Bethanechol-induced Contraction Research Articles

KV7 channels in the human detrusor: channel modulator effects and gene and protein expression.

Voltage-gated type 7 K+ (KV7 or KCNQ) channels regulate the contractility of various smooth muscles. With this study, we aimed to assess the role of KV7 channels in the regulation of human detrusor contractility, as well as the gene and protein expression of KV7 channels in this tissue. For these purposes, the isolated organ technique, RT-qPCR, and Western blot were used, respectively. XE-991, a selective KV7 channel blocker, concentration-dependently contracted the human detrusor; mean EC50 and Emax of XE-991-induced concentration-response curve were 14.1μM and 28.8% of the maximal bethanechol-induced contraction, respectively. Flupirtine and retigabine, selective KV7.2-7.5 channel activators, induced concentration-dependent relaxations of bethanechol-precontracted strips, with maximal relaxations of 51.6 and 51.8% of the precontraction, respectively. XE-991 blocked the relaxations induced by flupirtine and retigabine. All five KCNQ genes were found to be expressed in the detrusor with KCNQ4 being the most expressed among them. Different bands, having sizes similar to some of reported KV7.1, 7.4, and 7.5 channel subunit isoforms, were detected in the detrusor by Western blot with the KV7.4 band being the most intense among them. In conclusion, KV7 channels contribute to set the basal tone of the human detrusor. In addition, KV7 channel activators significantly relax the detrusor. The KV7.4 channels are probably the most important KV7 channels expressed in the human detrusor. These data suggest that selective KV7.4 channel activators might represent new pharmacological tools for inducing therapeutic relaxation of the detrusor.

Expression and motor functional roles of voltage-dependent type 7 K+ channels in the human taenia coli

Voltage-dependent type 7 K+ (KV7 or KCNQ) channels modulate the excitability of neurons and muscle cells. The aims of the present study were to investigate the motor effects of KV7 channel modulators and the expression of KV7 channels in the human taenia coli. The effects of KV7 channel modulators on the muscle tone of human taenia coli strips were investigated under nonadrenergic non-nitrergic conditions by organ bath studies. Gene expression and tissue localisation of channels were studied by real-time PCR and immunohistochemistry, respectively. Under basal conditions, the KV7 channel blocker XE-991 induced concentration-dependent contractions, with mean EC50 and Emax of 18.7μM and 30.5% respectively of the maximal bethanechol-induced contraction, respectively. The KV7 channel activators retigabine and flupirtine concentration-dependently relaxed the taenia coli, with mean EC50s of 19.2μM and 29.9μM, respectively. Retigabine also relaxed bethanechol-precontracted strips, with maximal relaxations of 79.2% of the bethanecol-induced precontraction. The motor effects induced by the KV7 channel modulators were not affected by tetrodotoxin or ω-conotoxin GVIA. XE-991 greatly reduced retigabine- and flupirtine-induced relaxations. Transcripts encoded by all KCNQ genes were detected in the taenia coli, with KCNQ4 showing the highest expression levels. KV7.4 channels were clearly visualised by immunohistochemistry in colonic epithelium, circular muscle layer and taenia coli. KV7 channels appear to contribute to the resting muscle tone of the human taenia coli. In addition, KV7 channel activators significantly relax the taenia coli. Thus, they could be useful therapeutic relaxant agents for colonic motor disorders.

Impaired propulsive motility in the distal but not proximal colon of BK channel β1‐subunit knockout mice

