Potentiation of vagal contractile response by thromboxane mimetic U-46619

We studied the effect of the thromboxane mimetic U-46619 on tracheal smooth muscle contraction caused by bilateral stimulation of the vagus nerves in 14 mongrel dogs in situ. The parasympathetic contractile response was studied isometrically after beta-adrenergic blockade with 2 mg/kg iv propranolol plus 20 micrograms X kg-1 X min-1 continuous intravenous infusion and blockade of endogenous prostaglandin synthesis with 5 mg/kg iv indomethacin. An initial frequency-response curve was generated by electrical stimulation of the caudal ends of cut cervical vagi over the range of frequencies 2-25 Hz (constant 25 V) at 15-s intervals. In five dogs, 10(-10) to 10(-8) mol of the thromboxane mimetic (15S)-hydroxyl-11 alpha,9 alpha-(epoxymethano)prosta-5Z,13E-dienoic acid (U-46619) was injected selectively into the tracheal arterial circulation, causing a transient contractile response (less than or equal to 10 g/cm). Additional frequency response studies were generated 7 min before and 1, 15, 30, 45, and 60 min after U-46619. Substantial augmentation of tracheal contraction to efferent vagal stimulation was observed after U-46619 for all frequencies greater than 4 Hz (P less than 0.02). Augmentation of vagally mediated contraction was not observed in four other dogs after equivalent tracheal contraction was elicited without U-46619. Similarly, in four separate dogs, augmentation of tracheal contraction was not observed when acetylcholine was given instead of vagal stimulation after U-46619. We conclude that the thromboxane analogue, U-46619, causes augmentation of tracheal contractile response induced by efferent vagus nerve stimulation. Potentiation is caused by a prejunctional action of U-46619 and is not induced by nonspecific precontraction with another agonist.     

Effect of resting smooth muscle length on contractile response in resistance airways

We studied the effect of resting smooth muscle length on the contractile response of the major resistance airways (generations 0-5) in 18 mongrel dogs in vivo using tantalum bronchography. Dose-response curves to 10(-10) to 10(-7) mol/kg methacholine (MCh) were generated [at functional residual capacity (FRC)] by repeated intravenous bolus administration using tantalum bronchography after each dose. Airway constriction varied substantially with dose-equivalent stimulation and varied sequentially from trachea (8.8 +/- 2.2% change in airway diam) to fifth-generation bronchus (49.8 +/- 3.0%; P less than 0.001). Length-tension curves were generated for each airway to determine the airway diameter (i.e., resting in situ smooth muscle length) at which maximal constriction was elicited using bolus intravenous injection of 10(-8) mol/kg MCh. A Frank-Starling relationship was obtained for each airway; the transpulmonary pressure at which maximal constriction was elicited increased progressively from 2.50 +/- 1.12 cmH2O for trachea (approximately FRC) to 18.3 +/- 1.05 cmH2O for fifth-generation airways (approximately 50% TLC) (P less than 0.001). A similar relationship was obtained when change in airway diameter was plotted as a function of airway radius. We demonstrate substantial heterogeneity in the lung volumes at which maximal constriction is elicited and in distribution of parasympathomimetic constriction within the first few generations of resistance bronchi. Our data also suggest that lung hyperinflation may lead to augmented airway contractile responses by shifting resting smooth muscle length toward optimum resting smooth muscle length.     

PAF-induced contraction of canine trachea mediated by 5-hydroxytryptamine in vivo

