• The Korean Journal of Physiology
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• v.28 no.1
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• pp.51-59
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• 1994

This study was carried out to elucidate the excitatory mechanisms of Substance P in the antral circular muscle, using isometric contraction recording, conventional microelectrode method and whole-cell patch clamp technique. Substance P produced tonic and phasic contractions in a dose-dependent manner and depolarized membrane potential with increased amplitude of slow waves in muscle strips. Voltage-dependent $Ca^{2+}$ currents were increased by the application of Substance P from a holding potential of -60mV to 50mV in 10mV steps and this effect was blocked by the addition of an antagonist. Also Substance P increased transient and spontaneous oscillatory $K^+$ outward currents. The enhanced outward currents were abolished by apamin in dispersed single cells. These results suggest that the depolarization of membrane potential by Substance P activates voltage-dependent $Ca^{2+}$ channels, which represents an excitatory response in the antral circular muscle and led to an increase in $Ca^{2+}\;activated\;K^+\;currents$.

• Journal of Ginseng Research
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• v.24 no.1
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• pp.18-22
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• 2000

In previous reports we have shown that ginsenosides inhibit high threshold voltage-dependent $Ca^{2+}$ channels in neuronal cells. However, these studies did not show whether ginsenosides-induced inhibition of $Ca^{2+}$ currents discriminates among the various $Ca^{2+}$ channel subtypes, although it is known that there are at least five different $Ca^{2+}$ channel subtypes in neuronal cells. In this study we investigated the effect of ginsenosides on high threshold voltage-dependent $Ca^{2+}$ channel subtypes using their selective $Ca^{2+}$ channel blockers nimodipine (L-type), $\omega$-conotoxin GVIA (N-type), or $\omega$-agatoxin IVA (P-type) in bovine chromaffin cells. We could observe that ginsenosides inhibited high threshold voltage-dependent $Ca^{2+}$ currents in a dose-dependent manner. The $IC_{50}$/ was about 120 $\mu$g/ml. Nimodipine had no effect on ginsenosides response. However, the effect of ginsenosides on $Ca^{2+}$ currents was reduced by $\omega$-conotoxin GVIA, $\omega$-agatoxin IVA, and mixture of nimodipine, $\omega$-contoxin GVIA, and $\omega$-agatoxin IVA. These data suggest that ginsenosides are negatively coupled to three types of calcium channels in bovine chromaffin cell, including an $\omega$-conotoxin GVIA-sensitive (N-type) channel, an $\omega$-agatoxin IVA-sensitive (P-type) channel and nimodipine/$\omega$-conotoxin GVIA/$\omega$-agatoxin IVA-resistant (presumptive Q-type) channel.Q-type) channel.

• Progress in Medical Physics
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• v.15 no.4
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• pp.220-227
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• 2004

In N-type $Ca^{2＋}$ channels, the mechanism of inactivation - decline of inward current during a depolarizing voltage step- is still controversial between voltage-dependent inactivation and $Ca^{2＋}$ -dependent inactivation. In the previous paper we demonstrated that fast component of inactivation of N-type calcium channels does not involve classic $Ca^{2＋}$ -dependent mechanism and the slowly inactivating component could result from a $Ca^{2＋}$ -dependent process. However, there should be signal transduction pathway which enhances inactivation no matter what the inactivation mechanism is. We have investigated the effect of phosphorylation on calcium channels of rat sympathetic neurons. Intracellular dialysis with the phosphatase inhibitors okadaic acid markedly enhanced the inactivation. The rapidly inactivating component is N-type calcium current, which is blocked by $\omega$-conotoxin GVIA. Staurosporine, a nonselective protein kinase inhibitor, prevented the action of okadaic acid, suggesting that protein phosphorylation is involved. More specifically lavendustin C, inhibitor of CaM kinase II, prevented the action of okadaic acid, suggesting that calmodulin dependent pathway is involved in inactivation process. It is not certain to this point whether phosphorylation process is inactivation itself. Molecular biological research regarding binding site should be followed to address the question of how the divalent cation binding site is related to phoshorylation process.