Large-conductance Ca(2+) -activated K(+) (BK) channels regulate smooth muscle tone. The BK channel β1-subunit increases Ca(2+) sensitivity of the α-subunit in smooth muscle. We studied β1-subunit knockout (KO) mice to determine if gastrointestinal (GI) motility was altered. Colonic and intestinal longitudinal muscle reactivity to bethanechol and colonic migrating motor complexes (CMMCs) were measured in vitro. Gastric emptying and small intestinal transit were measured in vivo. Colonic motility was assessed in vivo by measuring fecal output and glass bead expulsion time. Myoelectric activity of distal colon smooth muscle was measured in vitro using intracellular microelectrodes. Bethanechol-induced contractions were larger in the distal colon of β1-subunit KO compared to wild type (WT) mice; there were no differences in bethanechol reactivity in the duodenum, ileum, or proximal colon of WT vsβ1-subunit KO mice. There were more retrogradely propagated CMMCs in the distal colon of β1-subunit KO compared to WT mice. Gastrointestinal transit was unaffected by β1-subunit KO. Fecal output was decreased and glass bead expulsion times were increased in β1-subunit KO mice. Membrane potential of distal colon smooth muscle cells from β1-subunit KO mice was depolarized with higher action potential frequency compared to WT mice. Paxilline (BK channel blocker) depolarized smooth muscle cells and increased action potential frequency in WT distal colon. BK channels play a prominent role in smooth muscle function only in the distal colon of mice. Defects in smooth muscle BK channel function disrupt colonic motility causing constipation.

Neostigmine-induced contraction and nitric oxide-induced relaxation of isolated ileum from STZ diabetic guinea pigs

Both delayed gastrointestinal transit and autonomic neuropathy have been documented in patients with diabetes mellitus. The mechanism of neostigmine, an agent that mimics release of acetylcholine from autonomic neurons by prokinetic agents, to contract smooth muscle, despite dysfunctional enteric neural pathways, was determined using isolated ilea from STZ-treated and control guinea pigs. Both bethanechol- and neostigmine-induced contractions were stronger in diabetic ileum. Bethanechol-induced contractions of control but not diabetic ileum were increased by low dose scopolamine suggesting reduced activation of presynaptic muscarinic autoreceptors in diabetic ileum. The muscarinic receptor antagonist 4-DAMP strongly, but the nicotinic receptor antagonist hexamethonium only weakly, reduced neostigmine-induced contractions of control and diabetic ilea. The amount of acetylcholine, inferred from tissue choline content, was increased in diabetic ileum. Nicotinic neural and noncholinergic postjunctional smooth muscle receptors contributed more strongly to neostigmine-induced contractions in diabetic than control ileum. Relaxation of diabetic ileum by exogenous nitric oxide generated from sodium nitroprusside was comparable to control ileum, but smooth muscle relaxation by l-arginine using neuronal nitric oxide synthase to generate nitric oxide was weaker in diabetic ileum with evidence for a role for inducible nitric oxide synthase. Despite autonomic neuropathy, neostigmine strongly contracted ileum from diabetic animals but by a different mechanism including stronger activation of postjunctional muscarinic receptors, greater synaptic acetylcholine, stronger activation of noncholinergic excitatory pathways, and weaker activation of inhibitory pathways. A selective medication targeting a specific neural pathway may more effectively treat disordered gastrointestinal transit in patients with diabetes mellitus.

The effect of boric acid on acethylcholine, bethanechol and potasssium‐evoked responses on ileum of rat

1 The aim of this study was to clarify the effect of boric acid on contractions of rat isolated ileum. 2 Contractile responses expressed as Emax and pD2 for acetylcholine (10(-3)-10(-8) m, Ach), bethanechol (10(-3)-10(-8) m) and potassium (10-80 × 10(-3) m, KCl) were determined in the absence and presence of boric acid (10(-3); 5 × 10(-4); 10(-4) m). 3 The contractile response to Ach in the presence of verapamil (10(-6) or 10(-8) m) or in calcium-free Tyrode's solution was also determined in the absence and presence of boric acid. 4 Boric acid did not affect the contractile response to Ach, bethanechol or KCl. Single or cumulative treatment of boric acid did not affect ileum muscle contraction evoked by KCl. The atropine-resistant component of Ach-induced contraction and 4-diphenyl-acetoxy-N-methyl-piperidine methiodide-resistant component of bethanechol-induced contraction were not inhibited by boric acid (10(-3) m). The contractile response to Ach was reduced in calcium-free Tyrode's solution, and the contractile response was not affected by (10(-8) m). The addition of boric acid (10(-3) m) in combination with verapamil (10(-8) m) did not significantly affect the contractile response to Ach. 5 In conclusion, boric acid does not affect contractions induced by Ach, bethanechol or potassium in rat isolated ileum.

