• 동의생리병리학회지
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• 제16권2호
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• pp.389-396
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• 2002

In the present work, we examined the mechanism of vasorelaxant effect of scopoletin, an active constituent of Artemisia capillaris on rat thoracic descending aortic rings. Scopoletin induced a concentration-dependent relaxation in rat thoracic descending aortic rings pre-contracted with phenylephrine (EC/sub 50/ = 238.94±37.4 μM), while it was less effective in rat thoracic descending aortic rings precontracted with high potassium solution (KCI 30 mM). Vasorelaxation by scopoletin was significantly inhibited after endothelial removal, but recovered at high concentration. Pretreatment of rat thoracic descending aortic rings with N/sup G/-nitro-L-arginine (100 μM), a nitric oxide synthase inhibitor, and atropine (1 μM), a muscarinic receptor antagonist, significantly inhibited scopoletin-induced relaxation of rat thoracic descending aortic rings. Neither indomethacin (3 μM), an inhibitor of cydooxygenase, nor propranolol (1 μM), a β -adrenoceptor antagonist, modified the effect of scopoletin. The combination of N/sup G/ -nitro-L-arginine (100 μ M) and miconazole (10 μ M), an inhibitor of cytochrome P 450, did not modify the effect of scopoletin, when compared with pretreatment with N/sup G/-nitro-L-arginine(100 μM) alone. Vasorelaxant effect of scopoletin was inverted by pretreatment with diltiazem (10 μM), a Ca/sup 2+/-channel blocker, at low concentration, while restored at high concentration. Apamin (K/sub ca/-channel blocker, 1 μM), 4-aminopyridine (4-AP, K/sub v/-channel blocker, 1 mM), and tetrodotoxin (TTX, Na/sup +/-channel blocker 1 μM) potentiated the vasorelaxant effect of scopoledn, but glibendamide (K/sub ATP/-channel blocker, 10 μM), tetraetylammonium(TEA, non-selective K-channel blocker, 10 mM) did not affect the relaxation of scopoletin. Free radical scavengers (TEMPO, catalase, mannitol) did not modify vascular tone. These results suggest that nitric oxide, Ca/sup 2+/ -channels play a role in endothelium-dependent relaxations to scopoletin in rat aortas, that apamin, 4-AP, TTX but not glibenclamide, TEA potentiated relaxation to scopoletin mediated by these channels, and that free radicals do not concern to the vasorelaxant effect of scopoletin.

• The Korean Journal of Physiology and Pharmacology
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• 제12권2호
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• pp.73-77
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• 2008

Recent studies have documented that testosterone relaxes several smooth muscles by modulating $K^+$ channel activities. Smooth muscles of seminal vesicles playa fundamental role in ejaculation, which might involve testosterone. This study was aimed to assess the role of testosterone in seminal vesicular motility by studying its effects on contractile agents and on the ion channels of single vesicular myocytes in a rabbit model. The contractile responses of circular smooth muscle strips of rabbit seminal vesicles to norepinephrine ($10{\mu}M$), a high concentration of KCI (70 mM), and testosterone ($10{\mu}M$) were observed. Single vesicular myocytes of rabbit were isolated using proteolytic enzymes including collagenase and papain. Inside-out, attached, and whole-cell configurations were examined using the patch clamp technique. The applications of $10{\mu}M$ norepinephrine or 70 mM KCl induced tonic contractions, and $10{\mu}M$ testosterone (pharmacological concentration) evoked dose-dependent relaxations of these precontracted strips. Various $K^+$ channel blockers, such as tetraethylammonium (TEA; $10{\mu}M$), iberiotoxin ($0.1{\mu}M$), 4-aminopyridine (4-AP, $10{\mu}M$), or glibenclamide ($10{\mu}M$) rarely affected these relaxations. Single channel data (of inside-out and attached configurations) of BK channel activity were also hardly affected by testosterone ($10{\mu}M$). On the other hand, however, testosterone reduced L-type $Ca^{2+}$ currents significantly, and found to induce acute relaxation of seminal vesicular smooth muscle and this was mediated, at least in part, by $Ca^{2+}$ current inhibition in rabbit.

