPA-type EPSCs (excitatory postsynaptic currents) during LTP.

• The Korean Journal of Physiology and Pharmacology
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• 제6권2호
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• pp.109-112
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• 2002

The signal pathways involved in the regulation of AP-1 DNA binding activity in long-term nicotine stimulated bovine adrenal medullary chromaffin (BAMC) cells have not been well characterized. To understand the involvement of second messengers in the regulation of AP-1 DNA binding activity, the present study was designed to define the time-course for inhibition of nicotine-induced responses by cholinergic antagonists, $Ca^{2+}$ and calmodulin (CaM) antagonists, and calcium/calmodulin-dependent protein kinase (CaMK) II inhibitor using electrophoretic mobility shift assay. Nicotine $(10{\mu}M)$ stimulation increased AP-1 DNA binding activity at 24 hr after treatment. Posttreatment with hexamethonium (1 mM) plus atropine $(1{\mu}M)$ (HA), nimodipine $(1{\mu}M),$ or calmidazolium $(1{\mu}M)$ at 0.5, 3, and 6 hr after the nicotine treatment significantly inhibited the AP-1 DNA binding activity increased by long-term nicotine stimulation. However, posttreatment with HA, nimodipine, or calmidazolium at 9 or 12 hr after the nicotine treatment did not affect the nicotine-induced increase of AP-1 DNA binding activity. The pretreatment of BAMC cells with various concentrations of KN-62 inhibited the increase of AP-1 DNA binding activity induced by nicotine in a concentration-dependent manner. KN-62 $(10{\mu}M)$ posttreatment beginning at 0.5, 3, or 6 hr after the nicotine treatment significantly inhibited the increase of AP-1 DNA binding activity. However, KN-62 posttreatment beginning at 9 or 12 hr after the nicotine treatment did not affect the increase of AP-1 DNA binding activity. This study suggested that stimulation (for at least 6 hr) of nicotinic receptors on BAMC cells was necessary for increase of AP-1 DNA binding activity, and activation of $Ca^{2+},$ CaM, and CaMK II up to 6 hr at least seemed to be required for the increase of nicotine-induced AP-1 DNA binding activity.

• The Korean Journal of Physiology and Pharmacology
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• 제6권4호
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• pp.187-191
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• 2002

Tamoxifen, an antiestrogen, has previously been shown to induce apoptosis in HepG2 human hepatoblastoma cells through activation of the pathways independent of estrogen receptors, i.e., intracellular $Ca^{2+}$ increase and generation of reactive oxygen species (ROS). However, the mechanism of tamoxifen to link increased intracellular $Ca^{2+}$ to ROS generation is currently unknown. Thus, in this study we investigated the possible involvement of calmodulin, a $Ca^{2+}$ activated protein, and $Ca^{2+}$/calmodulin-dependent protein kinase II in the above tamoxifen-induced events. Treatment with calmodulin antagonists (calmidazolium and trifluoroperazine) or specific inhibitors of $Ca^{2+}$/calmodulin-dependent protein kinase II (KN-93 and KN-62) inhibited the tamoxifen-induced apoptosis in a dose-dependent manner. In addition, these agents blocked the tamoxifen-induced ROS generation in a concentration-dependent fashion, which was completely suppressed by intracellular $Ca^{2+}$ chelation. These results demonstrate for the first time that, despite of its well-known direct calmodulin-inhibitory activity, tamoxifen may generate ROS and induce apoptosis through indirect activation of calmodulin and $Ca^{2+}$/calmodulin-dependent protein kinase II in HepG2 cells.

• The Korean Journal of Physiology
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• 제18권1호
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• pp.37-50
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• 1984

