• The Journal of Internal Korean Medicine
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• v.19 no.1
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• pp.409-427
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• 1998

Insamjungchuntang has been used in Korea for many centuries as a treatment for respiratory disease. The effect of Insamjungchuntang on tracheal smooth muscle is not known. The purpose of the present study is to determine the effect of Insamjungchuntang on histamine and acetylcholine induced tracheal smooth muscle contraction in rats and guinea pigs. Guinea pig (500 g, male) and Sprague Dawley rats (200 g, male) were killed by $CO_2$ exposure and a segment (8-10 mm) of the thoracic trachea from each rat and guinea pig was cut into equal segments and mounted 'in pairs' in a tissue bath. Contractile force was measured with force displacement transducers under 0.5 g loading tension. The dose of histamine (His) and acetylcholine (Ach) which evoked 50% of maximal response $(ED_{50})$ was obtained from cumulative dose response curves for histamine and acetylcholine$(10^{-7}{\sim}10^{-4}\;M)$. Contractions evoked by His ($ED_{50}$) and Ach $(ED_{50})$ were inhibited significantly by Insamjungchuntang. In guinea pig tracheal smooth muscle, the mean percent inhibition of acetylcholine induced contraction was $38.58\(p<0.05)\;after\;10{\mu}l/ml$ Insamjungchuntang, $90.75\(p<0.01)\;after\;30{\mu}l/ml$. Insamjungchuntang and $133.17\(p<0.01)\;after\;100{\mu}l/ml$ Insamjungchuntang. In rat tracheal smooth muscle, the mean percent inhibition of acetylcholine induced contraction was $10.0\(p<0.05)\;after\;10{\mu}l/ml$ Insamjungchuntang, $80.71\(p<0.01)\;after\;30{\mu}/ml$ Insamjungchuntang and $118.29\(p<0.01)\;after\;100{\mu}l/ml$ Insamjungchuntang. Also, in guinea pig tracheal smooth muscle, the mean percent inhibition of histamine induced contraction was $45.5\(p<0.01)\;after\;10{\mu}l/ml$ lnsamjungchuntang, and $93.17\(p<0.01)\;after\;30{\mu}l/ml$. lnsamjungchuntang $134.50\(p<0.01)\;after\;100{\mu}l/ml$ Insamjungchuntang. In rat tracheal smooth muscle, the mean percent inhibition of histamine induced contraction was $37.83\(p<0.01)\;after\;10{\mu}l/ml$ lnsamjungchuntang, $90.5\(p<0.01)\;after\;30{\mu}l/ml$ Insamjungchuntang and $135.17\(p<0.01)\;after\;100{\mu}l/ml$ Insamjungchuntang. Propranolol $(10^{-7}\;M)$ slightly but significantly attenuated the inhibitory effects of Insamjungchuntang. Following treatment with propranolol, the mean percent inhibition caused by $100{\mu}l/ml$. Insamjungchuntang fell to 46.42% in guinea pig induced by acetylcholine contraction and by $100{\mu}l/ml$ Insamjungchuntang fell to 5.43% (p<0.05) in rat induced by acetylcholine contraction and the mean percent inhibition caused by $100{\mu}l/ml$ Insamjungchuntang fell to 49.0% in guinea pig induced by histamine contraction and by $100{\mu}l/ml$ Insamjungchuntang fell to 48.6% (p<0.05) in rat induced by histamine contraction. Indomethacin and methylene blue $(10^{-7}\;M)$ did not significantly alter the inhibitory effect of lnsamjungchuntang. Also, I could find the effects of lnsamjungchuntang and Insamjungchuntanggamorphine on the tracheal smooth muscle in guinea pig and rat did not change significantly. These results indicate that Insamjungchuntang can relax histamine and acetylcholine-induced contraction of guinea pig and rat tracheal smooth muscle, and that this inhibition involves sympathetic effects.

