Effects of muscarinic receptor antagonists on acetylcholine-induced contractions of jejunal smooth muscle in horses

This study investigated the effects of a muscarinic type 1 (M(1)), 2 (M(2)), and 3 (M(3)) antagonists (4-DAMP, pirenzepine, and methoctramine, respectively) on acetylcholine (Ach)-induced contractions of longitudinal jejunal muscle strips of horses. Strips were irrigated with Krebs-Henseleit solution gassed with 95% O(2) and 5% CO(2), and the developed tension in response to Ach was recorded before and after incubation with increasing concentrations of 4-DAMP (10(-8)-10(-6) M), pirenzepine (10(-6)-10(-4) M), and methoctramine (10(-5)-10(-3) M). When competitive antagonism was characterized, the affinity constant (pA(2)) was calculated by Schild plots. A parallel rightward shift in the concentration-response curves was observed after 4-DAMP and pirenzepine. Methoctramine presented a dual effect on the concentration-response curves: lower concentrations induced a parallel rightward shift without altering the maximum intensity of contraction (E(max)), while the highest concentration increased slope of the concentration-response curve and increased E(max). The pA(2) for 4-DAMP and pirenzepine was 9.18 and 7.13, respectively. Acetylcholine-induced contractions of longitudinal jejunal smooth muscle are mediated mainly via M(3) receptors. The complex role of M(2) receptors in jejunal smooth muscle contractions was evident because methoctramine potentiated the contractile response to higher doses of Ach.     

Mediation of acetylcholine and substance P induced contractions by myosin light chain phosphorylation in feline colonic smooth muscle

OBJECTIVES: To determine the role of myosin light chain phosphorylation in feline colonic smooth muscle contraction. SAMPLE POPULATION: Colonic tissue was obtained from eight 12- to 24-month-old cats. PROCEDURE: Colonic longitudinal smooth muscle strips were attached to isometric force transducers for measurements of isometric stress. Myosin light chain phosphorylation was determined by isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Stress and phosphorylation were determined following stimulation with ACh or SP, in the absence or presence of a calmodulin antagonist (W-7; 0.1 to 1.0 mM), myosin light chain kinase inhibitor (ML-9; 1 to 10 microM), or extracellular calcium free solutions. RESULTS: Unstimulated longitudinal colonic smooth muscle contained low amounts (6.9+/-3.2%) of phosphorylated myosin light chain. Phosphorylation of the myosin light chains was dose and time dependent with maximal values of 58.5% at 30 seconds of stimulation with 100 microM Ach and 60.2% at 45 seconds of stimulation with 100 nM SP Active isometric stress development closely paralleled phosphorylation of the myosin light chains in ACh- or SP-stimulated muscle. W-7 and ML-9 dose dependently inhibited myosin light chain phosphorylation and isometric stress development associated with ACh or SP stimulation. Removal of extracellular calcium inhibited myosin light chain phosphorylation and isometric stress development in ACh-stimulated smooth muscle. CONCLUSIONS AND CLINICAL RELEVANCE: Feline longitudinal colonic smooth muscle contraction is calcium-, calmodulin-, and myosin light chain kinase-dependent. Myosin light chain phosphorylation is necessary for the initiation of contraction in feline longitudinal colonic smooth muscle. These findings may prove useful in determining the biochemical and molecular defects that accompany feline colonic motility disorders.     

Short chain fatty acids stimulate feline colonic smooth muscle contraction

The effect of short chain fatty acids (SCFA) on feline colonic smooth muscle contraction was evaluated in vitro. Colonic tissue was obtained from seven healthy male and female adult cats and seven healthy male and female kittens. Longitudinal and circular colonic smooth muscle strips from proximal and distal colon were incubated with SCFA (acetate, butyrate and propionate; 1-100mM). SCFA-induced contractions were compared to responses obtained using maximal concentrations (10(-4)M) of acetylcholine (ACh). The calcium dependence of the SCFA response was investigated by incubating with nifedipine (1 microM) or verapamil (1 microM). Acetate, butyrate and propionate elicited isometric stress responses (0.25-1.98 x 10(4)N/m(2)) in longitudinal, but not circular, smooth muscle from both the proximal and distal colon of adult cats. Maximal responses were attained at 50 and 100mM SCFA. Maximal butyrate and propionate responses were 29 and 19% of the maximal ACh response (10(-4)M), respectively. Acetate was least effective in stimulating contractile responses. Nifedipine and verapamil abolished all responses. Contractile responses in kittens were similar to those observed in adult cats, but were smaller in amplitude.Results of these studies have shown that SCFA stimulate longitudinal colonic smooth muscle contractions in kittens and adult cats in vitro. These SCFA-induced contractions involve activation of calcium influx. These in vitro findings may account for some of the effects of dietary fiber on feline colonic motility in vivo.     