We studied the secretory correlates of tracheal smooth muscle contraction caused by platelet-activating factor (PAF) in nine mongrel dogs in vivo. In five dogs, dose-response curves were generated by rapid intra-arterial injection of 10(-10) to 10(-6) mol PAF into the isolated tracheal circulation; tracheal contractile response was measured isometrically in situ. To examine the mechanism by which PAF elicits contraction of canine trachealis, concentrations of serotonin (5-HT) and histamine were assayed in the venous effluent as the arteriovenous difference (AVd) in mediator concentration across the airway for each level of contraction. PAF caused dose-related active tracheal tension to a maximum of 37.2 +/- 5.4 g/cm (10(-6) mol PAF). The AVd in 5-HT increased linearly from 0.20 +/- 0.05 (10(-9) mol PAF) to 3.5 +/- 0.3 ng/ml (10(-6) mol PAF) (P less than 0.005). In contrast, the AVd in histamine was insignificant and did not change with increasing doses of PAF. A positive correlation was obtained between the AVd in 5-HT and active tracheal tension (r = 0.92, P less than 0.001); there was no correlation between AVd in histamine and active tension (r = -0.16). PAF-induced parasympathetic activation was not mediated by 5-HT; contraction elicited by exogenous 5-HT was not affected by muscarinic blockade. We conclude that nonparasympathetically mediated contraction elicited acutely by PAF in dogs results at least in part from secondary release of serotonin and is not mediated by histamine.     

Bronchial smooth muscle mechanics of a canine model of allergic airway hyperresponsiveness.

Although we have reported that tracheal smooth muscle from sensitized dogs shows altered mechanical properties, we did not know, because of technical difficulties with the preparation, whether similar changes occur in the properties of sensitized central bronchial smooth muscle (BSM), the site at which the acute asthmatic response is believed to develop. We have now succeeded in developing a cartilage-free BSM preparation that retains optimal mechanical properties. Such strips were obtained from mongrel dogs that had been sensitized to ragweed pollen. Controls were littermates injected with adjuvant alone. Length-tension relationships were obtained for both control and sensitized BSM strips (CBSM and SBSM, respectively). The maximal active stresses were the same (P greater than 0.05) when normalized to muscle fraction in total tissue cross-sectional area [6.2 +/- 0.6 x 10(4) and 5.9 +/- 0.6 x 10(4) (SE) for SBSM and CBSM, respectively]. This suggests that optimal tension is an insensitive indicator of bronchial hyperresponsiveness and that isotonic studies might be more revealing. The maximal shortening velocity (Vo) for SBSM at 2 s [0.35 +/- 0.017 (SE) lo/s, where lo signifies optimal muscle length], in the course of a 10-s contraction, was significantly greater (P less than 0.05) than Vo measured for CBSM (0.27 +/- 0.015 lo/s). However, Vo did not differ at the 8-s point of contraction. The sensitized group demonstrated a statistically significantly greater maximal shortening capacity (0.67 +/- 0.04 lo) than the control group (0.51 +/- 0.04 lo). At 2 s of contraction, 80% of maximal SBSM shortening had been completed and was significantly greater than for CBSM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Augmentation of parasympathetic contraction in tracheal and bronchial airways by PGF2 alpha in situ

We studied the effect of exogenous prostaglandin F2 alpha (PGF2 alpha) on airway smooth muscle contraction caused by parasympathetic stimulation in 22 mongrel dogs in situ. Voltage (0-30 V, constant 20 Hz) and frequency-response (0-25 Hz, 25 V) curves were generated by stimulating the cut ends of both cervical vagus nerves. Airway response was measured isometrically as active tension (AT) in a segment of cervical trachea and as change in airway resistance (RL) and dynamic compliance (Cdyn) in bronchial airways. One hour after 5 mg/kg iv indomethacin, a cumulative frequency-response curve was generated in nine animals by electrical stimulation of the vagus nerves at 15-s intervals. Reproducibility was demonstrated by generating a second curve 7 min later. A third frequency-response curve was generated during active contraction of the airway caused by continuous intravenous infusion of 10 micrograms X kg-1 X min-1PPGF2 alpha. Additional frequency-response studies were generated 15 and 30 min after PGF2 alpha, when airway contractile response (delta RL = +2.8 +/- 0.65 cmH2O X 1(-1) X s; delta Cdyn = -0.0259 +/- 0.007 1/cmH2O) returned to base line. Substantial augmentation of AT, RL, and Cdyn responses was demonstrated in every animal studied (P less than 0.01 for all points greater than 8 Hz) 15 min after PGF2 alpha. At 30 min, response did not differ from initial base-line control. In four animals receiving sham infusion, all frequency-response curves were identical. We demonstrate that PGF2 alpha augments the response to vagus nerve stimulation in tracheal and bronchial airways. Augmentation does not depend on PGF2 alpha-induced active tone.     

