• 대한수의학회지
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• 제39권4호
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• pp.702-709
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• 1999

The present study was performed to elucidate the effects of extracellular $Ca^{2+}$ on contractile responses in isolated porcine coronary artery ring using by perivascular nerve stimulation (PNS). Especially, the study was focused on the source of $Ca^{2+}$ on $P_{2X}$-purinoceptor mediated muscle contraction which one of $P_2$-purinoceptor subtypes. The following results can be drawn from these studies : 1. The phasic contractions induced by PNS were inhibited with muscarinic receptor antagonist, atropine ($10^{-6}M$). 2. The phasic contractions induced by PNS were significantly inhibited by sequential treatment with atropine and adrenergic neural blocker, guanethidine ($10^{-6}M$). 3. The phasic contractions induced by PNS were inhibited with $P_{2X}$-purinoceptor desensitization by repetitive application of $\alpha$,$\beta$-Me ATP ($10^{-4}M$). 4. The phasic contractions induced by PNS were so weakened in calcium-free medium. 5. The phasic contractions induced by PNS were inhibited with calcium channel blocker, verapamil ($10^{-6}{\sim}5{\times}10^{-6}M$). 6. The phasic contractions induced by PNS on pretreated with verapamil ($10^{-6}{\sim}5{\times}10^{-6}M$) were not changed by $\alpha$,$\beta$-Me ATP ($10^{-4}M$). These results demonstrate that the neurogenic phasic contractions induced by PNS are due to adrenergic-, cholinergic- and $P_{2X}$-purinergic receptors and the origin of $Ca^{2+}$ on $P_{2X}$-purinoceptor mediated muscle contraction is extracellular $Ca^{2+}$ through plasmalemmal $Ca^{2+}$ channels.

• Toxicological Research
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• 제35권3호
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• pp.279-285
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• 2019

In this study, we investigated the therapeutic potential of Cinnamomum camphora leaves on allergic skin inflammation such as atopic dermatitis. We evaluated the effects of C. camphora leaves on human adult low-calcium high-temperature keratinocytes and atopic dermatitis mice. C. camphora leaves inhibited Macrophage-derived chemokine (an inflammatory chemokine) production in $interferon-{\gamma}$ (10 ng/mL) stimulated Human adult low-calcium high-temperature keratinocytes in a dose dependent manner. C. camphora leaves suppressed the phosphorylation of janus kinase signal transducer and activator of transcription 1. C. camphora leaves also suppressed the phosphorylation of extracellular signal-regulated kinase 1/2, a central signaling molecule in the inflammation process. These results suggest that C. camphora leaves exhibits anti-inflammatory effect via the phosphorylation of signal transducer and activator of transcription 1 and extracellular signal-regulated kinase 1/2. To study the advanced effects of C. camphora leaves on atopic dermatitis, we induced experimental atopic dermatitis in mice by applying 2,4-dinitrochlorobenzene. The group treated with C. camphora leaves (100 mg/kg) showed remarkable improvement of atopic dermatitis symptoms: reduced serum immunoglobulin E levels, smaller lymph nodes with reduced thickness and length, decreased ear edema, and reduced levels of inflammatory cell infiltration in the ears. Interestingly, the effects of C. camphora leaves on atopic dermatitis symptoms were stronger than those of hydrocort cream, a positive control. Taken together, C. camphora leaves showed alleviating effects on the inflammatory chemokine production in vitro and atopic dermatitis symptoms in vivo. These results suggest that C. camphora leaves help in the treatment of allergic inflammation such as atopic dermatitis.

