• Progress in Medical Physics
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• v.12 no.1
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• pp.59-70
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• 2001

Adrenal chromaffin cells secrete catecholamine in response to acetylcholine. The secretory response has absolute requirement for extracellular calcium, indicating that $Ca^{2+}$ influx through voltage operated $Ca^{2+}$ channels 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, there is still disagreement with the types of $Ca^{2+}$ channels existed in different species. Therefore, we have tried to identify several distinct types of $Ca^{2+}$ channels in rat chromaffin cells. By using nicardipine(L type channel blocker), $\omega$-CgTx GVIA(N type channel blocker), and $\omega$-AgaTx VIA(P type channel blocker), it was identified that L, N, and P type $Ca^{2+}$ channel exist in rat adrenal chromaffin cells and the order of contribution of each channel type to whole cell $Ca^{2+}$ current was L type> N type> P type. type> P type.

• Restorative Dentistry and Endodontics
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• v.27 no.3
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• pp.215-231
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• 2002

삼차신경절의 뉴론이 구강악안면영역에서의 촉각, 압각, 온도각 및 통각 등 다양한 감각을 중추신경계로 전달하는 역할을 하는 것은 주지의 사실이다. 이러한 신경전달에 있어서 이온통로는 감각정보를 전달하는데 핵심적인 역할을 수행한다. 이 중 소디움 통로는 활동전위의 발생에 중요하며, 칼슘 통로는 시냅스 전도에 있어서 필수적인 역할을 수행하고, 포타슘 통로는 안정막전압의 유지 및 재분극에 관여한다. 최근에 여러 가지의 이온통로들의 뇌조직내의 분포에 관한 연구가 시작되고 있는데 삼차신경의 일차구심뉴론이 종지하는 삼차신경핵 즉 삼차신경 척수감각핵, 삼차신경 주감각핵, 삼차신경 중뇌핵 및 삼차신경 운동핵에 존재하는 이온통로에 관한 연구는 매우 희소하여 본 연구에서는 횐쥐의 삼차신경 핵에 존재하는 소디움, 칼슘 및 포타슘 이온통로들을 면역조직화학적 방법으로 조사하여 다음과 같은 결과를 얻었다. (1) 소디움 통로는 삼차신경 척수감각핵, 삼차신경 주감각핵 및 삼차신경 운동핵 모두에서 강하게 염색되었다. (2) 칼슘 통로는 삼차신경 척수감각핵에서는 N-type 통로가 중등도로 염색되었으며 , P/Q-type 통로는 약하게 염색되었으나 R-type 통로는 거의 염색되지 않았다. 삼차신경 주감각핵에서는 P/Q-type 통로가 매우 약하게 염색되었다. (3) 포타슘 통로는 삼차신경 척수감각핵과 삼차신경 주감각핵에서 inwardly rectifying 포타슘 통로(Kir 2.1)가 중등도로 염색되었고, voltage-gated 포타슘 통로(Kv 4.2)가 약하게 염색되었으며, BKCa는 그 염색 정도가 매우 약하게 나타났다. 이상의 결과를 종합해 볼 때 삼차신경 감각핵에는 소디움 통로의 분포가 가장 많았으며, 칼슘통로에서는 N-type이, 포타슘 통로 중에는 inwardly rectifying 통로(Kir 2.1)가 가장 많이 분포함을 관찰할 수 있었다.

• The Zoological Society Korea : Newsletter
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• v.18 no.2
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• pp.38-44
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• 2001

전위활성화 칼슘통로를 통한 칼슘이온의 세포 내 유입은 근육수축, 시냅스 전달, 호르몬 분비, 효소의 활성도 및 유전자 발현을 조절한다. 이와 같이 중요한 생리적 기능을 조절하기 때문에 칼슘통로를 대상으로 한 다방면의 연구가 과거 20년간 활발히 진행되어 왔다 칼슘통로는 $\alpha$1, $\alpha$2-$\delta$, $\beta$로 구성되어 있으며, 이 중 $\alpha$1은 칼슘통로의 일반적 특성을 나타내는 기본 구조체이며, $\alpha$2-$\delta$와 $\beta$는 $\alpha$1을 조절하는 보조 기능을 한다. 지금까지 10개의 $\alpha$1 subunits(L-형: $\alpha$1S, $\alpha$1C, $\alpha$1D, $\alpha$1F; non-L-형: $\alpha$1A, a1B, $\alpha$1E; T-형: $\alpha$1G, $\alpha$1H, $\alpha$1I), 4종류의 $\beta$ subunits, 3 종류의 $\alpha$2-$\delta$ subunits가 클로닝되었으며, 이들 클론을 이용한 분자 수준에서의 연구가 활발히 이루어지고 있다. 본 논단에서는 칼슘통로의 구조, 기능 및 조절에 대한 연구가 전기생리학적, 분자생물학적 및 약리학적 방법을 사용하여 어떻게 수행되어왔는지 살펴보고, 최근 연구성과에 대해서도 소개하고자 한다.