Epithelial augmentation of trachealis contraction caused by major basic protein of eosinophils

We studied the effect of epithelial removal and intraepithelial administration of human eosinophil granule major basic protein (MBP) on the contraction of underlying canine tracheal smooth muscle in 23 dogs in vivo. A dual in situ tracheal preparation was utilized that allowed sharp excision of epithelium. The response to intra-arterial acetylcholine (ACh) was augmented substantially in five dogs receiving 200 micrograms MBP by intraepithelial instillation. Active tension elicited by 10(-8) mol intra-arterial ACh was 34.0 +/- 2.2 g/cm before and 46.1 +/- 2.6 g/cm 30 min after MBP (P less than 0.002). There was no change in active tension in the control segment in the same dogs after intraepithelial instillation of vehicle only (34.7 +/- 3.2 vs. 34.4 +/- 2.3 g/cm; P = NS). Instillation of MBP directly into the subepithelial tracheal smooth muscle did not alter contraction. To assess whether this augmentation was caused by inhibition of an epithelial-derived relaxant factor, additional studies were performed in nine other dogs in which the epithelium was excised discretely from one of the two tracheal segments. No significant differences in contractile response to ACh or relaxation response to isoproterenol were observed at 2, 15, 30, or 60 min after epithelial excision. We demonstrate that intraepithelial administration of MBP augments the contraction of underlying canine tracheal smooth muscle elicited by ACh. This augmentation is a direct effect of MBP and does not require antagonism of epithelial inhibition.     

Response of bronchial smooth muscle to mast cell degranulation in situ

We studied the response of bronchial smooth muscle to mast cell degranulation with Ascaris suum antigen (AA) and compound 48/80 (48/80) in 26 mongrel dogs in situ. Bronchial smooth muscle response was measured isometrically in situ from a segment of the right middle lobe bronchus; tracheal response was monitored isometrically as a control. After intra-arterial (ia) injection of AA into the bronchial circulation, bronchial contraction preceded tracheal contraction by 19.2 +/- 4.6 s (P less than 0.002). Bronchial contraction to AA (21.7 +/- 3.4 g) was substantially greater than to 48/80 (10.5 +/- 1.8 g, P less than 0.05) corresponding to differences in maximal systemic histamine concentrations (146 +/- 24.1 vs. 1000 +/- 236 ng/ml, P less than 0.01). In 5 dogs, the effect of leukotriene D4 (LTD4) and FPL 55712 was studied. Injection of 10(-8) mol ia LTD4 caused no bronchial contraction. In four dogs, 10(-7) mol FPL 55712 caused no bronchial relaxation after initial precontraction during immune degranulation with AA; intravenous chlorpheniramine (5 mg/kg) caused 69.7 +/- 9.4% relaxation. We demonstrate a model that permits selective immune degranulation of a single major resistance bronchus in situ. We conclude that AA-induced degranulation in dogs caused bronchial contraction predominantly by secretion of preformed mediator.     