In vitro effects of bethanechol on specimens of intestinal smooth muscle obtained from the duodenum and jejunum of healthy dairy cows

OBJECTIVE: To describe the in vitro effects of bethanechol on contractility of smooth muscle preparations from the small intestines of healthy cows and define the muscarinic receptor subtypes involved in mediating contraction. SAMPLE POPULATION: Tissue samples from the duodenum and jejunum collected immediately after slaughter of 40 healthy cows. PROCEDURES: Cumulative concentration-response curves were determined for the muscarinic receptor agonist bethanechol with or without prior incubation with subtype-specific receptor antagonists in an organ bath. Effects of bethanechol and antagonists and the influence of intestinal location on basal tone, maximal amplitude (A(max)), and area under the curve (AUC) were evaluated. RESULTS: Bethanechol induced a significant, concentration-dependent increase in all preparations and variables. The effect of bethanechol was more pronounced in jejunal than in duodenal samples and in circular than in longitudinal preparations. Significant inhibition of the effects of bethanechol was observed after prior incubation with muscarinic receptor subtype M(3) antagonists (more commonly for basal tone than for A(max) and AUC). The M(2) receptor antagonists partly inhibited the response to bethanechol, especially for basal tone. The M(3) receptor antagonists were generally more potent than the M(2) receptor antagonists. In a protection experiment, an M(3) receptor antagonist was less potent than when used in combination with an M(2) receptor antagonist. Receptor antagonists for M(1) and M(4) did not affect contractility variables. CONCLUSIONS AND CLINICAL RELEVANCE: Bethanechol acting on muscarinic receptor sub-types M(2) and M(3) may be of clinical use as a prokinetic drug for motility disorders of the duodenum and jejunum in dairy cows.     

Effect of short-chain fatty acids on contraction of smooth muscle in the canine colon

OBJECTIVE: To determine effects of short-chain fatty acids (SCFA) on canine colonic smooth muscle. SAMPLE POPULATION: Colonic tissue obtained from 14 healthy dogs. PROCEDURE: Short-chain fatty acid (SCFA; acetate, propionate, and butyrate; 1 to 100 mmol/L)-induced contractions were compared with responses obtained with acetylmethylcholine (AMCh; 10(-4) mol/L). Roles of enteric neurons, cholinergic receptors, calcium stores in the sarcoplasmic reticulum, and extracellular calcium in the SCFA-induced responses were investigated by incubating muscle strips with tetrodotoxin (1 micromol/L), atropine (1 micromol/L), ryanodine (10 micromol/L), nifedipine (1 micromol/L), ethylene glycol-bis (beta-aminoethylether)-N,N,N',N'-tetra-acetate (EGTA; 0.1 mmol/L), or an extracellular calcium-depleted (zero extracellular calcium) solution prior to the addition of propionate or butyrate. RESULTS: Incubation with SCFA elicited isometric stress responses (0.25 to 2.15 x 10(4) N/m2) in colonic longitudinal smooth muscle. Maximal responses to butyrate and propionate (50 mmol/L) were 37 and 23%, respectively, of the maximal AMCh response. Acetate was least effective in stimulating contractile responses. Tetrodotoxin and atropine did not affect SCFA-induced contractions. Nifedipine and zero extracellular calcium solution abolished responses to butyrate and propionate, whereas EGTA attenuated (> 60%) but did not abolish those responses. Ryanodine did not affect SCFA-induced contractile responses. The SCFA did not affect colonic circular smooth muscle. CONCLUSIONS AND CLINICAL RESPONSE: The SCFA stimulate longitudinal but not circular colonic smooth muscle contractions via a direct effect on smooth muscle. The mechanism of the SCFA effect appears to involve the influx of extracellular calcium. These findings may account for some of the effects of fiber on canine colonic motility [corrected].     