The effects of $Ca^{++}$ and its antagonists (verapamil and $La^{+++}$) upon the spontaneous contraction and the contracture induced by 60 mM K-Tyrode solution were studied in the isolated uterine muscle. Longitudinal muscle strips were prepared from the rat uteri at estrous stage. All experiments were performed in tris-buffered Tyrode solution which was aerated with 100% $O_2$ and kept at 35^{\circ}$. The results obtained were as follows: 1) In the uterine strips contracting spontaneously, both the amplitude of peak tension and the area of contraction curve increased dose-dependently in the range of $0.5${\sim}8$ mM $Ca^{++}$. The frequency of contraction increased as the concentration of $Ca^{++}$ increased up to 2 mM, but above this concentration the frequency decreased. In $Ca^{++}-free$ media, however, contraction did not develop. In the contracture induced by 60 mM K-Tyrode solution, the developed tension increased dose-dependently as the concentration of external $Ca^{++}$ increased to 8 mM. In the absence of external $Ca^{++}$ K-contracture appeared, but it was not sustained. 2) The spontaneous contraction of rat uterus was suppressed by verapamil in proportion to an increase of its concentration and totally abolished at the concentration of $3{\times}10^{-4}\;g/l$, but the spontaneous contraction re-appeared by addition of $Ca^{++}$. The amplitude of peak tension recovered completely but the recovery of frequency was incomplete. K-contracture decreased in a dose-dependent manner after the treatment with verapamil and totally disappeared at its concentration of $3{\times}10^{-4}\;g/l$. Even in this case contracture developed again by extra $Ca^{++}$. 3) The spontaneous contractile activity was inhibited by $La^{+++}$. At the concentration of $10^{-4}$M $La^{+++}$, fibrillation appeared. In the strip inhibited by $10^{-5}M\;La^{+++}$, contractility recovered completely by extra $Ca^{++}$ while in the $10^{-4}M\;La^{+++}$ treated preparation, the rhythmic spontaneous contraction did not develop even at the concentration of 16 mM $Ca^{++}$. After the initial transient depression of contracture tension by $10^{-3}M$ of $La^{+++}$, the strip stowed considerably large size of contracture, hardly influenced by external $Ca^{++}$ or verapamil. The results obtained in this experiment suggest that in the rat uterine muscle there would be some competitive actions between $Ca^{++}$ and its antagonists. It is speculated that $Ca^{++}$ plays an important role in the conduction of excitation, and $La^{+++}$ influences upon cellular $Ca^{++}$ mobilization and re-uptake process as well as transmembrane $Ca^{++}$ transport in a K-depolarized state.

• Biomolecules & Therapeutics
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• 제25권2호
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• pp.194-201
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• 2017

Lysophosphatidylethanolamine (LPE), a lyso-type metabolite of phosphatidylethanolamine, has been reported to be an intercellular signaling molecule. LPE mobilizes intracellular $Ca^{2+}$ through G-protein-coupled receptor (GPCR) in some cells types. However, GPCRs for lysophosphatidic acid (LPA) were not implicated in the LPE-mediated activities in LPA GPCR overexpression systems or in SK-OV3 ovarian cancer cells. In the present study, in human SH-SY5Y neuroblastoma cells, experiments with $LPA_1$ antagonists showed LPE induced intracellular $Ca^{2+}$ increases in an $LPA_1$ GPCR-dependent manner. Furthermore, LPE increased intracellular $Ca^{2+}$ through pertussis-sensitive G proteins, edelfosine-sensitive-phospholipase C, 2-APB-sensitive $IP_3$ receptors, $Ca^{2+}$ release from intracellular $Ca^{2+}$ stores, and subsequent $Ca^{2+}$ influx across plasma membranes, and LPA acted on $LPA_1$ and $LPA_2$ receptors to induce $Ca^{2+}$ response in a 2-APB-sensitive and insensitive manner. These findings suggest novel involvements for LPE and LPA in calcium signaling in human SH-SY5Y neuroblastoma cells.

• 대한약리학회지
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• 제23권1호
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• pp.33-44
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• 1987

면역보체가 결합되어 있는 zymosan 또는 열로 응집된 IgG에 의하여 활성화된 호중구에서의 NADPH oxidase 의존적인 $O_{\overline{2}}$의 생성과 탐식작용은 칼슘의 흡수과정과 일치하였다. 활성화된 백혈구의 반응은 세포외 칼슘 농도에 따라 항진되었으며, 이는 칼슘의 킬레이트제인 EGTA나 EDTA에 의하여 유의하게 억제되었다. 활성화된 백혈구로부터 $O_{\overline{2}}$의 생성은 dantrolene과 chlorpromazine에 의하여 억제되었다. 칼슘 길항제인 bepredil, diltiazem, verapamil, nifedipine, nimodipine은 효과적으로 활성화된 백혈구의 칼슘흡수, $O_{\overline{2}}$ 생성 그리고 탐식 작용을 억제하였고 NADPH oxidase 활성도 또한 억제하였다. 그러므로, 칼슘 길항제는 칼슘 유입을 억제하거나 칼슘의 세포내 재분포 및 NADPH oxidase 반응계에 작용하여 활성화된 백혈구에서의 $O_{\overline{2}}$의 생성과 백혈구의 탐식작용을 억제할 것으로 시사되었다.