M2028 The use of a New Wireless Motility Device (SmartPill®) for Measurement of Gastrointestinal Transit Time After Dietary Fiber Intervention

G A A b st ra ct s tension recordings of KCland bethanechol-induced contractions in circular orientated myenteric strips. Intracellular calcium was determined by fluorescencemicroscopy in isolated smooth muscle cells loaded with fura-2-AM (ExWL 340/380nm, EmWL 510nm) and calcium changes were expressed as the ratio between elicited fluorescence at 340nm and 380nm (F340/380). Mitochondrial membrane potential was measured in isolated colonic smooth muscle cells using the cell-permeant green fluorescent dye rhodamine 123 Results: Aging caused an increase in caffeine-induced Ca2+ release (140 % respect to adult cells) and a decrease in mitochondrial membrane potential, as indicated by the reduction of the heterogeneity of the rhodamine123 fluorescence signal in isolated cells. When mitochondria function was impaired using rotenone (1 μM) plus oligomycin (5 μM) the amplitude of Ca2+ release was significantly reduced (by 45%) and the [Ca2+]i decay showed a slower rate in senescent cells. Melatonin treatment protected from the deleterious effects of these agents. These changes in Ca2+ homeostasis did not translate into alterations in contractility, since pretreatment of both proximal and distal circular myenteric strips with rotenone, oligomicin of both for 30 min did not modify significantly the contractile response to neither KCl (60 mM) nor bethanechol (100 μM). Conclusion: Mitochondria play a more important role in Ca2+ homeostasis in aged than adult cells, however these changes are not enough to modify contractility, which suggest that in colonic smooth muscle mitochondrial respiratory chain is not essencial for contraction and that more sources of ATP than ATP synthase coexist in these cells. Supported by BFU2007-60563 and JEX

Quantitation of the Contractile Response Mediated by Two Receptors: M2and M3Muscarinic Receptor-Mediated Contractions of Human Gastroesophageal Smooth Muscle

Although muscarinic receptors are known to mediate tonic contraction of human gastrointestinal tract smooth muscle, the receptor subtypes that mediate the tonic contractions are not entirely clear. Whole human stomachs with attached esophagus were procured from organ transplant donors. Cholinergic contractile responses of clasp, sling, lower esophageal circular (LEC), midesophageal circular (MEC), and midesophageal longitudinal (MEL) muscle strips were determined. Sling fibers contracted greater than the other fibers. Total, M(2) and M(3) muscarinic receptor density was determined for each of these dissections by immunoprecipitation. M(2) receptor density is greatest in the sling fibers, followed by clasp, LEC, MEC, and then MEL, whereas M(3) density is greatest in LEC, followed by MEL, MEC, sling, and then clasp. The potency of subtype-selective antagonists to inhibit bethanechol-induced contraction was calculated by Schild analysis to determine which muscarinic receptor subtypes contribute to contraction. The results suggest both M(2) and M(3) receptors mediate contraction in clasp and sling fibers. Thus, this type of analysis in which multiple receptors mediate the contractile response is inappropriate, and an analysis method relating dual occupation of M(2) and M(3) receptors to contraction is presented. Using this new method of analysis, it was found that the M(2) muscarinic receptor plays a greater role in mediating contraction of clasp and sling fibers than in LEC, MEC, and MEL muscles in which the M(3) receptor predominantly mediates contraction.

Nicotine-Induced Neurogenic Relaxation in the Mouse Colon: Changes With Dextran Sodium Sulfate–Induced Colitis

Nicotine has been shown to reduce both tone and muscular activity in the human colon by releasing nitric oxide (NO) from nerves. To our knowledge, however, the effect of nicotine on mouse colon has not been elucidated, and the response in tissue from ulcerative colitis (UC) has not been investigated. We examined nicotine-induced responses in colon from control mice and mice with dextran sodium sulfate (DSS)-induced UC. In controls, bath application of nicotine caused a transient relaxation in longitudinal preparations from the transverse and distal colons but not from the rectum. The response was observed in the presence of bethanechol, abolished by treatment with tetrodotoxin and hexamethonium, and mediated partially (>50%) by the NO pathway. In longitudinal preparations of the distal colon from DSS-treated mice, spontaneous contractions decreased markedly, and nicotine caused contraction without relaxation in half of the preparations tested. Nicotine-induced relaxation in the presence of bethanechol was significantly decreased in the DSS-treated distal colon without changing bethanechol-induced contractions. These data suggest that 1) responses to nicotine differ dependent on colon regions, 2) DSS treatment predominantly caused nicotine-sensitive neurogenic changes in distal colon, and 3) DSS treatment may reverse the direction of nicotine-evoked responses in the colon, in mice.