Autonomic response characteristics of porcine airway smooth muscle in vivo

We studied the autonomic response characteristics of airways in 65 swine in vivo. Tracheal smooth muscle response was measured isometrically in situ; bronchial response was measured simultaneously as change in airway resistance and dynamic compliance. To determine the optimal resting length at which maximal tracheal contraction was obtained, length-tension studies were generated in four animals using maximal electrical stimulation of the vagus nerves determined from stimulus-response characteristics in eight other swine. Pharmacological studies were performed in 25 animals to determine the relative potency and intrinsic activity of agonists (acetylcholine greater than histamine much greater than norepinephrine) causing contraction of trachea and bronchial airways. In 13 swine, the effects of autonomic stimulation were studied by intravenous administration of dimethylphenylpiperazinium (DMPP) after muscarinic blockade with 1.5 mg/kg iv atropine. Tracheal contraction caused by topical application of 3.4 X 10(-4) mol histamine (13.4 +/- 1.54 g/cm) was 96 +/- 7.2% blocked by 25 micrograms/kg iv DMPP in adrenal-intact animals; minimal relaxation was demonstrated in adrenalectomized animals, indicating absence of substantial sympathetic innervation to porcine trachea. Nonadrenergic innervation was not demonstrated. After beta-adrenergic blockade, sympathetic stimulation caused alpha-adrenergic contraction in bronchial airways but not in trachea. These data define the unique response characteristics of the airways of swine and demonstrate their utility for acute experimental study of airway responses in vivo.     

Cholinergic neuromodulation by prostaglandin D2 in canine airway smooth muscle

To determine whether prostaglandin D2 (PGD2) modulates cholinergic neurotransmission in airway smooth muscle and, if so, what the mechanism of action is, we studied bronchial segments from dogs under isometric conditions in vitro. PGD2 (10(-8)-10(-5) M) elicited dose-dependent muscle contraction, which was reduced after blockade of muscarinic receptors, so that 50% effective dose (ED50) increased from 1.3 +/- 0.3 X 10(-6) to 3.9 +/- 1.0 X 10(-6) M by atropine (10(-6) M) (mean +/- SE, P less than 0.05). Physostigmine, at a concentration insufficient to alter base-line tension (10(-8) M), enhanced the PGD2-induced contraction and decreased ED50 to 6.4 +/- 0.5 X 10(-7) M (P less than 0.05). When added at the highest doses that did not cause spontaneous contraction (1.9 +/- 0.5 X 10(-7) M), PGD2 increased the contractile response to electrical field stimulation (1-50 Hz) by 21.9 +/- 6.6% (P less than 0.001). In contrast to this effect, the response to administered acetylcholine was not affected by PGD2. On the other hand, PGD2-induced augmentation of the response to electrical field stimulation (5 Hz) was further increased from 23.6 +/- 3.0 to 70.4 +/- 8.8% in the presence of physostigmine (10(-8) M) and was abolished by atropine but not affected by the alpha-adrenergic antagonist phentolamine or the histamine H1-blocker pyrilamine. These results suggest that the contraction of airway smooth muscle induced by PGD2 is in in part mediated by a cholinergic action and that PGD2 prejunctionally augments the parasympathetic contractile response, likely involving the accelerated release of acetylcholine at the neuromuscular junction.     

Ca2+-dependent facilitated shortening in isotonic contraction of trachealis muscle

We compared isotonic shortening with isometric force generation as a function of external Ca2+ in 166 tracheal smooth muscle (TSM) strips from 27 mongrel dogs in vitro. Concentration-response curves were generated with muscarinic stimulation (acetylcholine, ACh), alpha-adrenergic receptor activation (norepinephrine after beta-adrenoceptor blockade, NE), serotonin (5-HT), and KCl-substituted Krebs-Henseleit solution. The concentrations of 5-HT causing half-maximal shortening (ECS50, 1.54 +/- 0.14 X 10(-7) M) and half-maximal active isometric tension (ECT50, 1.72 +/- 0.30 X 10(-7) M) were similar (P = NS). Likewise, ECS50 (21.9 +/- 0.7 mM) and ECT50, (22.0 +/- 0.9 mM) were similar for KCl. In contrast, facilitated isotonic shortening (i.e., greater isotonic shortening for comparable degrees of force generation) was elicited with ACh and NE for all levels of force generation between 15 and 85% of maximum and for all concentrations of ACh from 3 X 10(-8) to 3 X 10(-5) M (P less than 0.05 for all points). Facilitated isotonic shortening also was elicited for all concentrations of NE from 10(-8) to 10(-6) M (P less than 0.05 for all points). Removal of Ca2+ from the perfusate substantially reduced the potency of ACh (P less than 0.001) and abolished differences between ECS50 (2.23 +/- 0.28 X 10(-5) M) and ECT50 (2.50 +/- 0.46 X 10(-5) M, P = NS). We demonstrate that for comparable degrees of force generation, muscarinic and alpha-adrenergic receptor activation cause greater isotonic shortening than KCl or 5-HT and that this facilitated shortening is associated with the concentration of external Ca2+.     