• 대한약리학회지
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• 제32권1호
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• pp.67-74
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• 1996

고양이 담낭근에서 효소학적으로 분리한 평활근 세포는 cholecystokinin octapeptide (CCK-8), acetylcholine (ACh) 및 KCl에 의하여 용량에 의존하여 수축하였다. 이들 효현제 (CCK-5, ACh 및 KCl)에 의한 평활근 세포의 최대수축은 각각$10^{-9}M$, $10^{-5}M$ 및 20mM 농도에서 야기되었다. CCK-8에 의하여 야기되는 이들 평활근 세포의 수축은 HEPES 완충액에 $Ca^{2+}$을 제거시킴에 의하여 영향을 받지 아니하였으나, $Ca^{2+}$ 대신에 strontium을 첨가시켰을때 수축반응이 완전하게 억제되었다 (p<0.001). 이와는 반대로 KCl에 의한 수축반응은 strontium 치환에 의하여 영향을 받지 아니하고 HEPES 완충액에 $Ca^{2+}$을 제거시킴에 의하여 억제되었다 (p<0.01). ACh에 의하여 야기되는 수축반응은 세포 외액의 $Ca^{2+}$을 제거시킴에 의하여 중등도의 억제반응이 야기되었으나 (p<0.05) strontium에 의하여 영향을 받지 아니하였다. Saponin으로 세포 투과성 변동을 야기시킨 근세포에서 inositol 1,4,5-trisphosphate $(IP_3)$와 CCK-8은 수축반응을 일으켰고, 이러한 수축반응은 calmodulin 길항제인 CGS 9343B에 의하여 차단되었으며 (p<0.001), heparin은 CCK-8 및 $IP_3$의 작용을 완전하게 봉쇄하였다 (p<0.001). 그러나 이러한 수축반응에 있어서 protein kinase C 길항제인 H7은 아무런 작용을 나타내지 못하였다. 이러한 결과로 볼 때 CCK-8에 의하여 야기된 고양이 담낭근 세포의 수축반응은 $IP_3$에 의하여 세포내 저장소로부터 유리된 $Ca^{2+}$과 calmodulin에 의존적인 과정에 의하여 매개되어 지는 것으로 생각된다. 또한 ACh는 세포외액의 $Ca^{2+}$ 뿐만 아니라 세포내 저장소의 $Ca^{2+}$ 모두를 이용하며, KCl은 전적으로 세포외액의 $Ca^{2+}$에 의존적인 형태로 calmodulin과는 무관하게 고양이 담낭근 세포의 수축반응을 야기시키는 것으로 사료된다.

• Journal of Plant Biotechnology
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• 제34권3호
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• pp.213-221
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• 2007

분리된 무 자엽 조직에 산화질소 (nitric oxide: NO) 공여체인 sodium nitroprusside (SNP) 처리 시 농도 의존방식으로 부정근의 발달을 증진시켰다. 그러나 이러한 NO 증진 효과는 세포외 칼슘 chelator인 0.5 mM EGTA 또는 세포막 칼슘채널 차단제인 0.1 mM $LaCl_3$를 각각 $50\;{\mu}M$ SNP와 함께 혼합처리 시 반전되었다. 또한, 뿌리 발생에서 중심적 역할을 수행하는 것으로 알려진 guaiacol peroxidase (GPX)와 syringaldazine peroxidase (SPX)의 활성도가 SNP 단독 처리된 자엽에서 부정근이 형성되는 동안 현저히 증가하였다. 그러나, SNP와 $LaCl_3$ 혼합처리 시 SNP에 의해 유도된 GPX와 SPX 활성도 증가가 거의 증류수 대조구 수준으로 억제되었다. calmodulin의 anatagonist인 trifuoperazine 역시 SNP로 처리된 자엽에서 부정근 형성을 억제하여 발생된 뿌리의 개수와 길이를 감소시켰으며 동시에 GPX와 SPX를 불활성화 하였다. 결론적으로, 이들 결과는 칼슘이 GPX와 SPX 활성도 조절을 통해 부정근 유도를 이끄는 NO 반응에 포함되어 있음을 나타내는 것이다.