Effects of ( R,R)- and ( R,R/S,S)-Formoterol on Airway Relaxation and Contraction in an Experimental Rat Model

Background: Racemic ( R,R/S,S)-formoterol is a long-acting β-agonist composed of a 50:50 mixture of ( R,R)- and ( S,S)-enantiomers. Objective: The aim of this study was to determine whether ( R,R)-formoterol and ( R,R/S,S)-formoterol have differing effects on airway contraction and relaxation in vitro. Methods: Cylindrical airway segments 3-mm long were isolated from the mid-trachea of healthy Sprague-Dawley rats and placed in a modified Krebs-Henseleit solution. Dose-response curves of bethanechol-induced contraction (measured as milligrams of tension) and the concentration of bethanechol that elicited 50% to 75% of maximal contraction (EC 50–75) were determined. The air-way cylinders were then precontracted with bethanechol at the EC 50–75 and exposed to different concentrations of ( R,R)-formoterol (0.0001–1.0 μM) or ( R,R/S,S)-formoterol (0.0002–2.0 μM). Each concentration of the 2 formoterol formulations contained the same amount of ( R,R)-enantiomer (eg, [ R,R]-formoterol 0.0001 μM and [ R,R/S,S]-formoterol 0.0002 1JM contained the same amount of [ R,R]-enantiomer). The relaxation percentage in response to formoterol was calculated as a reduction in tension (in milligrams) in relation to baseline tension in the precontracted state, with each tracheal cylinder serving as its own control. To determine the effect of ( R,R)-formoterol on airway contraction, tracheal cylinders were incubated with ( R,R)- or ( R,R/S,S)-formoterol before electrical field stimulation (EFS). Results: Tracheae from 56 three-week-old Sprague-Dawley rats were used in the study. The relaxation percentage of precontracted trachea was significantly greater after exposure to ( R,R)-formoterol than to ( R,R/S,S)-formoterol at a 2-fold higher concentration ( P = 0.03; general linear model with repeated measures analysis comparing the 2 groups of animals). However, in a post hoc analysis, the mean (SE) relaxation percentage of precontracted trachea was significantly greater only after exposure to ( R,R)-formoterol 0.01 μM than to ( R,R/S,S)-formoterol 0.02 μM (15.6% [5.8%] vs 39.0% [5.6%]; P < 0.05, unpaired t test). EFS-induced airway contraction was significantly less in tracheal cylinders incubated in ( R,R)-formoterol compared with those incubated in ( R,R/S,S)-formoterol at a 2-fold higher concentration ( P = 0.05; general linear model with repeated measures analysis comparing the 2 groups of animals). However, in the post hoc analysis, mean (SE) EFS-induced tracheal contraction was significantly less only in ( R,R)-formoterol 0.01 μM compared with ( R,R/S,S)-formoterol 0.02 μM at 10 V (1070 [55] mgvs 1225 [28] mg; P < 0.05, unpaired t test). Conclusion: We found that ( R,R)-formoterol may induce greater relaxation of precontracted airway smooth muscle cells than ( R,R/S,S)-formoterol and that ( R,R)-formoterol may have a greater inhibitory effect on the endogenous cholinergic and excitatory nonadrenergic, noncholinergic contractile airway responses than ( R,R/S,S)-formoterol. We speculate that the presence of the ( S,S)-enantiomer in ( R,R/S,S)-formoterol may impair airway relaxation of pre-contracted trachea in rats.

