• BMB Reports
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• v.41 no.11
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• pp.803-807
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• 2008

We investigated the expression of muscarinic acetylcholine receptors (mAChRs) and their possible involvement in the regulation of cell proliferation in four colon cancer cell lines (SNU-61, SNU-81, SNU-407, and SNU-1033) derived from Korean colon carcinoma patients. A ligand binding assay showed that all four cell lines expressed mAChRs. Treatment of the four cell lines with the cholinergic agonist carbachol led to the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2). In SNU-407 cells, carbachol significantly stimulated cell proliferation, which could be abolished by the muscarinic antagonist atropine and the ERK1/2 kinase inhibitor PD98059. These results indicate that mAChRs specifically mediate the proliferation of SNU-407 colon cancer cells via the ERK1/2 pathway.

• BMB Reports
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• v.34 no.2
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• pp.182-187
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• 2001

Phospholiapse D (PLD), and phosphatidic acid generated by it, have been implicated in receptor-mediated intracellular signaling. Carbachol (CCh) is known to activate PLD1, and protein kinase C (PKC) is known to mediate in this signaling pathway In recent reports (Kim et al., 1999b; Kim et al., 2000), we published our observations of the direct phosphorylation of PLD1 by PKC and we described the phosphorylation-dependent regulation of PLD1 activity. In this study, we investigated the phasphorylation and compartmentalization of PLD1 in terms of CCh signaling in M3 muscarinic receptor (M3R)-expressing COS-7 cells. CCh treatment of COS-7 cells transiently coexpressing PLD1 and M3R stimulated PLD1 activity and induced direct phosphorylation of PLD1 by PKC. The CCh-induced activation and phosphorylation of PLD1 was completely blocked upon pretreatment of the cells with PKC-specific inhibitors. We looked at the localization of the PLD1 phosphorylation by PKC and found that PLD1 was mainly located in the caveolin-enriched membrane (CEM) fraction. Based on these results, we conclude that CCh induces the activation and phosphorylation of PLD1 via PKC and that the phosphorylation of PLD1 occurs in caveolae.

• Journal of Acupuncture Research
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• v.33 no.1
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• pp.9-21
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• 2016

Objectives : The aim of this study was to investigate the effect of low frequency electro-acupuncture at ST39 on intestinal motility in rats. Methods : Intestinal hypermotility and hypomotility in rats were induced by oral carbachol ingestion and loperamide injection. Rats were divided into seventeen experimental groups including the normal and holder groups. The rats were induced with intestinal hypermotility and hypomotility and divided into pre and post-treatment groups. I also carried out acupuncture (needle retention) and low frequency electro-acupuncture at ST39 or the sham point. I fed charcoal to rats after the treatment and calculated its distance travelled in the gastrointestinal tract, which was compared by groups so as to determine which treatment was more effective in increasing or decreasing intestinal motility. Results : 1. In normal rats, low frequency electro-acupuncture at ST39 showed no significant effect on intestinal motility. 2. Pre-treatment with acupuncture (needle retention) at ST39 on intestinal motility over-activated with carbachol significantly decreased intestinal motility in rats. 3. Pre-treatment with low frequency electro-acupuncture at ST39 on intestinal motility over-activated with carbachol significantly decreased intestinal motility in rats. 4. Pre-treatment with acupuncture (needle retention and low frequency electro-acupuncture) at ST39 showed no significant effect on intestinal hypomotility in rats that was induced by loperamide injection. Conclusions : These results suggest that acupuncture (needle retention) and low frequency electro-acupuncture at ST39 have preventive effects on intestinal hypermotility. Regardless of the stimulation method, ST39 showed an effect on intestinal motility. Further study is required to confirm other effects of ST39.

• Korean Journal of Oriental Medicine
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• v.2 no.1
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• pp.236-254
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• 1996

This study was carried out to investigate the tracheal muscle relaxation of water extracts of three kinds of Coptis rhizoma and also compare their effects with berberine and palmatine, which are the important alkaloid components of Coptis rhizoma, in isolated rat tracheal smooth muscle. As a result Coptis chinenis showed significant tracheal muscle relaxation on KCI-induced contraction and more markedly on carbachol-induced contraction in dose-dependent manner. Coptis japoinin exhibited also tracheal muscle relaxation on both contraction with slightly weaker activities than those of Coptis chinensts. The activities of the root hair of Coptis chinensis were the most weakest in three rhizomas. Berberine and palmatine had strong tracheal muscle relaxation, especially, on carbachol-induced contraction, however, their activities on KCI-induced contraction were weaker than that of Coptis chinenis. The inhibitory effects of three kinds of Coptis rhizoma on the contraction by KCI and carbachol were investigated to explain the process of their tracheal muscle relaxation. All rhizomas inhibited the contraction by both agents, among them, Coptis japonica showed the most significant effect, so indicating that the tracheal muscle relaxation of Coptis rhizoma may be concerned with the blocking of muscarine receptor and $Ca^{2+}$channel.