• The Journal of Internal Korean Medicine
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• v.18 no.2
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• pp.1-26
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• 1997

Chunggeumtang has been used in Korea for many centuries as a treatment for respiratory disease. The effect of Chunggeumtang on tracheal smooth muscle is not konwn. The purpose of the present study is to determine the effect of Chunggeumtang on histamine and acetylcholine induced tracheal smooth muscle contraction in rats and guinea pigs. Guinea pig(500g, male) and Sprague Dawley rats (250g, male) were killed by $CO_2$ exposure and a segment (8-10mm) of the thoracic trachea from each rat and guinea pig was cut into equal swegments and mounted 'in pairs' in a tissue bath. Contractile force was measured with force diplacement transducers under 0.5g loading tension. The dose of histamine (His) and acetylcholine (Ach) which evoked 50% of maximal response ($ED_{50}$) was obtained from cumulative dose response curves for histamine and acetylcholine($10^{-7}{\sim}10^{-4}M$). Contractions evoked by His ($ED_{50}$) and Ach ($ED_{50}$) were inhibited significantly by Chunggeumtang. In guinea pig tracheal smooth muscle, the mean percent inhibition of acetylcholine induced contraction was $6.1%after\;30{\mu}l/ml$ Chunggeumtang, and 49.4% (p<0.01) after $100{\mu}l/ml$ Chunggeumtang. In rat tracheal smooth muscle, the mean percent inhibition of acetylcholine induced contraction was $6.7%\;after\;30{\mu}l/ml$ Chunggeumtang, and $54.2%\;(p<0.01)\;after\;100{\mu}l/ml$ Chunggeumtang. Also, in guinea pig tracheal smooth muscle, the mean percent inhibition of histamine induced contraction was $30.6%\;(p<0.05)\;after\;30{\mu}l/ml$ Chunggeumtang, and $53.0%\;(p<0.01)\;after\;100{\mu}l/ml$ Chunggeumtang. In rat tracheal smooth muscle, the mean percent inhibition of histamine induced contraction was $24.1%\;(p<0.05)\;after\;30{\mu}l/ml$ Chunggeumtang, and $55.3%\;(p<0.01)\;after\;100{\mu}l/ml$ Chunggeumtang. Propranolol and indomethacin($10^{-7}M$) slightly but significantly attenuated the inhibitory effects of Chunggeumtang. Following treatment with propranolol, the mean percent inhibition caused by $100{\mu}l/ml$ Chunggeumtang fell to 27.6% in guinea pig induced by acetylcholine contraction and by $100{\mu}l/ml$ Chunggeumtang fell to 28.7% (p<0.05) in rat induced by acetylcholine contraction and the mean percent inhibition caused by $100{\mu}l/ml$ Chunggeumtang fell to 16.2% in guinea pig induced by histamine contraction and by $100{\mu}l/ml$ Chunggeumtang fell to 28.7% (p<0.05) in rat induced by histamine contraction. Indomethacin, the mean percent inhibition caused by $100{\mu}l/ml$ Chunggeumtang fell to 20.0% in guinea pig induced by acetylcholine contraction and by $100{\mu}l/ml$ Chunggeumtang fell to 16.9% (p<0.05) in rat induced by acetylcholine contraction and the mean percent inhibition caused by $100{\mu}l/ml$ Chunggeumtang fell to 16.4% in guinea pig induced by histamine contraction and by $100{\mu}l/ml$ Chunggeumtang fell to 23.1% (p<0.05) in rat induced by histamine contraction. Methylene blue($10^{-7}M$) did not significantly alter the inhibitory effect of Chunggeumtang. Also, I could find the effects of Chunggeumtang and Chunggeumtanggamorphine on the tracheal smooth muscle in guinea pig and rat did not change significantly. These results indicate that Chunggeumtang can relax histamine and acetylcholine-induced contraction of guinea pig and rat tracheal smooth muscle, and that this inhibition involves sympathetic effects and the release of cyclooxygenase products.