Effect of intravenous infusion of proglumide on ruminal motility in conscious sheep (Ovis aries)

The effects of intravenous infusion of proglumide on regular ruminal contractions were examined in conscious sheep using doses that inhibit pancreatic exocrine secretion. After a control period of 20 min, proglumide was infused intravenously for 40 min at a dose of 15, 30 or 60 micromol/kg per min and venous blood was collected. The intravenous infusion of proglumide significantly increased the frequency of ruminal contractions at 15 micromol/kg per min without altering the amplitude, while it significantly decreased the frequency and amplitude of ruminal contractions at 30 and 60 micromol/kg per min in a dose-dependent manner. Proglumide did not increase contractile activity of the omasum, abomasum and duodenum or the plasma concentration of immunoreactive cholecystokinin (CCK). Application of proglumide at 1-30 mmol/L inhibited bethanechol-induced contraction in both longitudinal and circular muscle strips of the dorsal sac of the rumen. These results suggest that proglumide at a low dose acts indirectly on the rumen as a CCK receptor antagonist to increase the frequency of contractions, whereas at higher doses it inhibits cholinergic-induced contraction of the ruminal muscles or acts as an agonist to inhibit contractions in sheep. Hence, proglumide at high doses seems unsuitable for research or therapeutic use as a CCK receptor blockade in sheep.

Hyperoxic conditions inhibit airway smooth muscle myosin phosphatase in rat pups

Exposure of rat pups to 100% oxygen is a model for studying neonatal lung injury. Airway reactivity is increased in this model, in part due to impaired airway smooth muscle (ASM) relaxation. We compared biochemical determinants of ASM contractility in rat pups exposed to 100% oxygen for 7 days vs. littermates raised in room air. The baseline quantities of ASM contractile proteins, extent of phosphorylation of the 20-kDa myosin regulatory light chain (LC(20)), and amount of the myosin-binding subunit of smooth muscle myosin phosphatase (MYPT) were all comparable between the two groups. Bethanechol-induced contraction increased the extent of phosphorylation of both LC(20) and MYPT in the hyperoxic group (45% and 70% over control, respectively). Relaxation after electrical field stimulation demonstrated greater phosphorylation of both LC(20) and MYPT in the hyperoxic group compared with controls (67% and 84%, respectively). To determine if hyperoxia induced changes in the isoforms of MYPT, isoform expression was also compared but differences were not found. To determine potential mechanisms whereby MYPT phosphorylation was increased by hyperoxia, separate tracheas were treated with the Rho kinase inhibitor Y-27632. This treatment completely eliminated differences in MYPT phosphorylation between the groups. Because phosphorylation of MYPT impairs the phosphatase activity of myosin phosphatase, these data suggest that hyperoxic conditioning during early postnatal life impairs relaxation through prolonging LC(20) phosphorylation. This mechanism might contribute to increased ASM reactivity seen in bronchopulmonary dysplasia.

Effects of ethanol and its metabolite acetaldehyde on responses of the rat bladder.

To investigate the pharmacological effects of ethanol and its metabolite acetaldehyde on isolated rat bladder muscle, and thus assess the potential influence of ethanol ingestion on the risk of urinary retention in patients with benign prostatic hyperplasia. Isometric tension changes of isolated rat bladder muscle strips were recorded in an organ bath using a pressure transducer. The acute or prolonged effects of ethanol (1-4%) or acetaldehyde (0.01, 0.1 or 1 mmol/L) were assessed on resting tension, electrical field stimulation (EFS), and bethanechol- (0.5 mmol/L), ATP- (2 mmol/L) or KCl- (127 mmol/L) induced contraction. To determine the mechanism of acetaldehyde-induced stimulation, an antihistamine, diphenhydramine was used after bethanechol stimulation. At the concentrations used, ethanol and acetaldehyde did not change the pH of the bathing medium. The resting tension of the muscle was not changed by ethanol, and acetaldehyde caused only a small increase in baseline tone at 1 mmol/L. Incubation with ethanol or acetaldehyde significantly suppressed contractility induced by EFS, bethanechol, ATP or KCl at each concentration (P<0.05). Contractions induced by all drugs were not changed significantly by the acute application of ethanol and the acute application of acetaldehyde did not affect contractions induced by ATP or KCl. However, EFS- or bethanechol-induced contractions were significantly enhanced (P<0. 05). The acetaldehyde-induced effects were completely blocked by the H1 antagonist, diphenhydramine (10 micromol/L). Ethanol did not affect resting tension but acetaldehyde and ethanol suppressed bladder muscle contractions. However, direct acetaldehyde-stimulation may release histamine and enhance contractility. This suggests that chronic alcoholism rather than acute intoxication is more likely to provoke urinary retention.