• Archives of Pharmacal Research
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• v.10 no.4
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• pp.212-218
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• 1987

Intact brain cell aggregates were dissociated from adult rat brains without cerebellum using a sieving technique. This proparation was used to elucidate the binding characteristics of agonist to muscarinic acetylcholine receptors (mAchR) in brain. Incubation of cells with carbamylcholine (carbachol) was shown agonist-induced receptor down-regulation depending on the concentration of agonist, not depending on the incubation time. This effect of carbachol was due to a reduction in the maximal binding capacity ($B_{max}$) to the mAchR without decreasing the affinity of the remaining receptors in incubation at 37.deg.C but was not apparent inincubation at $15^{\circ}}C$In addition, it was abolished when the receptors were blocked by atropine. The decline in ($^3H$)N-methylscopolamine (($^3H$)NMS) binding induced by agonist was reflected as a significant reduction in the receptor density with no change in receptor affinity, suggesting that 'true' receptor down-regulation takes place. Moreover, when the receptors were labeled with the lipophilic antagonist ($^3H$) quinuclidinyl benzilate (($^3H$) QNB) insted of the hydrophilic ligand ($^3H$)NMS, the magnitude of the observed receptor down-regulation was significantly lower in case of the former than the latter. This suggested that exposure of intact brain cells to muscarinic agonists might induce a slight degree of accumulation of receptors in intracellular sites before the receptors are actually degraded.

• Journal of Acupuncture Research
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• v.25 no.4
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• pp.11-29
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• 2008

Objectives : The purpose of this study was to observe the effects of acupuncture and electro-acupuncture at $LI_4$ on intestinal motility in rat. Methods : We made over-activated state of intestinal motility with carbachol and suppressed state of intestinal motility with loperamide in rat and carried out needle retention acupuncture, low frequency electro-acupuncture and high frequency electro-acupuncture at $LI_4$ or sham point in rat devided into pre-treatment group and post-treatment group. and we resulted as follow for measuring charcoal travel rate to observe intestinal motility. Results : 1. Any of needle retention acupuncture, low frequency electro-acupuncture and high frequency electro-acupuncture at $LI_4$ have no significant influences on intestinal motility of rat in normal state. 2. All of the pre-treatment and post-treatment of needle retention acupuncture, low frequency electro-acupuncture and high frequency electro-acupuncture didn't significantly decrease intestinal motility in rat which over-activated with carbachol. 3. Pre-treatment of needle retention acupuncture and high frequency electro-acupuncture significantly increased intestinal motility in rat which suppressed with loperamide. 4. Post-treatment of needle retention acupuncture, low frequency electro-acupuncture and high frequency electro-acupuncture significantly increased intestinal motility in rat which suppressed with loperamide. Conclusions : These results suggest that acupuncture on $LI_4$ have preventive effect and therapeutic effect on suppressed intestinal motility, and especially high frequency electro-acupuncture is more effective.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.3
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• pp.353-359
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• 1998

In gastrointestinal smooth muscle, muscarinic stimulation by carbachol (CCh) activates nonselective cation channel current ($I_{CCh}$) which is facilitated by intracellular [$Ca^{2+}$] increase. Caffeine is widely used in experiments to mobilize $Ca^{2+}$ from intracellular stores. This study shows a strong inhibitory effect of caffeine on $I_{CCh}$ in guinea-pig gastric myocyte. In this study, the underlying mechanism of the inhibitory effect of caffeine was investigated. $I_{CCh}$ was completely suppressed by the addition of caffeine (10 mM) to the superfusing solution. Inhibition of $I_{CCh}$ by caffeine was not related to the intracellular cAMP accumulation which was expected from the phosphodiesterase-inhibiting effect of caffeine. The blockade of $InsP_3-induced$ $Ca^{2+}$ release by heparin had no significant effects on the activation of $I_{CCh}$. When the same cationic current had been induced by intracellular dialysis of $GTP[{\gamma}S]$ in order to bypass the muscarinic receptor, the inhibitory effect of caffeine was significantly attenuated. The results of this study indicate that both intracellular signalling pathways for $I_{CCh}$, proximal and distal to G-protein activation, are suppressed by caffeine. A major inhibition was observed at the proximal level.

• Current Research on Agriculture and Life Sciences
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• v.7
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• pp.231-235
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• 1989