Neutral endopeptidase modulates endothelin-1-induced airway smooth muscle contraction in guinea-pig trachea.

This study was designed to evaluate the role of neutral endopeptidase (NEP) in modulating the airway smooth muscle contraction induced by endothelin-1 in isolated segments of guinea-pig trachea. Endothelin-1 (10(-9)-10(-6) M) produced a concentration-dependent contraction that reached a maximum by 30 min. The NEP inhibitor leucine-thiorphan (10(-5) M) significantly increased the contractile response to endothelin-1. The addition of leucine-thiorphan to tracheal segments precontracted by 10(-9) and 10(-8) M endothelin-1 increased isometric tension by 181 +/- 65% (mean +/- 1 S.E.M.; P less than 0.05) and by 138 +/- 49% (P less than 0.05), respectively. In contrast, the kininase II inhibitor captopril and the peptidase inhibitors leupeptin and bestatin had no effect. Preincubation of endothelin-1 with 1 microgram recombinant human NEP decreased the contractile activity of endothelin-1 by 72 +/- 9%, whereas no effect was observed using heat-inactivated NEP. We conclude that NEP modulates endothelin-induced contraction of airway smooth muscle in the guinea-pig trachea.     

Effect of M2 and M3 muscarinic receptors on airway responsiveness to carbachol in bronchial-hypersensitive (BHS) and bronchial-hyposensitive (BHR) guinea pigs.

The expression balance of M2 and M3 muscarinic receptor subtypes on the pathogenesis of airway hyperresponsiveness was investigated by using two congenitally related strains of guinea pigs, bronchial-hypersensitive (BHS) and bronchial-hyposensitive (BHR). CCh-induced airway responses in vivo and in vitro were investigated by comparing the effects of muscarinic receptor subtype antagonists, and the relative amounts of M2 and M3 muscarinic receptor mRNA in tracheal smooth muscle and lung tissue were investigated. After treatment with muscarinic receptor subtype antagonists, the ventilatory mechanics (VT, Raw, and Cdyn) of response to CCh aerosol inhalation were measured by the bodyplethysmograph method. The effects of these antagonists on CCh-induced tracheal smooth muscle contraction were also investigated. The effects of M2 muscarinic receptor blockade were less but the effects of M3 muscarinic receptors blockade on the airway contractile responses were greater in BHS than in BHR. In M3 muscarinic receptor blockades, CCh-induced tracheal contractions in BHS were significantly greater than those in BHR. In tracheal smooth muscle from BHS, the relative amount of M2 muscarinic receptors mRNA was less but that of M3 muscarinic receptor mRNA was more than those in BHR. These results suggest that the high ACh level as a consequence of dysfunction of M2 muscarinic autoreceptors and the excessive effect of M3 muscarinic receptors on the airway smooth muscle may play an important role in the pathogenesis of airway hyperresponsiveness.     

Physiological mechanisms mediating enhanced force generation during development and immune sensitization.