• Clinical and Experimental Reproductive Medicine
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• 제30권4호
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• pp.269-280
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• 2003

Objective: We reported the overcoming effect of $Ni^{2+}$ on the in vitro 2-cell block of mouse embryos. In this study, we aim to investigate whether $Ni^{2+}$ should induce intracellular $Ca^{2+}$ transient in the mouse embryos. Materials and Methods: Embryos were collected at post hCG 32hr from the oviduct of the ICR mouse and cultured in M2 medium omitted phenol red. Intracellular $Ca^{2+}$ was checked by using a confocal laser scanning microscope and fluo-3AM by using various intracellular $Ca^{2+}$ antagonists. Results: In 1mM $Ni^{2+}$ treated medium which contained $Ca^{2+}$(1.71mM), 75.7% of the embryos showed $[Ca^{2+}]i$ transient about 200 sec later. In the $Ca^{2+}$-free medium, 69.8% of the embryos showed $[Ca^{2+}]i$ transient. In U73122, phospholipaseC(PLC) inhibitor (5uM, 10min) pretreated group, 33.3% of the embryos showed $[Ca^{2+}]i$ transient. Heparine, inositol 1, 4, 5-triphosphate receptor(IP3R) antagonist preinjected embryos showed no response with 1mM $Ni^{2+}$. In danthrolene treatment, ryanodine receptor(RyR)-antagonist, 43% embryos showed $[Ca^{2+}]i$ transient but they showed delayed response about 340sec in the presence of $Ca^{2+}$. Conclusions: Summing up the above results, $Ni^{2+}$ seems to induce $Ca^{2+}$-release from the $Ca^{2+}$-store even in the $Ca^{2+}$-free medium. IP3 receptors of the mouse 2-cell embryos might have an essential role for the intracellular $Ca^{2+}$ increase by $Ni^{2+}$.

• Asian-Australasian Journal of Animal Sciences
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• 제15권7호
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• pp.949-956
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• 2002

There are several physiological and pharmacological evidences indicating that opening of voltage dependent $Ca^{2+}$ channels play a critical role in induction of acrosome reaction in mammalian sperm. We determined the intracellular free $Ca^{2+}$ concentration in ejaculated goat sperm using a fluorescent, $Ca^{2+}$-specific probe, Fura2/AM, after the suspension of sperm in KRB medium, capable of sustaining capacitation and the acrosome reaction. We used nifedipine, D-600 and diltiazem, the $Ca^{2+}$ channel antagonists belonging to the classes of dihydropyridines, phenylalkylamines and benzothiazepines, to investigate the possibility that L-type voltage gated $Ca^{2+}$ channels play a role in the progesterone-stimulated exocytotic response. Progesterone promoted a rise in intracellular $Ca^{2+}$ in goat sperm and addition of nifedipine (100 nM) just prior to progesterone induction, significantly inhibited both intracellular $Ca^{2+}$ rise and exocytosis suggesting that $Ca^{2+}$ channels are involved in the process. However, the intracellular $Ca^{2+}$ increase during the process of capacitation was not affected with the addition of nifedipine suggesting a role of focal channel for $Ca^{2+}$ during capacitation. Studies using monensin and nigericin, two monovalent cation ionophores showed that an influx of $Na^+$ also may play a role in the opening of $Ca^{2+}$ channels. These results strongly suggests that the entry of $Ca^{2+}$ channels with characteristics similar to those of L-type, voltage-sensitive $Ca^{2+}$ channels found in cardiac and skeletal muscle, is a crucial step in the sequence of events leading to progesterone induced acrosome reaction in goat sperm.

• 대한의생명과학회지
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• 제25권2호
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• pp.123-130
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• 2019

Platelet aggregation is essential for hemostatic process in case of blood vessels damages. However, excessive platelet aggregation can cause cardiovascular disorders including atherosclerosis, thrombosis and myocardial infarction. Scopoletin is usually found in the roots of genus Scopolia or Artemisia, and is known to have anticoagulant and anti-malarial effects. This study investigated the effect of scopoletin on human platelet aggregation induced by U46619, an analogue of thromboxane $A_2(TXA_2)$. Scopoletin had anti-platelet effects by down-regulating $TXA_2$ and intracellular $Ca^{2+}$ mobilization ($[Ca^{2+}]_i$), the aggregation-inducing molecules generated in activated platelets. On the other hand, scopoletin increased the levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), which are known to be intracellular $Ca^{2+}$ antagonists. This resulted in inhibition of fibrinogen binding to ${\alpha}IIb/{\beta}_3$ in U46619-induced human platelet aggregation. In addition, scopoletin inhibited the release of adenosine trisphosphate (ATP) in dose-dependent manner. This result means that the aggregation amplification activity through the granule secretion in platelets was suppressed by scopoletin. Therefore, we demonstrated that scopoletin has a potent antiplatelet effect and is highly likely to prevent platelet-derived vascular disease.