• 대한수의학회지
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• 제47권3호
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• pp.265-271
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• 2007

This study investigated the protective effects of ethylene glycol-bis(${\beta}$-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular calcium chelator, and ethylenediaminetetraacetic acid (EDTA), which chelates calcium and most metal ions, against carbon tetrachloride ($CCl_4$)-induced acute hepatotoxicity in mice. Mice were treated with EGTA or EDTA at a dose of 20 (low) or 100 mg/kg (high) subcutaneously 1h before $CCl_4$ administration. The mice were fasted and sacrificed 18h after $CCl_4$ treatment. Blood samples were collected from the carotid artery by decapitation under light ether anesthesia. Serum alkaline phosphatase (ALP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), triglyceride (TG), and cholesterol levels were measured. Malondialdehyde (MDA) production was determined as an index of lipid peroxidation in the liver. The liver, kidneys, and spleen were weighed. We also evaluated the histopathological changes in the liver in each group. The relative weights of the liver were significantly higher in the $CCl_4$-treatment group than in the normal group, except in the high-EDTA treatment group. EGTA and EDTA treatment caused a significant decrease in serum ALP, ALT, and AST levels. Of all of the doses of EGTA and EDTA tested, the high-EDTA dose resulted in the most remarkable inhibitory action. The protective effect in the high-EDTA-treatment group was confirmed histopathologically. The low-EGTA-treatment group showed a significant decrease in serum TG and cholesterol levels. Liver MDA levels were significantly decreased in the EGTA (20 mg/kg) and EDTA (20, 100 mg/kg) groups. These results suggest that EDTA, which chelates both calcium and metal ions, confers better protection in $CCl_4$-induced acute liver damage than does EGTA, a calcium chelator.

• Animal cells and systems
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• 제13권4호
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• pp.363-370
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• 2009

Since vitamin $D_3$ is an important regulator of osteoblastic differentiation, a presently-established vitamin $D_3$-entrapped calcium phosphate film (VCPF) was evaluated for hard tissue engineering. The entrapped vitamin $D_3$ more rapidly induced bone nodule formation. To characterize the cellular events leading to regulations including faster differentiation, signal transduction pathways were investigated in osteoblastic MG63 cells at a molecular level. Major signaling pathways for MG63 cell proliferation including phosphatidylinositol-3-kinase, extracellular signal-regulated kinase, c-Jun N-terminal kinase and focal adhesion kinase pathways were markedly down-regulated when cells were cultured on calcium phosphate film (CPF) and VCPF. This agreed with our earlier observations of the immediate delay in proliferation of MG63 cells upon culture on CPF and VCPF. On the other hand, the p38 mitogen-activated protein kinase (p38 MAPK) and protein kinase A (PKA) pathways were significantly up-regulated on both CPF and VCPF. CPF alone could simulate differential behaviors of MG63 cells even in the absence of osteogenic stimulation and entrapment of vitamin $D_3$ within CPF further amplified the signal pathways, resulting in continued promotion of MG63 cell differentiation. Interplay of p38 MAPK and PKA signaling pathways likely is a significant event for the promotion of differentiation and mineralization of MG63 cells.

• The Korean Journal of Physiology
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• 제28권2호
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• pp.151-157
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• 1994

Intracellular free $Ca^{2+}$ contributes to regulation of various events occurring in vascular smooth muscle cells. One of these events is modulating the membrane iou currents. Single smooth muscle cells were isolated from rabbit mesenteric artery. Three kinds of $Ca^{2+}-activated\;current$ were studied with the patch clamp method. $Ca^{2+}-activated\;K^+\;current$ with a large oscillation was recorded in the depolarized potential range. The single channel conductance of this current was about 250 pS. It was abolished by replacing intracellular $K^+\;with\;Cs^+$. A $Ca^{2+}-activated$ nonselective cation current was observed in both the depolarized and hyperpolarized potential ranges. And it was blocked by replacement of extracellular $Na^+$ with N-methylglucamine (NMG) or extracellular application of $Cd^{2+}$. $Ca^{2+}-activated\;Cl^-\;current$ was revealed in the whole voltage range and was blocked by niflumic acid. These results indicate that at least three kinds of $Ca^{2+}-activated$ ionic currents exist in smooth muscle cells from rabbit superior mesenteric artery.