We examined the development of acetylcholinesterase (AChase) activity and tracheal smooth muscle (TSM) contraction elicited by acetylcholine (ACh) in a swine model of maturation and a dog model of allergic bronchospasm. Strips of TSM were tethered isometrically at optimal length and responses were expressed as a percentage of the maximum to KCl-substituted perfusate (% KCl). Maximal contraction (ATmax) to ACh in 2-week-old swine (168 +/- 8% KCl) was greater than in 10-week-old swine (142 +/- 2% KCl; p less than 0.02). The AChase inhibitor, physostigmine, augmented ACh-elicited ATmax in 10-week-old (27% increase; p less than 0.01) but not in 2-week-old swine (2% increase; p is NS) and caused a greater increase in sensitivity to muscarinic activation in 2 versus 10 week-old swine (p less than 0.02), thus demonstrating increased contraction of TSM in 2 versus 10-week-old swine, which results at least in part from reduced AChase activity in immature animals. In another study, TSM from ragweed-sensitized dogs demonstrated augmented efficacy to ACh-elicited contraction (180 +/- 6% KCl) compared with TSM from sham-sensitized, littermate controls (163 +/- 4% KCl; p less than 0.05). In the presence of physostigmine, ATmax was not different between ragweed-sensitized and control TSM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cyclic nucleotide function in trachealis muscle of dogs with and without airway hyperresponsiveness

We examined basal adenosine 3',5'-cyclic monophosphate (cAMP) levels, isoproterenol (ISO)-stimulated cAMP responses, basal cAMP, and guanosine 3',5'-cyclic monophosphate (cGMP) phosphodiesterase (PDE) activities and protein-kinase (PK) activities in trachealis muscle from five Basenji-greyhound (BG) and four greyhound dogs to determine whether the inverse relationship between in vivo and in vitro airway responsiveness could be due to altered cyclic nucleotide metabolism. Basal cAMP levels were not significantly different (PNS) in muscle from BG (11.6 +/- 0.53 pmol/mg protein) and greyhound dogs (10.30 +/- 1.60 pmol/mg protein). The cAMP responses to stimulation with ISO were enhanced in BG compared with greyhound dogs. The low Michaelis constant (1) for Km-cAMP PDE activity (Km = 0.63 microM) was significantly less (P less than 0.005) in BG dogs (1.54 +/- 0.28 pmol.min-1.mg protein-1) than greyhounds (11.76 +/- 2.48). Endogenously active PK activity was significantly greater (P less than 0.005) in BG (54.74 +/- 5.39 pmol.min-1.mg protein-1) than in greyhound dogs (15.50 +/0 2.20). Increases in PK activity with 5 microM cAMP added were not significantly different between BG (14.79 +/- 6.00) and greyhound dogs (7.04 +/- 2.14). Approximately 90% of both endogenous PK activity and cAMP-activated PK activity in BG and greyhound dogs was inhibited by a cAMP-dependent PK inhibitor (PKI'). These data suggest that decreased cyclic nucleotide degradation due to decreased cyclic nucleotide PDE activity with increased PK could account for the in vitro hyporesponsiveness of airway smooth muscle in BG dogs as a protective adaptive mechanism.     

Physiological antagonism caused by adrenergic stimulation of canine tracheal muscle

We studied the simultaneous alpha- and beta-adrenergic response characteristics of canine tracheal smooth muscle in 398 strips from 67 dogs in vitro. Experiments were performed to determine the effects of beta-adrenergic blockade on the expression of the alpha-adrenoceptor contractile responses elicited by norepinephrine (NE), phenylephrine (PE), and clonidine (CLO). Maximal active tension caused by NE increased from 39.1 +/- 27.0 to 241 +/- 75.0 g/cm2 as the concentration of propranolol (PROP) was increased from 10(-6) to 10(-4) M. Augmentation of tracheal smooth muscle contraction caused by PE and CLO was also observed with progressive beta-adrenoceptor blockade; contraction to NE, PE, and CLO was blocked selectively with 3 X 10(-5) M phentolamine (PA) and phenoxybenzamine (PBZ). The beta-adrenergic relaxing properties of the same three agonists were also studied. After alpha-adrenergic blockade with PA or PBZ, all three agonists caused relaxation (NE greater than CLO greater than PE) of methacholine-induced contraction of tracheal smooth muscle that was reversed selectively with PROP. We demonstrate that NE, PE, and CLO cause simultaneous stimulation of both the alpha- and beta-adrenergic receptors in tracheal smooth muscle; the net response elicited is the result of adrenergic physiological antagonism and depends on the relative degree of alpha- and/or beta-adrenoceptor blockade.     