• The Korean Journal of Physiology and Pharmacology
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• 제1권6호
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• pp.783-796
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• 1997

The relaxation induced by stimulation of the inhibitory non-adrenergic, non-cholinergic (iNANC) nerve is mediated by the release of iNANC neurotransmitters such as nitric oxide (NO), vasoactive intestinal peptide (VIP) and adenosine triphosphate (ATP). The mechanisms of NO, VIP or ATP-induced relaxation have been partly determined in previous studies, but the detailed mechanism remains unknown. We tried to identify the nature of iNANC neurotransmitters in the smooth muscle of guinea pig ileum and to determine the mechanism of the inhibitory effect of nitric oxide. We measured the effect of NO-donors VIP and ATP on the intracellular $Ca^{2+}$ concentration$([Ca^{2+}]_i)$, by means of a fluorescence dye(fura 2) and tension simultaneously in the isolated guinea pig ileal smooth muscle. Following are the results obtained. 1. Sodium nitroprusside $(SNP:10^{-5}\;M)$ or S -nitro-N-acetyl-penicillamine $(SNP:10^{-5}\;M)$ decreased resting $[Ca^{2+}]_i$ I and tension of muscle. SNP or SNAP also inhibited rhythmic oscillation of $[Ca^{2+}]_i$ and tension. In 40mM $K^+$ solution or carbachol ($(CCh:10^{-6}\;M)$-induced precontracted muscle, SNP decreased muscle tension. VIP did not change $[Ca^{2+}]_i$ and tension in the resting or precontracted muscle, but ATP increased resting $[Ca^{2+}]_i$ and tension in the resting muscle. 2. 1H-[1,2,4]oxadiazol(4,3-a)quinoxalin-1-one $(ODQ:1\;{\mu}M)$, a specific inhibitor of soluble guanylate cyclase, limited the inhibitory effect of SNP 3. Glibenclamide $(10\;{\mu}M)$, a blocker of $K_{ATP}$ channel, and 4-aminopyridine (4-AP:5 mM), a blocker of delayed rectifier K channel, apamin $(0.1\;{\mu}M)$, a blocker of small conductance $K_{Ca}$ channel had no effect on the inhibitory effect of SNP. Iberiotoxin $(0.1\;{\mu}M)$, a blocker of large conductance $K_{Ca}$ channel, significantly increased the resting $[Ca^{2+}]_i$, and tension, and limited the inhibitory effect of SNP. 4. Nifedipine $(1\;{\mu}M)$ or elimination of external $Ca^{2+}$ decreased not only resting $[Ca^{2+}]_i$ and tension but also oscillation of $[Ca^{2+}]_i$ and tension. Ryanodine $(5\;{\mu}M)$ and cyclopiazonic acid $(10\;{\mu}M)$ decreased oscillation of $[Ca^{2+}]_i$ and tension. 5. SNP decreased $Ca^{2+}$ sensitivity of contractile protein. In conclusion, these results suggest that 1) NO is an inhibitory neurotransmitter in the guinea pig ileum, 2) the inhibitory effect of SNP on the $[Ca^{2+}]_i$ and tension of the muscle is due to a decrease in $[Ca^{2+}]_i$ by activation of the large conductance $K_{Ca}$ channel and a decrease in the sensitivity of contractile elements to $Ca^{2+}$ through activation of G-kinase.

• 대한본초학회지
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• 제25권4호
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• pp.17-21
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• 2010

Objectives : The aim of this study was to evaluate the differential mechnism of vasodilation of alcohol steamed Rhei Tangutici Radix et Rhizoma. (ART) and Rhei Tangutici Radix et Rhizoma. (RT) in rat thoracic aorta. Methods : Rat aortic ring preparations were mounted in organ baths with oxygenated (95% $O_2$-5% $CO_2$) Krebs-Ringer bicarbonate solutions at $37{\pm}0.5^{\circ}C$ and subjected to contractions or relaxations. Results : ART exerted vasorelaxation on phenylephrine(PE)-induced contraction in a dose dependent manner. Vasorelaxation effects of ART and RT were endothelium-independent. In the $Ca^{2+}$-free high KCl (60 mM) baths, the contraction of aortic rings induced by accumulative addition of $Ca^{2+}$ (0.3-10.0 mM) was significantly reduced by pre-treatment with both ART and RT for 10 min. The magnitude of vasodilatation was biggerin ART. Moreover, verapamil ($0.001{\mu}M$) and diltiazem ($10{\mu}M$), voltage operative $Ca^{2+}$channel blockers, attenuated the relaxation effect of ART but not that of RT. In the absence of extracellular $Ca^{2+}$, pre-incubation of the aortic rings with RT ($1.0mg/m{\ell}$) significantly reduced the contraction caused by PE but not that of ART. $K^+$ channel inhibitors such as glibenclamide (Gli, $10^{-5}M$), tetraethylammonium (TEA, 1 mM) and 4-aminopyridine (4-AP, 0.2 mM) significantly reduced the ART's vasorelaxation efficacy, but not that of RT. However, the relaxation effects of ART and RT were not inhibited by pre-treatment with indomethacin ($10^{-5}M$), and atropine ($10^{-6}M$). Conclusions : These results suggest that the endothelium-independent relaxation is due to inhibition of $Ca^{2+}$ influx via the suppression of $Ca^{2+}$ release from intracelluar store in RT but via both voltage operative $Ca^{2+}$channel blockage and $K^+$ channel activation in ART.