• Journal of Life Science
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• v.22 no.4
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• pp.499-507
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• 2012

Previous work has characterized myomodulin A (MMA, PMSMLRLamide) and myomodulin E (MME, GLQMLRLamide) purified from the central nervous systems of the sea hare, $Aplysia$ $Kurodai$, using the anterior byssus retractor muscle (ABRM) of the mussel, $Mytilus$ $edulis$. The amino acid sequences of MMA and MME were the same as those of the myomodulin family peptide found in other mollusks. In this study, we synthesized MME, its derivatives, and other neuropeptides to investigate the relationship between the structure and biological activity of MME. The primary structures of MME's derivatives, Des[$Gly^1$]-MME, Des[$Gly^1,Leu^2$]-MME, and Des[$Gly^1,Leu^2,Gln^3$]-MME, were LQMLRLamide, QMLRLamide, and MLRLamide, respectively. MMA and synthetic peptides were tested on ABRM in $M.$ $edulis$ as well as muscle preparations in $Achatina$ $fulica$. MME displayed an inhibitory effect on phasic contraction of the ABRM at $1{\times}10^{-9}$ M or higher. MME also had a relaxing effect on the catch-tension of AMRM at $1{\times}10^{-8}$ M. Both MMA and its analogs stimulated a contractile response on the crop and relaxed the catch-relaxing response on the penial retractor muscle of $A.$ $fulica$. These results suggest that MME and its analogs have modulatory effects on various muscles of mollusks. This study has also laid the groundwork for future neural and circuit modulation studies during animal behavioral changes.

• The Korean Journal of Pharmacology
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• v.29 no.2
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• pp.263-274
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• 1993

Potassium $(K^+)$ channels are present in airway smooth muscle cells, and their activation results in hyperpolarization and relaxation. Because these effects may have therapeutic relevance to hypersensitivity and asthma, we examined the effect of a potassium channel activator, cromakalim (BRL 34915, CK) on the release of mediators from superfused tracheal and parenchymal strips after passive sensitization with $IgG_1$ antibody. Both tissues were superfused with CK $(2{\times}10^{-6}\;M)$ for 30 min and challenged with CK and antigen (Ox-HSA). Using monodispersed, partially purified, highly purified guinea pig lung mast cells, we also examined the effect of CK on mediator release from these cells after passive sensitization with $IgG_{1}$ antibody $({\alpha}-OA)$. Guinea pig lung mast cells were purified using enzyme digestion method, count current elutriation, and discontinuous Percoll density gradient. After CK pretreatment, passively sensitized mast cells were challenged with varying concentration of antigen (OA, immunological stimuli) or with varying concentration of calcium ionophore (CaI, non-immunological stimuli). Histamine (Hist) release was determined by spectrophotofluorometry, and leukotrienes (LT) by radioimmunoassy. CK pretreatment decreased Hist by 35% and LT release by 40% in the antigen-induced tracheal tissue after $IgG_1$ sensitization but did not decrease the contractile response. In the antigen-induced parenchymal tissue CK decreased Hist release by 25% but poorly decreased LT. Both immunologic and non-immunologic stimuli caused a dose-dependent release of Hist and LT from monodispersed, partially purified and highly purified lung mast cells. Verification of LT release was obtained by the use of 5-lipoxygenase inhibitor, A64077 (Zileuton). CK decreased Hist and LT release by 20% respectively in the OA-induced guinea pig lung mast cells after $IgG_1$ sensitization. The inhibitory effects of CK on the Hist and LT release in the Ox-HSA-induced airway smooth muscle tissues or in the OA-induced and CaI-induced mast cells after $IgG_1$ sensitization were completely blocked by TEA and GBC. These studies show that guinea pig lung mast cells seem to be an important contributor to LT release, and that CK (which has been known as an airway smooth muscle relaxant) can in part act to inhibit mediator release in the antigen-induced airway smooth muscle, and that CK may also act to inhibit mediator release in the OA-induced and CaI-induced highly purified mast cells. These results suggest that Hist and LT release evoked by mast cell activation might in part be associated with $K{^+}4 channel activity.