Expression of airway contractile properties and acetylcholinesterase activity in swine

We studied the effect of maturation on contractile properties of tracheal smooth muscle from seventeen 2-wk-old swine (2ws) and fifteen 10-wk-old swine (10ws) in situ and in vitro. The response to parasympathetic stimulation was studied in situ in isometrically fixed segments. Contraction was elicited at lower frequencies [half-maximal response to electrical stimulation (ES50) = 6.7 +/- 0.05 Hz] in 2ws than in 10ws (ES50 = 9.1 +/- 0.4 Hz; P less than 0.01). Despite substantial differences in morphometrically normalized cross-sectional area in 2ws (0.012 +/- 0.003 cm2) and 10ws (0.028 +/- 0.001 cm2; P less than 0.01), maximal active tension elicited by parasympathetic stimulation was similar (12.4 +/- 3.2 g/cm in 2ws vs. 13.3 +/- 2.3 g/cm in 10ws; P = NS). In separate in vitro studies in 25 tracheal smooth muscle strips from 10 swine, concentration-response curves generated with potassium-substituted Krebs solution (KCl) were similar in 2ws and 10ws. In 58 other strips (10 swine), maximal active force elicited with acetylcholine (ACh) in 2ws was significantly greater than for 10ws (P less than 0.001). Removal of the epithelium had no effect. However, cholinesterase inhibition with 10(-7) M physostigmine augmented the response to ACh in 10ws (P less than 0.02) but not 2ws. We demonstrate increased force generation and sensitivity to vagal stimulation in 2ws vs. 10ws, which corresponds to increased reactivity to ACh in vitro. The relative hyperresponsiveness in 2ws is specific for cholinergic response and is attenuated at least in part by maturation of the activity of acetylcholinesterase enzyme.     

Prejunctional inhibitory muscarinic receptors on cholinergic nerves in human and guinea pig airways

We have investigated whether prejunctional inhibitory muscarinic receptors ("autoreceptors") exist on cholinergic nerves in human airways in vitro and whether guinea pig trachea provides a good model for further pharmacological characterization of these receptors. Pilocarpine was used as a selective agonist and gallamine as a selective antagonist of these autoreceptors. Acetylcholine (ACh) release from postganglionic cholinergic nerves was elicited by electrical field stimulation (EFS) (40 V, 0.5 ms, 32 Hz). In human bronchi, pilocarpine inhibited the contractile response to EFS in a dose-related fashion; the dose inhibiting 50% of the control contraction was 2.2 +/- 0.4 x 10(-7) (SE) M (n = 22), and the inhibition was 96% at 3 x 10(-5) M. The inhibitory effects of pilocarpine were antagonized by gallamine in a dose-related fashion. The results were qualitatively the same in the guinea pig. Gallamine significantly enhanced the contractile response to EFS in the guinea pig, whereas pirenzepine failed to do so, which suggests that M2-receptors are involved. We conclude that prejunctional muscarinic receptors that inhibit ACh release are present on cholinergic nerves in human airways and that guinea pig trachea is a good model for further pharmacological characterization of these receptors, which appear to belong to the M2-subtype.     

Beta-adrenergic relaxation of dog trachealis: contractile agonist-specific interaction

The phenomenon of contractile agonist-dependent relaxation by isoproterenol (ISO) of active tension elicited by acetylcholine (ACh), histamine (HIS), serotonin (5-HT), and potassium chloride-substituted Krebs-Henseleit solution (KCl) was studied in 210 tracheal smooth muscle (TSM) strips from 28 mongrel dogs in vitro. All TSM strips were contracted to similar active tensions [target tension (TT) = 50% of the maximal active tension elicited by 127 mM KCl] with ACh, HIS, 5-HT, or KCl and relaxed with either ISO, forskolin (FSK), N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (db-cAMP), or 3-isobutyl-1-methylxanthine (IMX). The concentrations of ISO causing 50% relaxation from TT (RC50) were ACh (2.9 +/- 1.1 x 10(-6) M) greater than 5-HT (8.4 +/- 1.5 x 10(-8) M) approximately KCl (8.1 +/- 2.1 x 10(-8) M) greater than HIS (1.6 +/- 0.2 x 10(-8) M). FSK and IMX relaxed TSM in the same rank order of potency as ISO. In contrast to the contractile agonist-dependent relaxation elicited by ISO, FSK, and IMX, db-cAMP was nearly equipotent in relaxing similarly contracted strips. These results are consistent with contractile agonist-specific interaction with cAMP production by ISO and FSK. These data demonstrate that the phenomenon of contractile agonist-dependent relaxation by ISO is not related specifically to the beta-adrenoceptor.     