Effect of histamine on lung contractile elements in growing cattle

OBJECTIVE: To determine the effect of histamine on the contractile elements of the respiratory tract in neonatal calves and young adult cattle. SAMPLE POPULATION: Samples of trachealis muscle, bronchi, and intrapulmonary arteries and veins dissected from the respiratory tracts of healthy bovids (2 to 8 days and 16 to 20 months old). PROCEDURE: Histamine cumulative concentration-effect curves (10(-6) to 10(-3) M) were constructed in duplicate smooth muscle samples mounted in organ baths. Contractile responses to histamine were compared with reference contractions elicited by methacholine (10(-5) M) for airways or KCl (127 mM) for vessels. RESULTS: In young adult cattle, trachealis muscle had a substantial contractile response to histamine (84% of methacholine-induced contraction), whereas bronchi reacted slightly (15 and 20% for large and small bronchi, respectively). Although contractile responses to KCl were comparable in arteries and veins, histamine-induced contractions were greater for intrapulmonary veins than for arteries (202 vs 48% of KCl-induced contraction). In neonatal calves, histamine-induced contraction of veins also exceeded that of arteries (230 vs 54% of KCl-induced contraction); however, unlike in young adult cattle, histamine produced notable contraction of large and small bronchi (48 and 60% of methacholine-induced contraction, respectively). CONCLUSIONS AND CLINICAL RELEVANCE: Compared with intrapulmonary arteries, intrapulmonary veins have greater contractile responses to histamine in neonatal and young adult cattle. Data suggest loss of histamine responsiveness in bronchial smooth muscle as neonatal calves grow to young adults. Venodilation may be useful in treatment of lung edema in cattle.     

In vitro effects of bethanechol on equine gastrointestinal contractility and functional characterization of involved muscarinic receptor subtypes

The goal of this study was to investigate the effect of bethanechol (BeCh) on contractility patterns of smooth muscle preparations of equine duodenum descendens, jejunum, caecum and pelvic flexure in vitro. Concentration-response relationships were developed for BeCh using in vitro assays with and without preincubation of muscarinic (M) receptor antagonists for M2 and M3 receptors. BeCh induced a significant, concentration-dependent increase in contractile response in equine intestine in specimens with circular orientation. The maximal effect was largest for jejunal specimens with no difference in EC50 within the different locations investigated. The M2 antagonist, AF-DX 116, caused a rightward shift of the concentration-response curve and the M3 antagonist, 4-DAMP (1,1-dimethyl-4-diphenylacetoxypiperidinium iodide), almost completely inhibited the effect of BeCh over the entire concentration-response curve. These data provide evidence that, although the effect of BeCh is predominantly mediated by M3 receptors, M2 muscarinic receptors also play a role in BeCh-induced contraction in specimens of equine intestine. The involvement of other muscarinic receptor subtypes cannot be excluded. Further studies are necessary to understand the effect of BeCh in vivo including diseased animals.     

Response of equine airway smooth muscle to acetylcholine and electrical stimulation in vitro

Smooth muscle strips from the midcervical portion of the trachea and bronchial smooth muscle strips from third-generation airways of horses were placed in tissue baths, and isometric contractile force was measured. Active force was measured in response to electrical stimulation and exogenous acetylcholine. Square-wave electrical stimuli were applied at various voltages (10, 12, 15, 18, 20, 25 V), frequencies (3, 5, 10, 15, 20, 25, 30 Hz), and pulse durations (0.2, 0.5, 1.0, 1.5, 2.0 ms). Isometric contractile force increased as voltage, frequency, and pulse duration increased. Maximal contractile response to electrical stimulation was obtained at 18 V, 25 Hz, and 0.5 ms. Atropine (10(-6)M) or tetrodotoxin (3 x 10(-6)M) blocked the contraction, indicating that the contractile response was attributable to the release of neurotransmitter from cholinergic nerves. Cumulative concentration-response curves to acetylcholine (10(-9)M through 10(-4)M) were determined. Isometric contractile force increased as acetylcholine concentration increased. There was a significant (P less than 0.05) difference in the 50% effective dose for acetylcholine in tracheal smooth muscle and bronchial smooth muscle. The mean (+/- SD) contractile response to maximal electrical stimulus was 89% (+/- 7.4%) of that in response to 10(-4)M acetylcholine in tracheal smooth muscle and was 68% (+/- 10.4%) of the response to 10(-4)M acetylcholine in bronchial smooth muscle.     