• 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.

• Journal of Embryo Transfer
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• v.21 no.1
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• pp.35-43
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• 2006

Ions play important roles in various cellular processes including fertilization and differentiation. However, it is little known whether how ions are regulated during early embryonic development in mammalian animals. In this study, we examined changes in $Ca^{2+}\;and\;K^+$ concentrations in embryos and oviduct during mouse early embryonic development using patch clamp technique and confocal laser scanning microscopy. The intracellular calcium concentration in each stage embryos did not markedly change. At 56h afier hCG injection when 8-cell embryos could be Isolated from oviduct, $K^+$ concentration in oviduct increased by 26% compared with that at 14h after injection of hCG During early embryonic development, membrane potential was depolarized (from -38 mV to -16 mV), and $Ca^{2+}$ currents decreased, indicating that some $K^+$ channel might control membrane potential in oocytes. To record the changes in membrane potential induced by influx of $Ca^{2+}$ in mouse oocytes, we applied 5 mM $Ca^{2+}$ to the bath solution. The membrane potential transiently hyperpolarized and then recovered. In order to classify $K^+$ channels that cause hyperpolarization, we first applied TEA and apamin, general $K^+$ channel blockers, to the bath solution. Interestingly, the hyperpolarization of membrane potential still appeared in oocytes pretreated with TEA and apamin. This result suggest that the $K^+$ channel that induces hyperpolarization could belong to another $K^+$ channel such as two-pore domain $K^+(K_{2P})$channel that a.e insensitive to TEA and apamin. From these results, we suggest that the changes in $Ca^{2+}\;and\;K^+$ concentrations play a critical role in cell proliferation, differentiation and reproduction as well as early embryonic development, and $K_{2P}$ channels could be involved in regulation of membrane potential in ovulated oocytes.

• Progress in Medical Physics
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• v.17 no.2
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• pp.96-104
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• 2006

Experiments from several groups Including ours have demonstrated that $Ca^{2+}$ can enhance the inactivation of N-type calcium channels. However, it is not clear if this effect can be ascribed to a 'classic' $Ca^{2+}$-dependent inactivation (CDI) mechanism. One method that has been used to demonstrate CDI of L-type calcium channels is to alter the intracellular and extracellular concentration of $Ca^{2+}$. In this paper we replaced the external divalent cation to monovalent ion ($MA^+$) to test CDI. In the previous paper, we could separate fast (${\tau}{\sim}150ms$) and slow (${\tau}{\sim}2,500ms$) components of inactivation in both $Ba^{2+}$ and $Ca^{2+}$ using 5-sec voltage step. Lowering the external divalent cation concentration to zero abolished fast inactivation with relatively little effect on slow inactivation. Slow inactivation ${\tau}$ correspond very well with provided the $MA^+$ data is shifted 10 mV hyperpolarized and slow inactivation ${\tau}$ decreases with depolarization voltage in both $MA^+\;and\;Ba^{2+}$, which consistent with a classical voltage dependent inactivation (VDI) mechanism. These results combined with those of our previous paper lead us to hypothesize that external divalent cations are required to produce fast N-channel inactivation and this divalent cation-dependent inactivation is a different mechanism from classic CDI or VDI.

• Restorative Dentistry and Endodontics
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• v.27 no.5
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• pp.451-462
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• 2002

삼차신경은 구강악안면영역의 운동 및 감각을 담당하고 있으므로 치과임상에서 매우 중요하다. 삼차신경근 중 삼차신경절에 세포체를 갖는 뉴론은 주로 체성 감각을 전달하는 1차 구심신경으로 악안면영역의 촉각, 압각, 진동감각 온도각 및 통각을 담당한다. 이러한 감각의 전달은 기본적으로 신경세포의 이온통로의 활동에 의존하는데 삼차신경절 세포에 여러 종류의 이온통로가 존재하는 것으로 알려져 있다. 본 연구에서는 항체 염색법을 이용하여 이온통로가 존재를 확인 하고자 한다. 횐쥐의 삼차신경절로부터 통법에 따라 뉴론을 단일 세포로 분리하고 immunocytochemistry 방법으로 세포를 염색하여 관찰한 바 다음과 같은 결과를 얻었다. 본 실험에서 이온전류의 측정 등으로 관찰된 여러 종류의 이온통로들을 면역 염색법으로 확인하였다. 횐쥐의 삼차신경절 뉴론에서 확인된 이온통로는 소디움통로와 N, P 및 Q-type의 칼슘통로 그리고 BK$_{Ca}$, Kv 4.2 및 Kir 2.1 등의 포타슘통로이었으며 이온통로의 종류에 따라 분포에 차이를 나타내었다.