A peptidergic component to vagally induced tracheal vasodilation in the dog.

The purpose of the study was to determine the extent that peptidergic afferent and efferent pathways contribute to vagally induced vasodilation in the trachea of the dog. The change in vascular resistance of the tracheal branch of the cranial thyroid artery and the trachealis responses were determined in 28 anesthetized, paralyzed, and mechanically ventilated dogs. After propranolol (2 mg/kg) and phentolamine (1.5 mg/kg), stimulation of the superior laryngeal nerves (NS; 15 Hz, 7 V, 2 ms, 30 s) caused a decrease in vascular resistance of 11.7 +/- 0.8% and a tracheal contraction of 5.2 +/- 4.7 cmH2O. Atropine (1.5 mg/kg) reduced the fall in vascular resistance to 4.7 +/- 0.8% (P less than 0.01), whereas tracheal contraction was abolished. Thiorphan (1.5 mg), a neutral endopeptidase inhibitor, augmented the decrease in vascular resistance (8.8 +/- 0.6%; P less than 0.01) to NS. After hexamethonium (0.5 mg/kg), NS still caused a small decrease in TVR (2.9 +/- 0.9%; P less than 0.05), which was abolished by capsaicin. In atropinized dogs, capsaicin reduced the fall in vascular resistance after NS; the residual vasodilation was virtually abolished by hexamethonium. Acetylcholine (10(-3) mg/kg) decreased vascular resistance (15.7 +/- 3.0%), and the effect was abolished by atropine. We conclude that there is noncholinergic nonadrenergic vagally induced tracheal vasodilation that is peptidergic. The peptidergic vasodilation appears to be mediated by both afferent and efferent pathways.     

Cyclooxygenase inhibitors increase canine tracheal muscle response to parasympathetic stimuli in situ

To determine whether cyclooxygenase inhibitors alter parasympathetic control of airway smooth muscle in situ, we pretreated anesthetized dogs with intravenous indomethacin, meclofenamate, or normal saline and measured the isometric contraction of tracheal muscle in response to electrical stimulation of the vagus nerves. Indomethacin and meclofenamate increase the response of airway smooth muscle to parasympathetic stimulation. In subsequent experiments to determine the site of action of cyclooxygenase inhibitors, we found that indomethacin does not alter the response of tracheal muscle to intra-arterial acetylcholine (a muscarinic agonist) but does augment the response to intra-arterial dimethylpiperaziniumiodide (a nicotinic agonist). Moreover, the response to parasympathetic stimulation after pretreatment with a combination of indomethacin and BW755C (a combined cyclooxygenase-lipoxygenase inhibitor) does not differ significantly from the response after indomethacin or meclofenamate alone. We conclude that cyclooxygenase inhibitors increase the sensitivity of the contractile response of tracheal smooth muscle to parasympathetic stimulation, that they exert their effect on the postganglionic parasympathetic neuron, and that their effect is prejunctional. The effect appears secondary to a decrease in cyclooxygenase products rather than to an increase in lipoxygenase products. These findings suggest that endogenous cyclooxygenase products may modulate parasympathetic control of airway smooth muscle in vivo. They may relate to the mechanisms that underlie airway hyperresponsiveness, by which mediators of inflammation modulate airway responsiveness and by which nonsteroidal anti-inflammatory drugs induce severe bronchoconstrictor responses in some persons who have asthma.