Influence of the autonomic nervous system in the horse urinary bladder

alpha and beta-adrenergic receptors in detrusor muscle and bladder base of horses were investigated by in vitro responses of smooth muscle strips to exogenous agonist and antagonist drugs. Noradrenaline, isoprenaline and salbutamol induced relaxation of detrusor muscle strips which was significantly inhibited by propranolol and butoxamine suggesting that the response is mediated by beta-2 adrenergic receptors. In the urinary bladder base noradrenaline, phenylephrine and B-HT 920 induced strong contractile effects. These contractile responses were inhibited by the alpha antagonist phenoxybenzamine, the alpha-1 selective antagonist prazosin and the alpha-2 selective antagonist yohimbine. The inhibitory action of prazosin was more potent than that observed with yohimbine suggesting that the response in the bladder base of horses is mediated predominantly by alpha-1 adrenergic receptors, although alpha-2 receptors also participate.     

In vitro effects of bethanechol on smooth muscle preparations from abomasal fundus, corpus, and antrum of dairy cows

Abomasal displacement has been associated with gastric hypomotility. The supply of prokinetic drugs available to address this problem is insufficient. The goal of the study was to investigate the effect of the muscarinic agonist bethanechol (BeCh) on contractility parameters of smooth muscle preparations from several regions of the bovine abomasum (fundus, corpus, and antrum). Cumulative concentration-response curves were constructed using BeCh in vitro with and without pre-incubation with antagonists targeted at M(2) and M(3) muscarinic acetylcholine receptor (mAChR) subtypes. In all preparations investigated, BeCh induced a significant and concentration-dependent increase in all contractility parameters investigated. The maximal attainable effect (V(max)) was more pronounced in circular specimens, and V(max) of antral specimens in circular orientation were significantly lower when compared to the other preparations. Both antagonists caused a rightward shift of the concentration-response curve, suggesting that the effect of BeCh is mediated at least partly by M(2) and M(3) AChRs.     

Effects of Alcohol Extract of Aster tataricus L.f.on the Contraction of Guinea Pig Tracheal Smooth Muscle in vitro

The aim of the experiment was to study the effect of alcohol extract of Aster tataricus L.f.on the contraction of isolated guinea pig tracheal smooth muscle.The guinea pig tracheal smooth muscle was prepared by isolated tracheal thermostatic perfusion method.The effects of Aster tataricus L.f.on tracheal smooth muscle under the resting state and contraction stimulated by acetylcholine (Ach),histamine (His),CaCl₂,Ca²⁺ release and influx in cells without calcium were measured by BL-420F biological function experiment systems.The results showed that Aster tataricus L.f.could induce the contraction of isolated trachea at resting state at a low concentrations of 0.002 to 0.008 g/mL and exerted a relaxation on the isolated trachea when at the higher concentrations of 0.008 to 0.196 g/mL.All of the three concentrations of Aster tataricus L.f.could inhibit the tension stimulated by Ach,His and CaCl₂ which were antispasmodic agents,led to the non-parallelled rightward moving of cumulative concentration-response curve and the decline of maximal responses.The suppressive effect on His was the strongest,followed by Ach and CaCl₂,and the suppression could counteract the contraction induced by extracellular Ca²⁺influx significantly,and they were all expressed concentration-dependent manner.These results indicated the alcohol extract of Aster tataricus L.f.could relax tracheal smooth muscle by acting on M-receptor and histamine receptor and blocking Ca²⁺ passage,thus inhibiting extracellular Ca²⁺influx.     

紫菀乙醇提取物对豚鼠离体气管平滑肌收缩功能的影响

本试验旨在研究紫菀乙醇提物对豚鼠离体气管平滑肌收缩功能的影响。采用离体气管恒温灌流的方法,将豚鼠气管制成气管螺旋条,通过 BL-420F 生物机能实验系统测定其张力的变化,观察紫菀乙醇提物对离体气管平滑肌静息状态下的舒张作用,以及在氯化乙酰胆碱(Ach)、磷酸组胺(His)、CaCl₂条件下和无钙下Ach诱导细胞内钙释放和外钙内流所致两种收缩条件下对离体气管平滑肌张力变化的影响。试验结果显示,低浓度紫菀乙醇提取物(0.002~0.008 g/mL)对静息状态下豚鼠离体气管平滑肌具有一定的收缩作用,高浓度(0.008~0.196 g/mL)时具有舒张作用;低、中、高3个剂量组均可抑制Ach、His和CaCl₂引起的气管平滑肌收缩强度,使各致痉剂的量效曲线非平行右移并降低最大效应。紫菀乙醇提取物对His的抑制作用强度最大,其次为Ach,最后是CaCl₂,且均呈剂量依赖性。以上结果表明紫菀乙醇提物具有舒张气管平滑肌的作用,其机制可能与抑制豚鼠气管平滑肌M受体、H₁受体和阻断Ca²⁺通道从而抑制细胞Ca²⁺内流有关。     