• The Korean Journal of Pharmacology
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• v.32 no.1
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• pp.39-49
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• 1996

In the present study, we observed change in intracellular $Ca^{2+}$$([Ca^{2+}]_i)$ as measured with the fluorescent $Ca^{2+}-indicator$ fura-2 in association with force development of the rat basilar arteries during activation by$K^+$ depolarizing solution and U46619, a thromboxane analogue, in the absence and the presence of calcitonin-gent related peptide (CGRP). CGRP (30 and 100 nM) caused a concentration-dependent inhibition of U46619-induced contraction with decrease in $[Ca^{2+}]_i$, whereas it did not exert any effect on the $K^+$ (90 mM)-induced contraction and increase in $[Ca^{2+}]_i$, Further, $[Ca^{2+}]_i-force$ relationships were determined by plotting the ratio of $F_{340}/F_{380}$ $([Ca^{2+}]_i)$ as a function of the force induced by U46619, and the results were compared with those obtained in the presence of CGRP. The curves obtained in the presence of CGRP (30 and 100 nM) were significantly moved to downward without right shift of the curves suggesting that CGRP inhibited the U46619-induced contraction only by mediation of reduction in $[Ca^{2+}]_i$ with out any change in the sensitivity of contractile apparatus to $Ca^{2+}$. The CGRP-induced attenuation of $[Ca^{2+}]_i$ and force development was significantly inhibited under pretreatment with CGRP $(8{\sim}37)$ fragment (100 nM), a CGRP1 receptor antagonist. Both the reduced contraction and reduction in $[Ca^{2+}]_i$ caused by CGRP were fully reversed by pretreatment with charybdotoxin (100 nM) and iberiotoxin (100 nM), large conductance $Ca^{2+}-activated$ $K^+$ channel blockers, but not by apamin (300 nM), a small conductance $Ca^{2+}-activated$ $K^+$ channel blocker, and glibenclamide ( 1 ${\mu}M$), an ATP-sensitive $K^+$ channel blocker. In conclusion, it is suggested that the CGRP1 receptor, upon activation by CGRP, are coupled to opening of $Ca^{2+}-activated$ $K^+$ channel and cause to decrease in $[Ca^{2+}]_i$, thereby leading to vasodilation of the rat basilar artery. However, it is not defined that the mechanism underlying vasodilation whether the $K^+$ channel blockers, charybdotoxin and iberiotoxin directly block the CGRP receptors and that CGRP-evoked relaxation is dependent on the cyclic AMP or $K^+$ channel opening or both actions.

• Journal of Chest Surgery
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• v.32 no.7
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• pp.603-612
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• 1999

Background : It has been documented that brief repetitive periods of ischemia and reperfusion (ischemic preconditioning, IP) enhances the recovery of post-ischemic contractile function and reduces infarct size after a longer period of ischemia. Many mechanisms have been proposed to explain this process. Recent studies have suggested that transient increase in the intracellular calcium may have triggered the activation of protein kinase C(PKC); however, there are still many controversies. Accordingly, the author performed the present study to test the hypothesis that preconditioning with high concentration of calcium before sustained subsequent ischemia(calcium preconditioning) mimics IP by PKC activation. Material and Method : The isolated hearts from the New Zealand White rabbits(1.5∼2.0 kg body weight) Method: The isolated hearts from the New Zealand White rabbits(1.5∼2.0 kg body weight) were perfused with Tyrode solution by Langendorff technique. After stabilization of baseline hemodynamics, the hearts were subjected to 45-minute global ischemia followed by a 120-minute reperfusion with IP(IP group, n=13) or without IP(ischemic control, n=10). IP was induced by single episode of 5-minute global ischemia and 10-minute reperfusion. In the Ca2+ preconditioned group, perfusate containing 10(n=10) or 20 mM(n=11) CaCl2 was perfused for 10 minutes after 5-minute ischemia followed by a 45-minute global ischemia and a 120-minute reperfusion. Baseline PKC was measured after 50-minute perfusion without any treatment(n=5). Left ventricular function including developed pressure(LVDP), dP/dt, heart rate, left ventricular end-diastolic pressure(LVEDP) and coronary flow(CF) was measured. Myo car ial cytosolic and membrane PKC activities were measured by 32P-${\gamma}$-ATP incorporation into PKC-specific pepetide. The infarct size was determined using the TTC (tetrazolium salt) staining and planimetry. Data were analyzed using one-way analysis of variance(ANOVA) variance(ANOVA) and Tukey's post-hoc test. Result: IP increased the functional recovery including LVDP, dP/dt and CF(p<0.05) and lowered the ascending range of LVEDP(p<0.05); it also reduced the infarct size from 38% to 20%(p<0.05). In both of the Ca2+ preconditioned group, functional recovery was not significantly different in comparison with the ischemic control, however, the infarct size was reduced to 19∼23%(p<0.05). In comparison with the baseline(7.31 0.31 nmol/g tissue), the activities of the cytosolic PKC tended to decrease in both the IP and Ca2+ preconditioned groups, particularly in the 10 mM Ca2+ preconditioned group(4.19 0.39 nmol/g tissue, p<0.01); the activity of membrane PKC was significantly increased in both IP and 10 mM Ca2+ preconditioned group (p<0.05; 1.84 0.21, 4.00 0.14, and 4.02 0.70 nmol/g tissue in the baseline, IP, and 10 mM Ca2+ preconditioned group, respectively). However, the activity of both PKC fractions were not significantly different between the baseline and the ischemic control. Conclusion: These results indicate that in isolated Langendorff-perfused rabbit heart model, calcium preconditioning with high concentration of calcium does not improve post-ischemic functional recovery. However, it does have an effect of limiting(reducing) the infart size by ischemic preconditioning, and this cardioprotective effect, at least in part, may have resulted from the activation of PKC by calcium which acts as a messenger(or trigger) to activate membrane PKC.

