Effect of lung inflation in vivo on airways with smooth muscle tone or edema

Brown, Robert H., Wayne Mitzner, Yonca Bulut, and ElizabethM. Wagner. Effect of lung inflation in vivo on airways with smoothmuscle tone or edema. J. Appl.Physiol. 82(2): 491-499, 1997.Fibrousattachments to the airway wall and a subpleural surrounding pressurecan create an external load against which airway smooth muscle mustcontract. A decrease in this load has been proposed as a possible causeof increased airway narrowing in asthmatic individuals. To study theinteraction between the airways and the surrounding lung parenchyma, weinvestigated the effect of lung inflation on relaxed airways, airwayscontracted with methacholine, and airways made edematous by infusion ofbradykinin into the bronchial artery. Measurements were made inanesthetized sheep by using high-resolution computed tomography tovisualize changes in individual airways. During methacholine infusion,airway area was decreased but increased minimally with increases intranspulmonary pressure. Bradykinin infusion caused a 50% increase inairway wall area and a small decrease in airway luminal area. Incontrast to airways contracted with methacholine, the luminal areaafter bradykinin increased substantially with increases intranspulmonary pressure, reaching 99% of the relaxed area at totallung capacity. Thus airway edema by itself did not prevent fulldistension of the airway at lung volumes approaching total lungcapacity. Therefore, we speculate that if a deep inspiration fails torelieve airway narrowing in vivo, this must be a manifestation ofairway smooth muscle contraction and not airway wall edema.

     

Reflex changes in tracheal smooth muscle tone during high-frequency oscillation

We examined the effect of high-frequency oscillatory ventilation (HFOV) on tracheal smooth muscle tension and upper airway resistance in anesthetized dogs. The animals were ventilated via a low tracheostomy by HFOV or conventional intermittent positive pressure ventilation (IPPV) with and without added positive end-expiratory pressure (PEEP). The transverse muscle tension of the trachea above the tracheostomy was measured and found to be lower during HFOV when compared with IPPV or IPPV with PEEP. When both vagi were cooled to 8 degrees C to interrupt afferent traffic from the lungs, there was no longer any difference between the modes of ventilation. In a second series of experiments, the airflow resistance of the upper airway above the tracheostomy was measured (Ruaw). During HFOV, Ruaw was significantly lower than during either IPPV or IPPV with PEEP. We conclude that HFOV induces a relaxation of tracheal smooth muscle and a reduction of upper airway resistance through a vagally mediated mechanism.     

Effect of inflation on trachealis muscle tone in canine tracheal segments in vitro

Isolated tracheal segments were studied in vitro to determine how inflation affects the length and tension of the contracted and relaxed trachealis muscle. Circumferential trachealis muscle lengths were measured from cross-sectional radiographs taken during stepwise inflation of intact 20-cm-long tracheal segments to an inflation pressure of 25 cmH2O. A tracheal length spanning two cartilage rings was then cut out and mounted in a tissue bath using clips attached at the points of muscle insertion into the cartilage. The ring was stretched open along the axis of the muscle, and the resulting forces of the relaxed and contracted muscle and the cartilage were measured. Muscle lengths and tensions during inflation of the trachea were determined by comparing pressure vs. length and force vs. length measurements. During inflation from 0 to 25 cmH2O, the circumferential length of the trachealis muscle contracted with 10(-5) M acetylcholine increased from 48 to 70% of its length of maximal active tension (Lmax), while the relaxed muscle increased from 80 to 93% Lmax. The length of the contracted muscle was maintained at a nearly constant proportion of its relaxed length at each pressure.     

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.     

Endogenous AA metabolites and their possible role in tracheal smooth muscle tone in guinea pigs

The effects of endogenous arachidonic acid (AA) metabolites on inherent tone and histamine-induced constriction were studied in guinea pig tracheal smooth muscle. Inhibitors of either cyclooxygenase (indomethacin) or lipoxygenase (AA 861) significantly diminished the inherent tone of the muscle. Antagonists of prostaglandins (SC 19220) or leukotrienes (FPL 55712) also diminished the inherent tone, whereas an inhibitor of thromboxane synthase (OKY 046) had no significant effect. These results show that the metabolites of the lipoxygenase pathway as well as prostaglandins also participate in the maintenance of inherent tone. To reexamine the previously reported augmentation of histamine constriction induced by the inhibitors and the antagonists, we compared the active tension of the muscle measured from the maximum relaxed level as the base line to eliminate the fluctuation of inherent tone. Such comparison revealed that the inhibitors and the antagonists have no augmentative effect on either the maximum response to histamine or the concentration required to produce 50% of maximum active tension and that there is functional synergism between the exogenously added histamine and the endogenously produced AA metabolites. Therefore the zero active tension is useful as a base line to compare the contractile response of a drug-treated preparation with that of a nontreated preparation.     