Adrenergic, cholinergic, and nonadrenergic-noncholinergic intrinsic innervation of the jejunum in horses.

OBJECTIVE: To determine the major neurotransmitters that regulate contractile activity in the jejunum of horses. SAMPLE POPULATION: Jejunal specimens from 65 horses without gastrointestinal tract lesions. PROCEDURE: Jejunal smooth muscle strips, oriented in the plane of the circular or longitudinal muscular layer, were suspended isometrically in muscle baths. Neurotransmitter release was induced by electrical field stimulation (EFS) delivered at 30 and 70 V intensities and at various frequencies on muscle strips maintained at low or high muscle tone. To detect residual nonadrenergic-noncholinergic neurotransmission, the response of muscle to EFS in the presence of adrenergic and cholinergic blockade was compared with the response in the presence of tetrodotoxin. RESULTS: Atropine (ATR) decreased the contractile response of muscle strips to EFS under most conditions. However, ATR increased the contractile response of high-tone circular muscle. Adrenergic blockade generally increased the muscle responses to 30 V EFS and in high-tone longitudinal muscle but decreased contractile responses in high-tone circular muscle. Tetrodotoxin significantly altered the responses to EFS, compared with adrenergic and cholinergic receptor blockade. CONCLUSIONS: Acetylcholine and norepinephrine appear to be important neurotransmitters regulating smooth muscle contractility in the equine jejunum. They induce contraction and relaxation, respectively, in most muscle preparations, although they may cause opposite effects under certain conditions. In addition, nonadrenergic-noncholinergic excitatory and inhibitory influences were detected. CLINICAL RELEVANCE: Acetylcholine or norepinephrine release within the myenteric plexus of horses may alter gastrointestinal motility.     

奶牛输卵管平滑肌前列腺素类受体的分析研究

本研究选用了前列腺素类化合物(Prostaglandin E2（PGE2）、Prostaglandin F2α（PGF2α）、Prostaglandin D2（PGD2)）和受体选择性激动剂（Butaprost、U-46619）,用多道生理信号采集系统测定其对体外分离的奶牛输卵管峡部及壶腹部平滑肌收缩的影响,以此初步揭示受体的种类。结果发现,PGE2和Butaprost均抑制了奶牛输卵管峡部、壶腹部的自发性收缩运动,呈现明显的浓度依存性舒张反应;PGF2α在高浓度(1～10 μmol/L)下,使峡部和壶腹部的自发性收缩运动有了显著增强;PGD2在低浓度时轻微抑制了壶腹部平滑肌的收缩,但在高浓度(1～10 μmol/L)时加强了峡部和壶腹部平滑肌的收缩;U-46619从低浓度便引起了输卵管峡部和壶腹部平滑肌的强烈收缩。结果表明,EP4、EP2、FP、DP和TP受体同时存在于奶牛输卵管平滑肌上,能与内源性前列腺素类结合并参与调节输卵管平滑肌的自发性收缩。     