• The Korean Journal of Pharmacology
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• v.25 no.1
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• pp.53-58
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• 1989

For several years, we investigated the pharmacological action of several substances isolated from Buxus microphylla var koreana Nakai, which had been used as folk remedies of malaria and venereal disease. Cyclobuxine $D(C_{25}H_{42}ON_2)$, a steroidal alkaloid, exerted an antiinflammatory action, hypotensive and bradycardic effects in rats. In the present study, we isolated alkaloid from the acetone-insoluble fraction of the strong bases of this plants. This alkaloid $(C_{25}H_{38}ON_2)$ was identified as a steroidal alkaloid contained a cyclopropane ring by physical and chemical methods. It is a derivative of cyclobuxine D and named cyclobuxine E. We examined the effect of cyclobuxine E on the contractile response induced by acetylcholine and two distinct types of potassium-activated calcium channels in an intestinal smooth muscle of the rat. Cyclobuxine E inhibited significantly the Ach-induced contraction. The isolated longitudinal muscle from the rat duodenum was immersed calcium-depleted potassium depolarizing solution. Ten minutes after, 1.8 mM $CaCl_2$ was added to muscle bath and elicited a biphasic increase in muscle tension. Cyclobuxine E produced an appreciable inhibition of both components of the mechanical response. In addition, Cyclobuxine E introduced at a point when the tonic response had reached its maximum level, caused the muscle to exhibit a rapid loss of tension. Based on these experimental results, we proposed the possibility that the inhibitory action of cyclobuxine E on the isolated rat duodenum may be due to inhibiting the transmembrane fluxes of calcium ion in potassium-activated calcium channels.

• Journal of fish pathology
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• v.14 no.1
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• pp.46-53
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• 2001

The present study was undertaken to investigate and compare the effect and mode of action of tachykinins on isolated strip preparations of the intestinal smooth muscle from the nile tilapia, Oreochromis niloticus and the Israel carp, Cyprinus carpio. Both of neurokinin 1(NK-1) receptor agonist, substance P(SP) and neurokinin 2(NK-2) receptor agonist, neurokinin A(NKA) caused concentration-dependent contractions of intestinal smooth muscle in the nile tilapia and the israel carp. The efficiency and potency of these agonists varied between two fish species. In the nile tilapia intestine, the efficiency and potency of SP were greater than those of NKA. However, the efficiency and potency of SP were similar to those of NKA. In the nile tilapia intestine and the israel carp intestine, the contractile responses of SP and NKA were noncompetitively antagonized by NK-1 receptor antagonist, L-732, 138 but unaffected by NK-2 receptor antagonist, MDL 29913. In addition, SP-induced contractions in the both of preparation were significantly inhibited by muscarinic antagonist, atropine($5{\times}10^{-7}$M) and ganglionic inhibitor, tetrodotoxin($2{\times}10^{-7}$M) but NKA-induced contractions were unaffected by those. These results indicate that two tachykinin agonists, SP and NKA predominately modulate the mechanical activity of isolated preparation from the nile tilapia and the israle carp directly through the activation of NK-1 receptor on the intestinal smooth muscle cells, but in the case of SP action, the indirect action through activation of cholinergic nerve terminals seems to be also implicated.