• Proceedings of the Korean Biophysical Society Conference
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• 2001.06a
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• pp.26-26
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• 2001

Inactivation of N-type calcium current has been reported to be both voltage dependent and Ca$\^$2＋/ dependent. We have investigated the effects of Ba$\^$2＋/ and Ca$\^$2＋/ on N-channel inactivation in rat superior cervical ganglion neurons using the whole cell configuration of patch clamp technique. Inactivation was larger in Ca$\^$2＋/ than in Ba$\^$2＋/ even with 20 mM BAPTA.(omitted)

• Molecules and Cells
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• v.21 no.1
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• pp.121-128
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• 2006

Maitotoxin (MTX) is known as one of the most potent marine toxins involved in Ciguatera poisoning, but intracellular signaling pathways caused by MTX was not fully understood. Thus, we have investigated whether intracellular reactive oxygen species (ROS) are involved in MTX-induced cellular responses in human umbilical vein endothelial cells. MTX induced a dose-dependent increase of intracellular [$Ca^{2+}$]. MTX stimulated the production of intracellular ROS in a dose- and time-dependent manner, which was suppressed by BAPTA-AM, an intracellular $Ca^{2+}$ chelator. Ionomycin also elevated the ROS production in a dose-dependent manner. MTX elevated transamidation activity in a time-dependent manner and the activation was largely inhibited by transfection of tissue transglutaminase siRNA. The activation of tissue transglutaminase and ERK1/2 by MTX was suppressed by BAPTA-AM or ROS scavengers. In addition, MTX-induced cell death was significantly delayed by BAPTA-AM or a ROS scavenger. These results suggest that [$Ca^{2+}$]-dependent generation of intracellular ROS, at least in part, play an important role in MTX-stimulated cellular responses, such as activation of tTGase, ERK phosphorylation, and induction of cell death, in human umbilical vein endothelial cells.

• Journal of Life Science
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• v.27 no.3
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• pp.310-317
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• 2017

Mammalian cells control cellular homeostasis using a variety of defensive enzymes in order to combat against environmental oxidants and electrophiles. NF-E2-related factor-2 (NRF2) is a transcription factor that, in response to an exposure to oxidative stress, translocates into the nucleus and modulates the inducible expression of various phase II cytoprotective enzymes by binding to the antioxidant response element (ARE). In the present study, we have acquired 400 ethanol extracts of traditional medicinal plants and attempted to find out possible extract(s) that can increase the NRF2/ARE-dependent gene expression in human keratinocytes. As a result, we have identified that ethanol extracts of Rheum undulatum and Inula japonica strongly activated the ARE-dependent luciferase activity in HaCaT- ARE-luciferase cells. Exposure of ethanol extracts of Rheum undulatum and Inula japonica increased the viability and activated transcription and translation of NRF2-dependent phase II cytoprotective enzymes in HaCaT cells, such as heme oxygenase-1 (HO-1) and NAD[P]H:quinone oxidorecutase-1 (NQO1). In addition, ethanol extracts of Rheum undulatum and Inula japonica suppressed 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced generation of intracellular reactive oxygen species (ROS), thereby inhibiting the formation of 8-hydroxyguanosine (8-OHG) and 4-hydroxynonenal (4-HNE) in HaCaT cells. Together, our results demonstrate that ethanol extracts of Rheum undulatum and Inula japonica exert anti-oxidant effects via the induction of NRF2/ARE-dependent gene expression in human keratinocytes.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.5
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• pp.403-408
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• 2000

The outward currents elicited in hamster eggs by depolarizing pulses were studied. The currents appeared to comprise at least two components, a transient outward component $(I_{to})$ and a steady-state outward component $(I_{\infty}).\;I_{to}$ was transiently followed by the cessation of inward $Ca^{2+}$ current $(I_{Ca}),$ and its current-voltage (I-V) relation was a mirror image of that of $(I_{Ca}).$ Either blockade of $(I_{Ca})$ by $Co^{2+}$ or replacement of $Ca^{2+}$ with $Sr^{2+}$ abolished $I_{to}$ without change in $I_{\infty}.$ Intracellular EGTA (10 mM) inhibited $I_{to}$ but not $I_{\infty}.$ suggesting strongly that generation of $I_{to}$ requires intracellular $Ca^{2+}.$ Apamin (1 nM) abolished selectively $I_{to},$ indicatingthat $I_{to}$ is $Ca^{2+}-dependent\;K^+$ current. On the other hand, $I_{\infty}$ was $Ca^{2+}-independent.$ Both $I_{to}$ and $I_{\infty}$ were completely inhibited by internal $Cs^+$ and external TEA. The estimated reversal potential of $I_{to}$ was close to the theoretical $E_K.$ Taken together, both outward currents were carried by $K^+$ channels. From these results, $I_{to}$ is likely to be a current responsible for the hyperpolarizing responses seen in hamster eggs at fertilization.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.4
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• pp.411-417
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• 1998