Evaluation of substance P as a neurotransmitter in equine jejunum

OBJECTIVE: To determine whether substance P (SP) functions as a neurotransmitter in equine jejunum. SAMPLE POPULATION: Samples of jejunum obtained from horses that did not have lesions in the gastrointestinal tract. PROCEDURE: Jejunal smooth muscle strips, oriented in the plane of the circular or longitudinal muscle, were suspended isometrically in muscle baths. Neurotransmitter release was induced by electrical field stimulation (EFS) delivered at 2 intensities (30 and 70 V) and various frequencies on muscle strips that were maintained at low tension or were under contraction. A neurokinin-1 receptor blocker (CP-96,345) was added to baths prior to EFS to interrupt SP neurotransmission. Additionally, direct effects of SP on muscle strips were evaluated, and SP-like immunoreactivity was localized in intestinal tissues, using indirect immunofluorescence testing. RESULTS: Substance P contracted circularly and longitudinally oriented muscle strips. Prior treatment with CP-96,345 altered muscle responses to SP and EFS, suggesting that SP was released from depolarized myenteric neurons. Depending on orientation of muscle strips and stimulation variables used, CP-96,345 increased or decreased the contractile response to EFS. Substance P-like immunoreactivity was detected in the myenteric plexus and circular muscle layers. CONCLUSIONS AND CLINICAL RELEVANCE: Substance P appears to function as a neurotransmitter in equine jejunum. It apparently modulates smooth muscle contractility, depending on preexisting conditions. Effects of SP may be altered in some forms of intestinal dysfunction. Altering SP neurotransmission in the jejunum may provide a therapeutic option for motility disorders of horses that are unresponsive to adrenergic and cholinergic drugs.     

5-Hydroxytryptamine potentiates contraction mediated by the intramural cholinergic nerve in the longitudinal smooth muscle of the ruminant forestomach

The effects of 5-hydroxytryptamine (5-HT) on the longitudinal smooth muscle from the rumen and reticulum of the bovine forestomach were investigated. 5-HT (0.25–490 μM) caused a contraction and a relaxation of the ruminal strips while it produced only an excitatory effect on the reticular strips. These effects were not affected by tetrodotoxin, hexamethonium, atropine or morphine, but were blocked by methysergide, LSD-25 or phenoxybenzamine. 5-HT potentiated the contraction evoked by stimulation of the intramural cholinergic nerves but did not show any effect on the relaxation produced by the non-adrenergic inhibitory nerves' excitation. The 5-HT-induced potentiation was not affected by morphine, LSD-25, methysergide and hexamethonium or high concentration of nicotine. Nicotine and dimethylphenylpiperazinium also caused a transient augmentation of the nerve-mediated contraction, but these effects were abolished by the competitive ganglionic blockers. The evoked contraction was depressed in high-Mg²⁺ solution, but this depression was antagonized partly by 5-HT. The affinity of the cholinomimetics to post-synaptic muscarinic receptor was not affected by 5-HT. It is concluded that contractions or relaxations of bovine forestomach strips induced by 5-HT are mediated through activation of D-receptors in the smooth muscle, and the 5-HT-induced potentiation of the evoked contraction may be elicited through presynaptic neural effects of 5-HT on the cholinergic nerves.     

Effects of a selective and a nonselective muscarinic cholinergic antagonist on heart rate and intestinal motility in dogs

The effects of methoctramine, a cardioselective muscarinic cholinergic antagonist, on heart rate and small intestinal motor activity were compared to those of the nonselective competitive muscarinic antagonist, atropine. Methoctramine or atropine, 6, 10, 30, 60 μg/kg, or sterile isotonic saline, was administered intravenously to six conscious dogs in cross-over studies. Methoctramine administration caused dose-dependent tachycardia without affecting intestinal motility, while atropine administration caused dose-dependent tachycardia accompanied by significant reductions in small intestinal motility. Additionally, methoctramine did not inhibit intestinal smooth muscle contractile activity initiated by the muscarinic agonist bethanechol, while atropine inhibited bethanechol-induced contractile activity in a dose-dependent manner. Calculated dosages of methoctramine and atropine required to produce a 50% increase in heart rate over baseline were 35.1 ± 5.3 and 39.5 ± 6.2 μg/kg, respectively. This dosage of atropine caused a 93 ± 13.9% reduction in intestinal motility. These findings suggest that selective muscarinic antagonists may be useful drugs for those veterinary patients in which nonselective muscarinic antagonists have the potential to produce untoward effects on intestinal motility.     