• Progress in Medical Physics
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• v.11 no.1
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• pp.29-38
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• 2000

The signal transduction pathway for most neurotransmitter induced inhibition of $Ca^{2+}$ channels in sympathetic neurons involves a G-protein mediated, membrane-delimited mechanism without the participation of any known protein kinase. However, activation of protein kinase C (PKC) has been proposed as one of the intracellular mechanisms mediating some neurotransmitter induced $Ca^{2+}$ channel inhibition. In the present study, we investigated the effects of phorbol-12, 13-dibutyrate (PDBu) on $Ca^{2+}$ channel currents of acutely dispersed neurons from adult rat superior cervical ganglion (SCG) neurons using whole cell variant of the patch clamp technique. PDBu (500 nM), the activator of PKC, increased $Ca^{2+}$ channel currents and retarded the deactivation of tail currents. The effects of PDBu were voltage dependent and the maximal increase in the current amplitudes was observed between -10 to 10 mV (n=4). PDBu attenuated $Ca^{2+}$ current inhibition induced by norepinephrine (NE), which modulates $Ca^{2+}$ channels via a pertussis toxin (PTX)-sensitive pathway. Inhibition of PDBu by staurosporine (1 $\mu$M) blocked the effects of PDBu on current amplitudes and NE-induced G-protein mediated inhibition of $Ca^{2+}$ currents. Further experiment should be done to know if G-protein or $Ca^{2+}$ channel itself is the target of PKC phosphorvlation.phosphorvlation.

• Progress in Medical Physics
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• v.25 no.3
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• pp.157-166
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• 2014

ATP in quantity co-stored with neurotransmitters in the secretory vesicles of neurons, by being co-released with the neurotransmitters, takes an important role to modulate the stimulus-secretion response of neurotransmitters. Here, in this study, the modulatory effect of ATP was studied in $Ca^{2+}$ channels of cultured rat adrenal chromaffin cells to investigate the physiological role of ATP in neurons. The $Ca^{2+}$ channel current was recorded in a whole-cell patch clamp configuration, which was modulated by ATP. In 10 mM $Ba^{2+}$ bath solution, ATP treatment (0.1 mM) decreased the $Ba^{2+}$ current by an average of $36{\pm}6%$ (n=8), showing a dose-dependency within the range of $10^{-4}{\sim}10^{-1}mM$. The current was recovered by ATP washout, demonstrating its reversible pattern. This current blockade effect of ATP was disinhibited by a large prepulse up to +80 mV, since the $Ba^{2+}$ current increment was larger when treated with ATP ($37{\pm}5%$, n=11) compared to the control ($25{\pm}3%$, n=12, without ATP). The $Ba^{2+}$ current was recorded with $GTP{\gamma}S$, the non-hydrolyzable GTP analogue, to determine if the blocking effect of ATP was mediated by G-protein. The $Ba^{2+}$ current decreased down to 45% of control with $GTP{\gamma}S$. With a large prepulse (+80 mV), the current increment was $34{\pm}4%$ (n=19), which $25{\pm}3%$ (n=12) under control condition (without $GTP{\gamma}S$). The $Ba^{2+}$ current waveform was well fitted to a single-exponential curve for the control, while a double-exponential curve best fitted the current signal with ATP or $GTP{\gamma}S$. In other words, a slow activation component appeared with ATP or $GTP{\gamma}S$, which suggested that both ATP and $GTP{\gamma}S$ caused slower activation of $Ca^{2+}$ channels via the same mechanism. The results suggest that ATP may block the $Ca^{2+}$ channels by G-protein and this $Ca^{2+}$ channel blocking effect of ATP is important in autocrine (or paracrine) inhibition of adrenaline secretion in chromaffin cell.

• Journal of Life Science
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• v.25 no.2
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• pp.231-236
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• 2015

Angiotensin II (AngII) is an essential hormone that affects vascular physiology. For example, stimulation of vascular smooth muscle cells (VSMCs) rapidly induces vasoconstriction and results in the up-regulation of blood pressure. Chronic stimulation of VSMCs with AngII also results in hypertrophy. In this study, we confirmed an involvement of phosphatidylinositol 3-kinase (PI3K)-dependent calcium mobilization in AngII-induced generation of reactive oxygen species (ROS). Stimulation of rat aortic smooth muscle cells (RASMCs) with AngII significantly induced the generation of ROS in a dose- and time-dependent manner. AngII-induced generation of ROS was completely abolished by pharmacological inhibition of PI3K (with LY294002), but inhibition of the ERK signaling pathway had no effect. AngII-induced calcium mobilization was completely blocked by inhibition of PI3K, whereas inhibition of the ERK signaling pathway by PD98059 was ineffective. Depletion of extracellular calcium or inhibition of the L-type calcium channel by nifedipine completely blocked AngII-induced calcium mobilization. Depletion of extracellular calcium by EGTA and incubation of RASMCs with calcium-free medium both significantly blocked AngII-induced ROS generation. Inhibition of the L-type calcium channel also significantly blocked AngII-induced ROS generation. These results suggest that AngII-induced ROS generation is regulated by calcium mobilization, which, in turn, is modulated by a PI3K/L-type calcium channel signaling pathway.