• Journal of Yeungnam Medical Science
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• 제9권2호
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• pp.359-381
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• 1992

Benzodiazepine계 약물들은 진정 최면제의 대표적인 약물로서, 중추신경계에서의 그 작용은 gamma amino butyric acid(GABA) 수용체와 짝지워져 있는 benzodiazepine 수용체를 통해서 나타나며 또한 뇌에 있는 synaptosome에서 전위 의존성 calcium channel을 통한 calcium의 섭취를 억제함으로써 진정작용 및 최면 작용이 나타난다. 이와 아울러 말초 장기에서도 benzodiazepine 수용체와 GABA 수용체가 발견 되었는데 이들의 기능과 상호관계는 잘 알려져 있지 않다. 이에 본 실험에서는 benzodiazepine계통의 대표적인 약물이며 중추신경과 말초 장기에 동시에 작용하는 diazepam이 흰쥐 적출자궁의 자발 수축 및 oxytocin 유발 수축에 미치는 영향을 검색하고, 이러한 diazepam의 효과와 GABA 수용체 및 calcium과의 상호관계를 검색함으로써 그 작용기전을 추구해 보기 위하여 다음과 같은 실험을 하였다. 난소를 제거한 후 estrogen(17 beta-estradiol : $500{\mu}g/kg/day$)을 4일 동안 전 처치한 흰쥐의 자궁을 적출하여 등척성 장력을 측정함으로써 그 수축력의 변화를 관찰하였다. Diazepam과 GABA 수용체 효현제 및 그 봉쇄제들이 자궁절편의 자발 수축과 oxytocin 유발 수축에 미치는 영향을 검색하였고, 또 이들 약물의 작용에 관련된 calcium 동원기전에 대하여 관찰하여 다음과 같은 결과를 얻었다. Diazepam은 흰쥐 적출자궁의 자발수축 및 oxytocin 유발수축을 농도 의존적으로 억제하였다. GABA, GABA A 수용체 효현제인 muscimol, GAGA A 수용체의 상경적 봉쇄제인 bicuculline, GABA A 수용체의 비상경적 봉쇄제인 picrotoxin, GABA B 수용체 효현제인 boclofen, 그리고 GABA B 수용체 봉쇄제인 delta-aminovaleric acid는 흰쥐 적출 자궁의 자발 수축 및 oxytocin 유발수축에 아무런 영향을 미치지 않았다. 자발 수축 및 oxytocin 유발수축에 대한 diazepam의 억제 작용은 GABA 수용체 효현제 및 봉쇄제의 영향을 받지 않았다. 그러나 bicuculline은 diazepam의 억제 작용에 상가적으로 작용하였는데, bicuculline의 이러한 작용은 muscimol에 의해서 길항되지 않았다. 정상 PSS 내에서 diazepam에 의해 억제되었던 자발수욱 및 oxytocin유발수촉은 calcium의 첨가 및 calcium inophore인 A23187의 첨가로 일부 회복되었다. Calcium 배제 용액내에서는 diazepam이 calcium 첨가로 인한 수축력 회복을 방해하였으며 calcium inophore인 A23187에 의한 수축력 증가는 막지 못하였다. 또 세포외액에 calcium이 결핍된 상태에서는 oxytocin 자체에 의한 수축을 방해하지 못하였으나 이어 첨가된 calcium에 의한 oxytocin 유발 수축의 증가는 일부 억제하였다. 이상의 실험결과로 미루어 볼 때 diazepam은 자궁의 자발수측 및 oxytocin 유발 수축을 억제할 수 있으며, 이러한 작용은 GABA 수용체 의존성이 아닌 세포외액의 calcium의 유입을 억제함으로써 나타나는 것으로 사료된다.