The role of phospholipase ($A_2\;PLA_2$) in tumor cell growth was investigated using SK-N-MC human neuroblastoma cells. 4-Bromophenacyl bromide (BPB) and mepacrine (Mep), known $PLA_2$ inhibitors, suppressed growth of the tumor cells in a dose-dependent manner without a significant cytotoxicity. Melittin (Mel), a $PLA_2$ activator, enhanced the cell growth in a concentration-dependent fashion. The growth-enhancing effects of Mel were significantly reversed by the co-treatment with $PLA_2$ inhibitors. In addition, Mel induced intracellular $Ca^{2+}$ release from internal stores like as did serum, a known intracellular $Ca^{2+}$ agonist in the tumor cells. Intracellular $Ca^{2+}$ release induced by these agonists was significantly blocked by $PLA_2$ inhibitors at growth-inhibitory concentrations. Arachidonic acid (AA), a product of the $PLA_2-catalyzed$ reaction, induced cell growth enhancement and intracellular $Ca^{2+}$ release. These effects of AA were significantly blocked by BAPTA/AM, an intracellular $Ca^{2+}$ chelator. Taken together, these results suggest that the modulation of $PLA_2$ activity may be one of the regulatory mechanisms of cell growth in human neuroblastoma cells. Intracellular $Ca^{2+}$ may act as a key mediator in the $PLA_2-induced$ growth regulation.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.3
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• pp.163-167
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• 2010

This study investigated the effects of the methanol extracts of Morinda citrifolia containing numerous anthraquinone and iridoid on phospholipase $A_2$ ($PLA_2$) isozyme. $PLA_2$ activity was measured using various $PLA_2$ substrates, including 10-pyrene phosphatidylcholine, 1-palmitoyl-2-[$^{14}C$]arachidonyl phosphatidylcholine ([$^{14}C$]AA-PC), and [$^3H$]arachidonic acid (AA). The methanol extracts suppressed melittin-induced [$_3H$]AA release in a concentration-dependent manner in RAW 264.7 cells, and inhibited $cPLA_2/sPLA_2$-induced hydrolysis of [$^{14}C$]AA-PC in a concentration- and time-dependent manner. A Dixon plot showed that the inhibition by methanol extracts on $cPLA_2$ and $sPLA_2$ appeared to be competitive with inhibition constants ($K_i$) of $3.7{\mu}g/ml$ and $12.6{\mu}g/ml$, respectively. These data suggest that methanol extracts of Morinda citrifolia inhibits both $Ca^{2+}$-dependent $PLA_2$ such as, $cPLA_2$ and $sPLA_2$. Therefore, Morinda citrifolia may possess anti-inflammatory activity secondary to $Ca^{2+}$-dependent $PLA_2$ inhibition.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.4
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• pp.419-425
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• 1998

To clarify the effect of extracellular ATP in cultured C6 glioma cells, ATP-induced cytosolic free calcium ($[Ca^{2+}]_i$) mobilization and cell proliferation were investigated. ATP-induced $[Ca^{2+}]_i$ increased in a dose-dependent manner $(10^{-7}\;M{\sim}10^{-3}\;M)$. ATP-induced $[Ca^{2+}]_i$ increases were slightly slowed in extracellular calcium-free conditions especially in sustained phase. ATP-induced $[Ca^{2+}]_i$ increment was also inhibited by the pretreatment of U73122, a phospholipase C (PLC) inhibitor, in a time-dependent manner. Suramin, a putative $P_{2Y}$ receptor antagonist, dose-dependently weakened ATP-induced $[Ca^{2+}]_i$ mobilization. Significant increases in cell proliferation were observed at 2, 3, and 4 days after ATP was added. Stimulated cell proliferation was also observed with adenosine at days 2 and 3. This cell proliferation was significantly inhibited by the treatment with suramin. Ionomycin also stimulated cell proliferation in a concentration-dependent manner. In conclusion, we suggest that extracellular ATP stimulates C6 glioma cell proliferation via intracellular free calcium mobilization mediated by purinoceptor.