Effect of Na-K adenosinetriphosphatase activity on relaxation of canine tracheal smooth muscle

The effect of Na-K adenosinetriphosphatase (ATPase) on relaxation induced by isoproterenol, prostaglandin E2, sodium nitroprusside, and forskolin, a specific stimulant of adenylate cyclase, was investigated in canine tracheal smooth muscle strips. Relaxation in response to isoproterenol, prostaglandin E2, and forskolin was significantly decreased after inhibition of the Na-K ATPase by ouabain or a potassium-free medium, but relaxation to sodium nitroprusside was not affected. Relaxation to isoproterenol was greater in muscles contracted by 5-hydroxytryptamine than in those contracted by acetylcholine. The stimulation of Na-K ATPase activity with potassium also caused differences in relaxation between tissues contracted with 5-hydroxytryptamine or acetylcholine. Relaxation caused by isoproterenol by activation of the Na-K-ATPase was also decreased by the Ca2+-channel antagonists, verapamil and diltiazem. The results suggest 1) Na-K ATPase activity modulates relaxation caused by isoproterenol, prostaglandin E2, and forskolin in canine tracheal smooth muscle, 2) isoproterenol or activation of the Na-K ATPase may cause relaxation partly by reducing Ca2+ influx through potential-dependent Ca2+ channels, and 3) the differences in the inhibitory effects of isoproterenol and Na-K ATPase activity on muscles contracted by acetylcholine and 5-hydroxytryptamine could be due to differences between these contractile agents in their dependence on extracellular Ca2+ for activation.     

Effect of 1,1-dimethylphenyl 1,4-piperazinium on mouse tracheal smooth muscle responsiveness

Bronchial hyperresponsiveness is one of the main features of asthma. A nicotinic receptor agonist, 1,1-dimethylphenyl 1,4-piperazinium (DMPP), has been shown to have an inhibitory effect on airway response to methacholine in an in vivo model of asthma. The aims of this study were to 1) verify whether nicotinic acetylcholine receptors (nAChR) were present on mouse tracheal smooth muscle, 2) verify whether bronchoprotection observed in mice was due to a direct effect on airway smooth muscle, and 3) compare the effects of nicotinic agonists to that of salbutamol. Alpha3-, alpha4-, and alpha7-nAChR subunits were detected by immunofluorescence on tracheal tissues from normal BALB/c mice. The effect of DMPP on tracheal responsiveness was verified by an isometric method. Tracheas were isolated from normal mice, placed in organ baths, and contracted with a single dose of methacholine. Cumulative doses of DMPP or salbutamol were added to the baths. Results show that mouse tracheal smooth muscle is positive for alpha4- and alpha7-nAChR subunits and that the epithelium is positive for alpha3-, alpha4-, and alpha7-subunits. DMPP induced a greater dose-dependent relaxation of tracheal smooth muscles precontracted with methacholine than with salbutamol. These results suggest that the smooth muscle-relaxing effect of DMPP could have some interest in the treatment of obstructive pulmonary diseases.     

Airway smooth muscle orientation in intraparenchymal airways

Lei, M., H. Ghezzo, M. F. Chen, and D. H. Eidelman.Airway smooth muscle orientation in intraparenchymal airways.J. Appl. Physiol. 82(1): 70-77, 1997.Airway smooth muscle (ASM) shortening is the central eventleading to bronchoconstriction. The degree to which airway narrowingoccurs as a consequence of shortening is a function of both themechanical properties of the airway wall as well as the orientation ofthe muscle fibers. Although the latter is theoretically important, ithas not been systematically measured to date. The purpose of this studywas to determine the angle of orientation of ASM () in normal lungs by using a morphometric approach. We analyzed the airway tree of theleft lower lobes of four cats and one human. All material was fixedwith 10% buffered Formalin at a pressure of 25 cmH₂O for 48 h. The fixed materialwas dissected along the airway tree to permit isolation ofgenerations 4-18 in the cats andgenerations 5-22 in the humanspecimen. Each airway generation was individually embedded in paraffin.Five-micrometer-thick serial sections were cut parallel to the airwaylong axis and stained with hematoxylin-phloxine-saffron. Each blockyielded three to five sections containing ASM. To determine , wemeasured the orientation of ASM nuclei relative to the transverse axisof the airway by using a digitizing tablet and a light microscope (×250) equipped with a drawing tube attachment. Inspection of thesections revealed extensive ASM crisscrossing without a homogeneous orientation. The  was clustered between 20° and 20°in all airway generations and did not vary much between generations inany of the cats or in the human specimen. When  was expressedwithout regard to sign, the mean values were 13.2° in the cats and13.1° in the human. This magnitude of obliquity is not likely toresult in physiologically important changes in airway length duringbronchoconstriction.