• The Korean Journal of Physiology and Pharmacology
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• 제21권2호
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• pp.233-239
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• 2017

Intracellular calcium ($Ca^{2+}$) oscillation is an initial event in digestive enzyme secretion of pancreatic acinar cells. Reactive oxygen species are known to be associated with a variety of oxidative stress-induced cellular disorders including pancreatitis. In this study, we investigated the effect of hydrogen peroxide ($H_2O_2$) on intracellular $Ca^{2+}$ accumulation in mouse pancreatic acinar cells. Perfusion of $H_2O_2$ at $300{\mu}M$ resulted in additional elevation of intracellular $Ca^{2+}$ levels and termination of oscillatory $Ca^{2+}$ signals induced by carbamylcholine (CCh) in the presence of normal extracellular $Ca^{2+}$. Antioxidants, catalase or DTT, completely prevented $H_2O_2$-induced additional $Ca^{2+}$ increase and termination of $Ca^{2+}$ oscillation. In $Ca^{2+}$-free medium, $H_2O_2$ still enhanced CCh-induced intracellular $Ca^{2+}$ levels and thapsigargin (TG) mimicked $H_2O_2$-induced cytosolic $Ca^{2+}$ increase. Furthermore, $H_2O_2$-induced elevation of intracellular $Ca^{2+}$ levels was abolished under sarco/endoplasmic reticulum $Ca^{2+}$ ATPase-inactivated condition by TG pretreatment with CCh. $H_2O_2$ at $300{\mu}M$ failed to affect store-operated $Ca^{2+}$ entry or $Ca^{2+}$ extrusion through plasma membrane. Additionally, ruthenium red, a mitochondrial $Ca^{2+}$ uniporter blocker, failed to attenuate $H_2O_2$-induced intracellular $Ca^{2+}$ elevation. These results provide evidence that excessive generation of $H_2O_2$ in pathological conditions could accumulate intracellular $Ca^{2+}$ by attenuating refilling of internal $Ca^{2+}$ stores rather than by inhibiting $Ca^{2+}$ extrusion to extracellular fluid or enhancing $Ca^{2+}$ mobilization from extracellular medium in mouse pancreatic acinar cells.

• The Korean Journal of Physiology and Pharmacology
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• 제22권2호
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• pp.215-223
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• 2018

Intracellular $Ca^{2+}$ mobilization is closely linked with the initiation of salivary secretion in parotid acinar cells. Reactive oxygen species (ROS) are known to be related to a variety of oxidative stress-induced cellular disorders and believed to be involved in salivary impairments. In this study, we investigated the underlying mechanism of hydrogen peroxide ($H_2O_2$) on cytosolic $Ca^{2+}$ accumulation in mouse parotid acinar cells. Intracellular $Ca^{2+}$ levels were slowly elevated when $1mM\;H_2O_2$ was perfused in the presence of normal extracellular $Ca^{2+}$. In a $Ca^{2+}-free$ medium, $1mM\;H_2O_2$ still enhanced the intracellular $Ca^{2+}$ level. $Ca^{2+}$ entry tested using manganese quenching technique was not affected by perfusion of $1mM\;H_2O_2$. On the other hand, $10mM\;H_2O_2$ induced more rapid $Ca^{2+}$ accumulation and facilitated $Ca^{2+}$ entry from extracellular fluid. $Ca^{2+}$ refill into intracellular $Ca^{2+}$ store and inositol 1,4,5-trisphosphate ($1{\mu}M$)-induced $Ca^{2+}$ release from $Ca^{2+}$ store was not affected by $1mM\;H_2O_2$ in permeabilized cells. $Ca^{2+}$ efflux through plasma membrane $Ca^{2+}-ATPase$ (PMCA) was markedly blocked by $1mM\;H_2O_2$ in thapsigargin-treated intact acinar cells. Antioxidants, either catalase or dithiothreitol, completely protected $H_2O_2-induced$ $Ca^{2+}$ accumulation through PMCA inactivation. From the above results, we suggest that excessive production of $H_2O_2$ under pathological conditions may lead to cytosolic $Ca^{2+}$ accumulation and that the primary mechanism of $H_2O_2-induced$ $Ca^{2+}$ accumulation is likely to inhibit $Ca^{2+}$ efflux through PMCA rather than mobilize $Ca^{2+}$ ions from extracellular medium or intracellular stores in mouse parotid acinar cells.