Cisapride Stimulates Contraction of Idiopathic Megacolonic Smooth Muscle in Cats

We have previously shown that cisapride, a substituted piperidinyl benzamide, stimulates contraction of healthy feline colonic smooth muscle. The purpose of the present investigation was to determine the effect of cisapride on feline idiopathic megacolonic smooth muscle function. Longitudinal smooth muscle strips from ascending and descending colon were obtained from cats with idiopathic megacolon, suspended in a 1.5 mM Ca²⁺-HEPES buffer solution (37°C, 100% O₂, pH 7.4), attached to isometric force transducers, and stretched to optimal muscle length (L₀). Control responses were obtained at each muscle site with acetylcholine (10–⁸ to 10 ⁴ M), substance P (10–¹¹ to 10^-7 M), or potassium chloride (10 to 80 mM). Muscles were then stimulated with cumulative (10–⁹ to 10–⁶ M) doses of cisapride in the absence or presence of tetrodotoxin (10–⁶ M) and atropine (10–⁶ M), or in a 0 calcium HEPES buffer solution. In cats with idiopathic megacolon, cisapride stimulated contractions of longitudinal smooth muscle from both the ascending and the descending colon. Cisapride-induced contractions were similar in magnitude to those induced by substance P and acetylcholine in the ascending colon, but were less than those observed in the descending colon. Cisapride-induced contractions in megacolonic smooth muscle were only partially inhibited by tetrodotoxin and atropine, but were virtually abolished by removal of extracellular calcium. We concluded that cisapride-induced contractions of feline megacolonic smooth muscle are largely smooth muscle mediated and dependent on influx of extracellular calcium. Cisapride-induced contractions in megacolonic smooth muscle are only partially dependent on enteric cholinergic nerves. Thus, cisapride may be useful in the treatment of cats with idiopathic megacolon.     

Estrous cycle-dependent differences in responsiveness to prostaglandins and contractile agents in sheep (Ovis aries) cervical smooth muscle

We investigated the influence of the phase of the estrous cycle on mechanical responses elicited in sheep cervix by potassium chloride (KCl), acetylcholine chloride (ACh), prostaglandin F2 alpha (PGF2 alpha) and prostaglandin E1 (PGE1). The cervix of adult ewes (n = 48) were classified according to the presence or absence of corpora lutea (luteal or follicular phase, respectively). Muscle strips of the circular and longitudinal layers were prepared in an organ bath and coupled to an isometric force transducer. Concentration-response curves were obtained noncumulatively. KCl and ACh produced concentration-dependent contractions in all preparations in both phases of the estrous cycle. However, maximum effect, EC50 and slope values of KCl and ACh were not significantly different between muscle layers, as well as between the phases of the estrous cycle. The prostanoid, PGF2 alpha, produced a significant reduction in the amplitude of spontaneous contractions for all preparations. The depressant effect of PGF2 alpha on spontaneous contractions of circular smooth muscle was significantly greater during the follicular than the luteal phase, whilst the depressant effect of PGF2 alpha on the longitudinal layer did not differ between phases of the estrous cycle. PGE1 significantly reduced the amplitude of spontaneous contractions on circular but not on longitudinal preparations. In conclusion, we have characterized with in vitro preparations of circular and longitudinal muscle layers of ewes during the follicular and luteal phases of the estrous cycle, the parameters of the K- and ACh-induced contractions on cervix and the efficacy of PGF2 alpha and PGE1 on inhibition spontaneous contractile activity.     

Effects of Pasteurella haemolytica culture supernate on bovine tracheal smooth muscle.

Pasteurella haemolytica leukotoxin is a ruminant specific leukotoxin that has been implicated in the pathogenesis of shipping fever in cattle. The present study was undertaken to determine the effect of this toxin on bovine airway smooth muscle. In vitro, the addition of culture supernate containing leukotoxin to bovine tracheal smooth muscle resulted in contraction of 55% of the muscle strips tested. Maximum responses were reached rapidly during cumulative additions of this material. In 95% of the muscle strips that responded, maximum responses were obtained after the addition of one or two cumulative doses. Repeated additions of culture supernate resulted in decreased responsiveness. Since responsiveness to other agonists was not affected, these results suggest the development of a condition similar to tachyphylaxis. The contractions were inhibited by antihistamines. Diphenhydramine, at a concentration of 10(-6) M (dose-ratio 7), and mepyramine, at a concentration of 2 x 10(-7) M (dose-ratio 56), blocked the contractions by 84% and 100% respectively. In addition, the contractions were blocked by the muscarinic antagonist atropine, but this inhibition was much weaker (46%) and was present at high concentrations only. Inhibition of the contractions by H1 receptor antagonists suggests that the contractions are mediated via H1 receptors. Since the dose-response relationship is not typical of a drug-receptor interaction, it appears unlikely that the leukotoxin is a direct agonist of H1 receptors. It is proposed that an indirect mechanism of action involving the release of histamine by tissue mast cells is responsible for the leukotoxin-induced contractions.