The effects of choline and choline esters(acetylcholine, methacholine, carbachol, bethanechol) on motility of isolated rabbit jejunum segment were examined and $pD_2$ values of each drugs were compared. The results were as follows. In choline, there were revealed that maximum effective concentration was $10^{-2}M$, $ED_{50}$ was $2.4{\times}10^{-3}M$, and $pD_2$ value was 2.619. In acetylcholine, there were revealed that maximum effective concentration was $10^{-4}M$, effect was hardly showin in $10^{-9}M$ concentration, $ED_{50}$ was $0.5{\times}10^{-5}M$, and $pD_2$ value was 5.154. In methacholine, there were revealed that maximum effective concentration was $10^{-5}M$, effect was hardly shown in $10^{-9}M$ concentration, $ED_{50}$ was $9{\times}10^{-7}M$, and $pD_2$ value was 6.045. In carbachol, there were revealed that maximum effective concentration was $10^{-5}M$, effect was hardly shown in $10^{-11}M$ concentration, $ED_{50}$ was $5.7{\times}10^{-7}M$, and $pD_2$ value was 6.244. In bethanechol, there were revealed that maximum effective concentration was $10^{-4}M$, effect was hardly shown in $10^{-8}M$ concentration, $ED_{50}$ was $3.3{\times}10^{-6}M$, and $pD_2$ value was 5.480. Choline and choline esters caused contraction on motility of isolated rabbit jejunum segment. The order of $pD_2$ values of drugs was carbachol, methacholine, bethanechol, acetylcholine and choline (in the descending order of potency).

• The Korean Journal of Pharmacology
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• v.24 no.1
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• pp.83-92
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• 1988

Although the release of lysosomal enzymes from activated PMN leukocyte can be regulated by intracellular cyclic nucleotide levels, other responses of PMN leukocyte according to the binding of neurotransmitters to either ${\beta}$-adrenergic or muscarinic receptors are still not clarified. In addition, the function of PMN leukocyte mediated by ${\alpha}$-adrenergic receptors is uncertain. Atropine, phentolamine and propranolol inhibited calcium uptake, superoxide generation, NADPH oxidase activity and phagocytic activity in activated PMN leukocyte, whereas carbachol and isoproterenol slightly further stimulated the responses of activated cells. Either carbachol or isoproterenol stimulated superoxide generation was inhibited by their antagonists, atropine and propranolol, respectively. The response of activated PMN leukocyte was inhibited by chlorpromazine, verapamil and dantrolene but slightly stimulated by lithium. On the other hand, chlorpromazine and dibucaine did not affect NADPH oxidase activity. Atropine, phentolamine and propranolol suppressed the calcium dependent phagocytic activity. Thus, the results suggest that atropine, phentolamine and propranolol may inhibit superoxide generation in activated PMN leukocyte by the inhibition of calcium influx and by their direct action on the NADPH oxidase system which is associated with autonomic receptors.

• Journal of Physiology & Pathology in Korean Medicine
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• v.24 no.1
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• pp.143-151
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• 2010

In Vivo and In vitro antispasmodic effects of Jun-Si-Baek-Chul-San, a Traditional Korean Polyherbal Medicineconsisted of 7 types of herbs were observed in the present study. To clarify the effects of Jun-Si-Baek-Chul-San, on accelerating small intestinal movement induced by the stimulation of cholinergic neurotransmission, we evaluated the effects of Jun-Si-Baek-Chul-San on In vivo carbachol (an acetylcholinergic agent)-accelerated mice small intestinal transit and on In vitro contractions induced by low-frequency electrostimulation, KCl, histamine or acetylcholine using isolated guinea pig ileum. To induce the acceleration of mice small intestinal transit, Carbachol 1 mg/kg was once subcutaneously dosed 15min before last administration of the test drugs. In the present study, Jun-Si-Baek-Chul-San 500, 250 and 125 mg/kg or domperidone 20 mg/kg were orally pretreated on the carbachol-accelerated mice small intestinal transit once a day for 7 days and the small intestinal transit rateof activated charcoal powder were monitored. In vitro assays, Jun-Si-Baek-Chul-San1, 0.1, 0.01 and 0.001 mg/ml or domperidone $2{\times}10^{-5}M$ were treated 10min before ileal contraction was induced by filed stimulation, acetylcholine, KCl and histamine, and the % changes of contractions were observed compared to the treatment of inducer alone. In spontaneous contraction, the % changes of contractions were observed compared to treatment of vehicle alone at 10min after Jun-Si-Baek-Chul-San or domperidone treatment. The efficacy of Jun-Si-Baek-Chul-San was compared to those of domperidone. High concentration, 1 mg/ml of Jun-Si-Baek-Chul-San was found to decrease the spontaneous contraction of the isolated guinea-pig ileum. In addition, Jun-Si-Baek-Chul-San decrease contractions induced by electrostimulation, acetylcholine, histamine and KCl in the isolated guinea-pig ileum. In addition, Jun-Si-Baek-Chul-San effectively inhibited the accelerated small intestinal movement induced by carbachol stimulation of cholinergic neurotransmission in In vivo. Based on the results, although the exact molecular or action mechanism and which herbs or compound in Jun-Si-Baek-Chul-San are responsible for actions, it was concluded that Jun-Si-Baek-Chul-San normalization in the accelerated intestinal motility might be interfere with a variety of muscarinic, adrenergic and histaminic receptor activities or with the mobilization of calcium ions required for smooth muscle contraction non-specificly. Therefore, it is expected that Jun-Si-Baek-Chul-San will be promising as a prescription of clinical treatment of digestive tract disorders such as accelerated the motility of intestine, diarrhea or intestinal painful contractions.