• The Korean Journal of Physiology and Pharmacology
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• 제16권1호
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• pp.37-42
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• 2012

The aim of the present study was to elucidate the direct effects of melatonin on bladder activity and to determine the mechanisms responsible for the detrusor activity of melatonin in the isolated rat bladder. We evaluated the effects of melatonin on the contractions induced by phenylephrine (PE), acetylcholine (ACh), bethanechol (BCh), KCl, and electrical field stimulation (EFS) in 20 detrusor smooth muscle samples from Sprague-Dawley rats. To determine the mechanisms underlying the inhibitory responses to melatonin, melatonin-pretreated muscle strips were exposed to a calcium channel antagonist (verapamil), three potassium channel blockers [tetraethyl ammonium (TEA), 4-aminopyridine (4-AP), and glibenclamide], a direct voltage-dependent calcium channel opener (Bay K 8644), and a specific calcium/calmodulin-dependent kinase II (CaMKII) inhibitor (KN-93). Melatonin pretreatment ($10^{-8}{\sim}10^{-6}M$) decreased the contractile responses induced by PE ($10^{-9}{\sim}10^{-4}M$) and Ach ($10^{-9}{\sim}10^{-4}M$) in a dose-dependent manner. Melatonin ($10^{-7}M$) also blocked contraction induced by high KCl ($[KCl]_{ECF}$; 35 mM, 70 mM, 105 mM, and 140 mM) and EFS. Melatonin ($10^{-7}M$) potentiated the relaxation response of the strips by verapamil, but other potassium channel blockers did not change melatonin activity. Melatonin pretreatment significantly decreased contractile responses induced by Bay K 8644 ($10^{-11}{\sim}10^{-7}M$). KN-93 enhanced melatonin-induced relaxation. The present results suggest that melatonin can inhibit bladder smooth muscle contraction through a voltage-dependent, calcium-antagonistic mechanism and through the inhibition of the calmodulin/CaMKII system.

• The Korean Journal of Physiology and Pharmacology
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• 제15권6호
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• pp.405-413
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• 2011

This study was designed to elucidate high-$K^+$ induced response of circular and longitudinal smooth muscle from human gastric corpus using isometric contraction. Contraction from circular and longitudinal muscle stripes of gastric corpus greater curvature and lesser curvature were compared. Circular smooth muscle from corpus greater curvature showed high $K^+$ (50 mM)-induced tonic contraction. On the contrary, however, longitudinal smooth muscle strips showed high $K^+$ (50 mM)-induced sustained relaxation. To find out the reason for the discrepancy we tested several relaxation mechanisms. Protein kinase blockers like KT5720, PKA inhibitor, and KT5823, PKG inhibitor, did not affect high $K^+$-induced relaxation. $K^+$ channel blockers like tetraethylammonium (TEA), apamin (APA), glibenclamide (Glib) and barium ($Ba^{2+}$) also had no effect. However, N(G)-nitro-L-arginine (L-NNA) and 1H-(1,2,4) oxadiazolo (4,3-A) quinoxalin-1-one (ODQ), an inhibitor of soluble guanylate cyclase (sGC) and 4-AP (4-aminopyridine), voltage-dependent $K^+$ channel (KV) blocker, inhibited high $K^+$ -induced relaxation, hence reversing to tonic contraction. High $K^+$-induced relaxation was observed in gastric corpus of human stomach, but only in the longitudinal muscles from greater curvature not lesser curvature. L-NNA, ODQ and KV channel blocker sensitive high $K^+$-induced relaxation in longitudinal muscle of higher portion of corpus was also observed. These results suggest that longitudinal smooth muscle from greater curvature of gastric corpus produced high $K^+$-induced relaxation which was activated by NO/sGC pathway and by $K_V$ channel dependent mechanism.