• Journal of the Korean Society of Food Science and Nutrition
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• v.26 no.2
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• pp.300-307
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• 1997

The effects of Aster scaber and Ixeris dentata on cadiovascular system in hyperlipidemic rats were examined. Five groups of thirty Sprague Dawley rats were fed with the diet contained 1% cholesterol, 0.25% sodium cholate, 10% coconut oil and 5% lard(control group) for 4 weeks. Each experimental diet group was added with 5% plant powder or extract of the 5% plant powder by dry weight. Contractile or relaxation responses in the isolated artria and thoracic aortae were measured and the morphological changes of the aortic endotherium from the rats fed the experimental diet were inspected. In response to isoproterenol, the number of right atrial spontaneous beat was significantly lower in Cham chyi powder group$(PP_{1})$ and Sumbagui powder group$(PP_{2})$ than control at $10^(-8)M$ concentration. The contraction forces by injection of phenylephrine and calcium in isolated thoracic aorta were significantly low in each experimental groups compared with the control. The relaxation rates by acetylcholine represented comparatively higher value in $PP_{1}$ than control. The morphological changes of endothelial cell surface was a little in $PP_{1}$ and $PP_{2}$ compared with control, while the damages were considerably advanced in Cham chyi and Sumgbagui extract diet group$(PE_{1},\;PE_{2})$.

• Journal of Yeungnam Medical Science
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• v.11 no.2
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• pp.293-302
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• 1994

The objective of this study was to establish a good methodology to isolate single smooth muscle cells that are alive and respond properly to pharmacological agents. Canine urinary bladders were employed as the source of single cells, and acetylcholine, atropine and imipramine were used as indicators of pharmacological responsiveness. Imipramine, an antidepressant drug exhibited the anticholinergic and calcium antagonizing properties on rat detrusor muscle. To establish a control value for a further experiment to elucidate the mechanism of action of imipramine on detrusor muscle, we measured the concentration-response of single cells to acetylcholine in the presesnce of imipramine by length of the cells and compared the result with the response in the presence of atropine. Tiny chops of smooth muscle taken from anesthetized canine urinary bladder were incubated in collagenase solution at $36^{\circ}C$ for 17-20 minutes. The collagenase solution included collagenase 1.2 mg/ml, soybean tryspin inhibitor 0.08 mg/ml, bovine serum albumin 2% in 10 ml Krebs-Henseleit buffer solution aerated with a consistent breeze of 95/5% $O_2/CO_2$, to maintain the pH at 7.4. After washing with plain K-H solution on 450 mesh, cells were dissociated from the digested tissue for 12-15 minutes. Cell suspension was transfered in 5 ml test tubes and acetylcholine was added for the final concentration to be $10^{-14}M{\sim}10^{-9}M$. To find the optimal time to fix the cells to determine the contractile responses, 1% acrolein was added 5, 10, 20, 30, 60 and 120 seconds after the administration of ACh. The length of cells fixed by acrolein were measured by microscaler via CCTV camera on phaes-contrast microscope. The average length of 50 cells from a slide glass was taken as the value of a sample at the very concentration point. Single cells were isolated from canine detrusor. The length of untreated cells varied from 82 ${\mu}m$ to 94 ${\mu}m$. The maximal response to actylcholine $10^{-9}M$ was accomplished within 5 seconds of exposure, and the shortening was $19{\pm}3$%. Atropine reduced the contraction of the cells concentration-dependently. Imipramine which exerts a cholinergic blocking action on some smooth muscles also reduced the contraction concentration-dependently and by a similar pattern as atropine. These findings document that imipramine may exerts a cholinergic blocking activity in the single smooth muscle cells isolated from canine urinary bladder.