     

Inspiratory rhythm in airway smooth muscle tone

In anesthetized paralyzed open-chested cats ventilated with low tidal volumes at high frequency, we recorded phrenic nerve activity, transpulmonary pressure (TPP), and either the tension in an upper tracheal segment or the impulse activity in a pulmonary branch of the vagus nerve. The TPP and upper tracheal segment tension fluctuated with respiration, with peak pressure and tension paralleling phrenic nerve activity. Increased end-tidal CO2 or stimulation of the carotid chemoreceptors with sodium cyanide increased both TPP and tracheal segment tension during the increased activity of the phrenic nerve. Lowering end-tidal CO2 or hyperinflating the lungs to achieve neural apnea (lack of phrenic activity) caused a decrease in TPP and tracheal segment tension and abolished the inspiratory fluctuations. During neural apnea produced by lowering end-tidal CO2, lung inflation caused no further decrease in tracheal segment tension and TPP. Likewise, stimulation of the cervical sympathetics, which caused a reduction in TPP and tracheal segment tension during normal breathing, caused no further reduction in these parameters when the stimulation occurred during neural apnea. During neural apnea the tracheal segment tension and TPP were the same as those following the transection of the vagi or the administration of atropine (0.5 mg/kg). Numerous fibers in the pulmonary branch of the vagus nerve fired in synchrony with the phrenic nerve. Only these fibers had activity which paralleled changes in TPP and tracheal tension. We propose that the major excitatory input to airway smooth muscle arises from cholinergic nerves that fire during inspiration, which have preganglionic cell bodies in the ventral respiratory group in the region of the nucleus ambiguus and are driven by the same pattern generators that drive the phrenic and inspiratory intercostal motoneurons.     

Effects of tracheal irritation and hypercapnia on tracheal smooth muscle in humans

Both hypercapnia and tracheal irritation are known to constrict the airways in animals. To see whether similar responses occur in humans, we investigated tracheal smooth muscle (TSM) responses to hypercapnia and tracheal irritation with water in 14 paralyzed and anesthetized humans. TSM tone was monitored by measuring the pressure in the saline-filled cuff of the endotracheal tube. Although, tracheal irritation caused TSM constriction in 10 of 14 patients, 4 patients showed no TSM response. Administration of intravenous atropine attenuated the TSM constriction response. Hypercapnia did not cause any change in TSM tone in any of the 14 patients. These results indicate that in paralyzed and anesthetized humans, there exist interindividual differences in the TSM responses to tracheal irritation and that hypercapnia cannot be an effective stimulus for the TSM constriction.     

Respiratory epithelium inhibits bronchial smooth muscle tone

The aim of the present study was to determine whether or not the respiratory epithelium can modulate the responsiveness of bronchial smooth muscle. Paired rings of canine bronchi (4-6 mm OD), in some of which the epithelium had been removed mechanically (by rubbing the luminal surface), were mounted in physiological saline solution, gassed with 95% O2-5% CO2, and maintained at 37 degrees C. The presence or absence of the epithelium was confirmed by histological examination. Removal of the epithelium increased the contractile responses evoked by acetylcholine, histamine, and 5-hydroxytryptamine. Transmural nerve stimulation evoked similar peak responses in the presence and absence of epithelium. In unrubbed preparations, the peak response was followed by a gradual decrease when the stimulation was continued. This decrease, which persisted in the presence of propranolol, was not observed in epithelium-denuded preparations. In bronchial rings contracted with acetylcholine, isoproterenol produced concentration-dependent relaxations which were significantly greater in rings with epithelium compared with denuded rings. These results suggest that respiratory epithelial cells may generate an inhibitory signal to decrease the responsiveness of bronchial smooth muscle to contractile agonists and augment the effectiveness of inhibitory stimuli.     

Parasympathetic innervation of canine tracheal smooth muscle

To investigate whether efferent parasympathetic fibers to the trachealsmooth muscle course through the pararecurrent nerve rather than therecurrent or the superior laryngeal nerve, we stimulated all threenerves in anesthetized dogs. We also recorded the pararecurrentnerve activity response to bronchoconstrictor stimuli and compared itwith pressure changes inside a saline-filled cuff of an endotrachealtube. Electrical stimulation (30 s, 100 Hz, 0.1 ms, 10 mA) increasedtracheal cuff pressure by 21.0 ± 3.2 and 1.3 ± 0.7 cmH₂O for the pararecurrent and the recurrent laryngealnerve, respectively. Stimulation of the superior laryngeal nerveincreased tracheal cuff pressure before, but not after, sectioning ofthe ramus anastomoticus, which connects it to the pararecurrent nerve.Intravenous administration of sodium cyanide increased pararecurrentnerve activity by 208 ± 51% and tracheal cuff pressure by14.4 ± 3.5 cmH₂O. Elevation of end-tidalPCO₂ to 50 Torr increased pararecurrent nerveactivity by 49 ± 19% and tracheal cuff pressure by 8.4 ± 3.6 cmH₂O. Further elevation to 60 Torr increasedpararecurrent nerve activity by 101 ± 33% and tracheal cuffpressure by 11.3 ± 2.9 cmH₂O. These results lead usto the conclusion that parasympathetic efferent fibers reach the smoothmuscle of the canine trachea via the pararecurrent nerve.