• The Korean Journal of Physiology and Pharmacology
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• 제17권4호
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• pp.359-365
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• 2013

Plasma pH can be altered during pregnancy and at labor. Membrane excitability of smooth muscle including uterine muscle is suppressed by the activation of $K^+$ channels. Because contractility of uterine muscle is regulated by extracellular pH and humoral factors, $K^+$ conductance could be connected to factors regulating uterine contractility during pregnancy. Here, we showed that TASK-2 inhibitors such as quinidine, lidocaine, and extracellular acidosis produced contraction in uterine circular muscle of mouse. Furthermore, contractility was significantly increased in pregnant uterine circular muscle than that of non-pregnant muscle. These patterns were not changed even in the presence of tetraetylammonium (TEA) and 4-aminopyridine (4-AP). Finally, TASK-2 inhibitors induced strong myometrial contraction even in the presence of L-methionine, a known inhibitor of stretch-activated channels in myometrium. When compared to non-pregnant myometrium, pregnant myometrium showed increased immunohistochemical expression of TASK-2. Therefore, TASK-2, seems to play a key role during regulation of myometrial contractility in the pregnancy and provides new insight into preventing preterm delivery.

• The Korean Journal of Physiology and Pharmacology
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• 제3권1호
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• pp.1-10
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• 1999

We sought to find out the mechanism of vascular relaxation by extracellular $K^+$ concentration $([K^+]_o)$ in the cerebral resistant arteriole from rabbit. Single cells were isolated from the cerebral resistant arteriole, and using voltage-clamp technique barium-sensitive $K^+$ currents were recorded, and their characteristics were observed. Afterwards, the changes in membrane potential and currents through the membrane caused by the change in $[K^+]_o$ was observed. In the smooth muscle cells of cerebral resistant arteriole, ion currents that are blocked by barium, 4-aminopyridine (4-AP), and tetraethylammonium (TEA) exist. Currents that were blocked by barium showed inward rectification. When the $[K^+]_o$ were 6, 20, 60, and 140 mM, the reversal potentials were $-82.7{\pm}1.0,\;-49.5{\pm}1.86,\;-26{\pm}1.14,\;-5.18{\pm}1.17$ mV, respectively, and these values were almost identical to the calculated $K^+$ equilibrium potential. The inhibition of barium-sensitive inward currents by barium depended on the membrane potential. At the membrane potentials of -140, -100, and -60 mV, $K_d$ values were 0.44, 1.19, and 4.82 ${\mu}M,$ respectively. When $[K^+]_o$ was elevatedfrom 6 mM to 15 mM, membrane potential hyperpolarized to -50 mV from -40 mV. Hyperpolarization by $K^+$ was inhibited by barium but not by ouabain. When the membrane potential was held at resting membrane potential and the $[K^+]_o$ was elevated from 6 mM to 15 mM, outward currents increased; when elevated to 25 mM, inward currents increased. Fixing the membrane potential at resting membrane potential and comparing the barium-sensitive outward currents at $[K^+]_o$ of 6 and 15 mM showed that the barium- sensitive outward current increased at 15 mM $K^+.$ From the above results the following were concluded. Barium-sensitive $K^+$?channel activity increased when $[K^+]_o$ is elevated and this leads to an increase in $K^+-outward$ current. Consequently, the membrane potential hyperpolarizes, leading to the relaxation of resistant arteries, and this is thought to contribute to an increase in the local blood flow of brain.

• The Korean Journal of Physiology and Pharmacology
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• 제14권1호
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• pp.29-35
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• 2010

We have shown that myosin light chain kinase (MLCK) was required for the off-contraction in response to the electrical field stimulation (EFS) of feline esophageal smooth muscle. In this study, we investigated whether protein kinase C (PKC) may require the on-contraction in response to EFS using feline esophageal smooth muscle. The contractions were recorded using an isometric force transducer. On-contraction occurred in the presence of $N^G$-nitro-L-arginine methyl ester (L-NAME), suggesting that nitric oxide acts as an inhibitory mediator in smooth muscle. The excitatory composition of both contractions was cholinergic dependent which was blocked by tetrodotoxin or atropine. The on-contraction was abolished in $Ca^{2+}$-free buffer but reappeared in normal $Ca^{2+}$-containing buffer indicating that the contraction was $Ca^{2+}$ dependent. 4-aminopyridine (4-AP), voltage-dependent $K^+$ channel blocker, significantly enhanced on-contraction. Aluminum fluoride (a G-protein activator) increased on-contraction. Pertussis toxin (a $G_i$ inactivator) and C3 exoenzyme (a rhoA inactivator) significantly decreased on-contraction suggesting that Gi or rhoA protein may be related with $Ca^{2+}$ and $K^+$ channel. ML-9, a MLCK inhibitor, significantly inhibited on-contraction, and chelerythrine (PKC inhibitor) affected on the contraction. These results suggest that endogenous cholinergic contractions activated directly by low-frequency EFS may be mediated by $Ca^{2+}$, and G proteins, such as Gi and rhoA, which resulted in the activation of MLCK, and PKC to produce the contraction in feline distal esophageal smooth muscle.