Participation of nitric oxide in the relaxation of the rat gastric corpus.

Nitric oxide is an important mediator of the relaxation in the rat gastric fundus. The present study investigates the role of NO in the rat gastric corpus in vitro, since the corpus differs from the fundus with regard to its physiological function and its spontaneous motor behaviour. In the presence of guanethidine electrically induced relaxations of circular, mucosa-free corpus strips precontracted with bethanechol were concentration-dependently reduced by the NO-synthase inhibitors L-NG-nitro-arginine (L-NNA) or L-NG-nitro-arginine-methyl-ester (L-NAME). The D-enantiomers were markedly less active. The inhibitory effect of L-NAME could be prevented by L-arginine. L-NNA and L-NAME, however, did not influence spontaneous motility or the bethanechol-induced contraction. Vasoactive intestinal polypeptide or sodium nitroprusside also relaxed the muscle strips, but these relaxations were not affected by L-NAME. When the corpus strips were stimulated electrically from baseline, they reacted with a contraction followed by relaxation. L-NNA or L-NAME blocked the relaxatory and enhanced the contractile component. In strips that also reacted with a rebound contraction, it was blunted by L-NAME. These effects of the NO-synthase inhibitors were abolished in the presence of atropine. Apamin increased the electrically induced contraction of the muscle strips. Inhibition of the relaxation together with a further shift to contraction could only be seen when apamin was combined with L-NNA. The inhibitory action of apamin and apamin + L-NNA was not influenced by atropine. The results demonstrate a role of NO in the relaxation of the circular muscle of the rat gastric corpus both at a postsynaptic site and via inhibition of acetylcholine release. The relaxation induced by vasoactive intestinal polypeptide does not involve NO.

Gastrokinetic effects of erythromycin: myogenic and neurogenic mechanisms of action in rabbit stomach.

The aims of this study were to determine regional differences and the mechanism of gastric contractile effects of erythromycin. Rabbit gastric circular muscle strips were studied in vitro. The threshold dose for erythromycin was significantly less and the maximum contraction greater in the antrum (1 microM and 0.9 +/- 0.3 kg/cm2) than in the fundus (10 microM and 0.3 +/- 0.1 kg/cm2). Erythromycin-induced antral contractions were decreased by motilin tachyphylaxis but unaffected by tetrodotoxin, atropine, hexamethonium, or ondansetron. At a subthreshold dose (0.1 microM), erythromycin increased the frequency, but not the amplitude, of bethanechol (10 +/- 3%)-and substance P-induced (13 +/- 5%) phasic antral contractions. This chronotropic effect was inhibited with tetrodotoxin, atropine, or motilin tachyphylaxis. Erythromycin (10 microM) and motilin (1 microM) enhanced the amplitude of substance P-induced tonic fundic contractions by 38 and 32%, respectively, without effect on bethanechol-induced contractions. In summary, erythromycin contracts antral muscle more potently and forcefully than fundic muscle. Erythromycin increases antral contractility by two mechanisms: an inotropic effect acting on smooth muscle motilin receptors, and, at lower doses, a cholinergic chronotropic effect mediated through neuronal motilin receptors.

Thyrotropin-releasing hormone: an inhibitory regulatory peptide of feline lower esophageal sphincter.