     

Effects of extracellular calcium on canine tracheal smooth muscle

Strips of canine tracheal smooth muscle were studied in vitro to determine the effects of changes in the extracellular calcium (Cao) concentration on tonic contractions induced by acetylcholine and 5-hydroxytryptamine. Strips were contracted with graded concentrations of the above agents in 2.4 mM Ca, after which CaCl2 was administered to achieve final concentrations of 5.0, 10.0, and 20.0 mM. Increases in Cao to 5 mM or above caused significant relaxation of muscles contracted with 5-hydroxytryptamine but did not significantly relax muscles contracted with acetylcholine. Increases in Cao also caused significant relaxation of muscles contracted with low concentrations of K+ (20 or 30 mM). However, in 60 or 120 mM K+, increases in Cao resulted predominantly in muscle contraction. Inhibition of the Na+-K+-ATPase by ouabain (10(-5) M) or K+ depletion reversed the effects of Cao from relaxation to contraction in tissues contracted with 5-hydroxytryptamine. Increases in Cao also caused contraction rather than relaxation in the presence of verapamil (10(-6) M). We conclude that calcium has both excitatory and inhibitory effects on the contractile responses of canine tracheal smooth muscle. The inhibitory effects of Ca2+ appear to be linked to the activity of the membrane Na+-K+-ATPase.     

Reflex regulation of airway smooth muscle tone.

Autonomic nerves in most mammalian species mediate both contractions and relaxations of airway smooth muscle. Cholinergic-parasympathetic nerves mediate contractions, whereas adrenergic-sympathetic and/or noncholinergic parasympathetic nerves mediate relaxations. Sympathetic-adrenergic innervation of human airway smooth muscle is sparse or nonexistent based on histological analyses and plays little or no role in regulating airway caliber. Rather, in humans and in many other species, postganglionic noncholinergic parasympathetic nerves provide the only relaxant innervation of airway smooth muscle. These noncholinergic nerves are anatomically and physiologically distinct from the postganglionic cholinergic parasympathetic nerves and differentially regulated by reflexes. Although bronchopulmonary vagal afferent nerves provide the primary afferent input regulating airway autonomic nerve activity, extrapulmonary afferent nerves, both vagal and nonvagal, can also reflexively regulate autonomic tone in airway smooth muscle. Reflexes result in either an enhanced activity in one or more of the autonomic efferent pathways, or a withdrawal of baseline cholinergic tone. These parallel excitatory and inhibitory afferent and efferent pathways add complexity to autonomic control of airway caliber. Dysfunction or dysregulation of these afferent and efferent nerves likely contributes to the pathogenesis of obstructive airways diseases and may account for the pulmonary symptoms associated with extrapulmonary disorders, including gastroesophageal reflux disease, cardiovascular disease, and rhinosinusitis.     

Neural control of relaxation in cat airways smooth muscle

Functional innervation of cat airways smooth muscle was examined in isolated segments of trachea and bronchi using electrical field stimulation (EFS) techniques. Field stimulation caused contraction in tissues at resting tone and biphasic responses (contraction followed by relaxation) in tissues precontracted with 5-hydroxytryptamine (5-HT). Contractions were abolished by 10(-6) M atropine. Inhibitory responses were dependent on impulse voltage, duration, and frequency. At low voltages (less than or equal to 10 V) and pulse durations (less than or equal to 0.3 ms), EFS induced relaxations were abolished by 3 X 10(-6) M tetrodotoxin (TTX). Greater stimulus parameters elicited TTX-resistant relaxations. Pretreatment of the tissues with 10(-6) M propranolol and 10(-5) M guanethidine caused rightward shifts in relaxation frequency-response curves. These findings indicate that cat airways are innervated by excitatory cholinergic, inhibitory adrenergic, and inhibitory nonadrenergic noncholinergic (NANC) nerves. Pretreatment of the tissues with hexamethonium, cimetidine, indomethacin, or nordihydroguaiaretic acid did not affect NANC relaxation responses. It is concluded that NANC inhibitory responses in cat airway smooth muscle are mediated through intrinsic postganglionic nerve fibers and occur independently of histamine H2-receptor activation and without involvement of cyclooxygenase or lipoxygenase products of arachidonic acid metabolism.