The functional role of thyrotropin-releasing hormone (TRH) at the lower esophageal sphincter (LES) was examined in the cat. The specific aims of this study were to determine: 1) the relative distribution of TRH throughout the feline gastrointestinal tract and 2) the effect of TRH on LES basal pressures and its response to exogenously induced contractions. TRH concentrations were determined by radioimmunoassay in tissue extracts from 12 sites. The mean concentration of TRH at the manometrically determined LES was 240 +/- 85 pg/g wet wt tissue, and the maximal concentration was just distal to the LES (659 +/- 189 pg/g wet wt). TRH concentration was higher in the mucosa than the underlying muscle layer of the fundus, antrum, duodenum, and ileum. In physiological studies, TRH given selectively via the left gastric artery had no effect on basal LES or esophageal pressures. TRH (2.8 x 10(-8) mol/kg) decreased the LES response to the D50 of substance P by 47.2% (34.8 +/- 3.1 to 18.4 +/- 2.9 mmHg, P < 0.01). In the presence of tetrodotoxin, TRH gave a similar inhibition of substance P-induced contractions (53.5%). TRH also decreased bombesin-induced contractions by 47.5% (29.6 +/- 6.0 to 15.8 +/- 3.9 mmHg, P < 0.025). TRH, however, had no effect on bethanechol-induced contractions. We conclude that 1) TRH is present throughout the gastrointestinal tract, with highest concentrations in the region distal to the LES; 2) TRH has no effect on basal LES tone; and 3) TRH inhibits the LES response to endogenously released and exogenous substance P but not the LES response to bethanechol.(ABSTRACT TRUNCATED AT 250 WORDS)

Actions of potassium channel openers in rat detrusor urinae.

This study was performed to investigate the action of potassium channel openers on the mechanical activity of detrusor muscle isolated from rats. Detrusor muscle strips, 15 mm in length, were myographied isometrically in an isolated organ bath. P 1060, RP 49356 and BRL 38277, potassium channel activators, reduced the basal tone and diminished the phasic activity of detrusor concentration-dependently. P 1060, RP 49356 and BRL 38227 suppressed the maximal responses to bethanechol and shifted the concentration-response curves of bethanechol-induced contraction to the right. RP 49356 and BRL 38227 reduced the contraction by low (20 mM) concentration of potassium. P 1060, however, diminished the high (80 mM) and low (20 mM) concentration of potassium-induced contraction. Glibenclamide, an inhibitor of ATP-dependent potassium channel, antagonized the suppressive action of P 1060, RP 49356 and BRL 38227 on the basal tone. Apamin or procaine did not antagonize it significantly. Based on these results, it is suggested that the relaxation of detrusor muscle strip caused by P 1060, RP 49356 and BRL 38227 may predominantly involve opening of the same potassium channel, i.e., the ATP-dependent potassium channel.

Existance of cholinergic and purinergic receptor on the detrusor muscle of rat urinary bladder

This study was aimed at investigation of the stimulatory innervations on the rat urinary bladder. Detrusor muscle strips of 15 mm long were suspended in isolated muscle chambers containing 1 ml of PSS maintained at and aerated with 95% . Isometric myography was perfomed, and the results were as followings : Muscle strips showed on-contraction by electric field stimulation (EFS) frequency-dependently. The EFS-induced contraction was not affected by hexamethonium, a ganglion blocker, but abolished, by tetrodotoxin, a nerve conduction blocker. Physostigmine, a cholinesterase inhibitor enhanced the EFS-induced contraction which was inhibited by hemicholinium, an inhibitor of choline uptake at the cholinergic nerve ending. Such an EFS-induced contraction was antagonized by atropine only partially, and the atropine-resistant portion was completely abolished by the desensitization of purinergic receptors by prolonged incubatin of the strips in the presence of high concentratin of ATP. Bethanechol, a cholinergic agonist, elicited concentration-dependent contraction. Adenosine triphosphate (ATP), a purinergic agonist, induced a weak but concentration-dependent contraction of short duration. Bethanechol-induced contraction was not affected by ATP-desensitization, and ATP-induced contraction was not affected by tetrodotoxin. These results suggest that there are at least two main stimulatory components of innervations in the detrusor muscle, cholinergic muscarinic and purinergic ; and those receptors are independent each other.

Effect of selective muscarinic antagonists on peristaltic contractions in opossum smooth muscle.

In this study we examined the role of M1- and M2-muscarinic receptors in the mediation of circular smooth muscle esophageal contractions elicited by pharmacological cholinergic stimulation and during peristalsis in anesthetized opossums. Esophageal-body contractions were induced by bethanechol administration, whereas peristalsis was elicited by pharyngeal stroking or cervical vagal stimulation. Contractions were measured by a low-compliance manometric recording system. The incidence and amplitude of bethanechol-induced contractions were antagonized by 4-diphenylacetoxy-n-methylpiperidine (4-DAMP) and atropine but not pirenzepine. 4-DAMP and atropine caused an increased velocity, decreased amplitude, and preferential reduction of the incidence of primary peristaltic contractions in the proximal smooth muscle esophagus. During long-train vagal stimulation, intra-stimulus A-waves had a velocity similar to primary peristalsis, whereas poststimulus B-waves showed a velocity considerably faster than primary peristalsis. Short-train vagal stimulation produced a contraction sequence, termed an "S-wave," that had a velocity similar to that of the A-wave. At low doses 4-DAMP increased the velocity and decreased the amplitude of A-wave and S-wave contractions, and at high doses 4-DAMP abolished both the A-wave and S-wave contractions. B-wave contractions were minimally affected by 4-DAMP. Pirenzepine had no effect on contractions induced by swallows or vagal stimulation. We conclude that M2-muscarinic receptors mediate esophageal contractions in the circular smooth muscle during primary peristalsis and during A-waves and S-waves induced by vagal stimulation, and M1-receptors do not have any important role in the excitatory neural pathway to the esophagus.(ABSTRACT TRUNCATED AT 250 WORDS)

EFFECT OF TRIMEBUTINE MALEATE ON BETHANECHOL-INDUCED CONTRACTIONS OF GASTROINTESTINAL TRACT IN CONSCIOUS AND ANAESTHETIZED DOGS

The effect of trimebutine maleate (TM-906) on bethanechol-induced contractions of the gastrointestinal tract in dogs was studied by means of chronically implanted force transducers, and it was compared with that of metoclopramide and atropine. During the period of motor quiescence in the fasted state of conscious dog, a bolus i.v. injection of 10 to 50 μg/kg bethanechol caused rhythmic contractions in the stomach and small intestine, in which an increase in basal tone was often accompanied in the distal ileum. The stomach responded to a less extent than the small intestine. Intravenous infusion of TM-906 at 3.0 mg/kg-hr for 20 to 60 min reduced the gastric contractions induced by 20 to 30 μg/kg bethanechol in five out of six conscious animals examined. The drug, however, enhanced the contractions of the duodenum and jejunum in a majority of the animals. Metoclopramide (1.8 mg/kg-hr) showed a tendency to potentiate the bethanechol-induced contractions in the stomach and small intestine, and atropine (0.06 mg/kg-hr) diminished them. In pentobarbital-Na anaesthetized dogs, effects of TM-906 on bethanechol-induced contractions resembled those in conscious dogs, inhibition in the stomach and potentiation in the small intestine.

Some in vitro and in vivo actions of the new histamine H2-receptor antagonist, ranitidine.

1 Ranitidine has been investigated as an antagonist of the H2-receptor-mediated responses to histamine of guniea-pig atrium and rat uterus in vitro and as an inhibitor of gastric acid secretion in the rat. 2 Ranitidine competitively antagonized histamine-induced increases in contraction frequency of the guinea-pig isolated rat atrium. Ranitidine had a pA2 of 7.2 and was 7.9 and 4.5 times more potent than metiamide and cimetidine respectively. 3 Ranitidine competitively antagonized histamine-induced relaxations of the rat isolated uterine horn. Ranitidine had a pA2 of 6.95 and was 3.6 and 5.9 times more potent than metiamide and cimetidine respectively. 4 Ranitidine, even at high concentrations, did not affect responses of the guinea-pig isolated atrium or rat isolated uterus to (-)-isoprenaline. Similarly it was without effect on either histamine or bethanechol-induced contractions of guinea-pig isolated ileum. 5 Ranitidine inhibited histamine- and pentagastrin-induced gastric acid secretion in the perfused stomach preparation of the anaesthetized rat. Ranitidine was 5.2 and 7.0 times more potent on a molar basis than metiamide and cimetidine respectively, as an inhibitor of histamine-induced gastric acid secretion. 6 It is concluded that ranitidine is a potent, competitive and selective antagonist of histamine at H2-receptor sites and and effective inhibitor of gastric acid secretion in vivo.