Responsiveness of the isolated airway during simulated deep inspirations: effect of airway smooth muscle stiffness and strain.

In vivo, breathing movements, including tidal and deep inspirations (DIs), exert a number of beneficial effects on respiratory system responsiveness in healthy humans that are diminished or lost in asthma, possibly as a result of reduced distension (strain) of airway smooth muscle (ASM). We used bronchial segments from pigs to assess airway responsiveness under static conditions and during simulated tidal volume oscillations with and without DI and to determine the roles of airway stiffness and ASM strain on responsiveness. To simulate airway dilations during breathing, we cycled the luminal volume of liquid-filled segments. Volume oscillations (15 cycles/min) were set so that, in relaxed airways, they produced a transmural pressure increase of approximately 5-10 cmH(2)O for tidal maneuvers and approximately 5-30 cmH(2)O for DIs. ACh dose-response curves (10(-7)-3 x 10(-3) M) were constructed under static and dynamic conditions, and maximal response and sensitivity were determined. Airway stiffness was measured from tidal trough-to-peak pressure and volume cycles. ASM strain produced by DI was estimated from luminal volume, airway length, and inner wall area. DIs produced substantial ( approximately 40-50%) dilation, reflected by a decrease in maximal response (P < 0.001) and sensitivity (P < 0.05). However, the magnitude of bronchodilation decreased significantly in proportion to airway stiffening caused by contractile activation and an associated reduction in ASM strain. Tidal oscillations, in comparison, had little effect on responsiveness. We conclude that DI regulates airway responsiveness at the airway level, but this is limited by airway stiffness due to reduced ASM strain.     

Relationship of airway narrowing, compliance, and cartilage in isolated bronchial segments.

Structural components of the airway wall may act to load airway smooth muscle and restrict airway narrowing. In this study, the effect of load on airway narrowing was investigated in pig isolated bronchial segments. In some bronchi, pieces of cartilage were removed by careful dissection. Airway narrowing was produced by maximum electrical field stimulation. An endoscope was used to record lumen narrowing. The compliance of the bronchial segments was determined from the cross-sectional area of the lumen and the transmural pressure. Airway narrowing and the velocity of airway narrowing were increased in cartilage-removed airways compared with intact control bronchi. Morphometric assessment of smooth muscle length showed greater muscle shortening to acetylcholine in cartilage-removed airways than in controls. Airway narrowing was positively correlated with airway compliance. Compliance and area of cartilage were negatively correlated. These results show that airway narrowing is increased in compliant airways and that cartilage significantly loads airway smooth muscle in whole bronchi.     

Decreased airway narrowing and smooth muscle contraction in hyperresponsive pigs.

Increased smooth muscle contractility or reduced smooth muscle mechanical loads could account for the excessive airway narrowing and hyperresponsiveness seen in asthma. These mechanisms were investigated by using an allergen-induced porcine model of airway hyperresponsiveness. Airway narrowing to electric field stimulation was measured in isolated bronchial segments, over a range of transmural pressures (0-20 cmH(2)O). Contractile responses to ACh were measured in bronchial segments and in isolated tracheal smooth muscle strips isolated from control and test (ovalbumin sensitized and challenged) pigs. Test airways narrowed less than controls (P < 0.0001). Test pigs showed reduced contractility to ACh, both in isolated bronchi (P < 0.01) and smooth muscle strips (P < 0.01). Thus isolated airways from pigs exhibiting airway hyperresponsiveness in vivo are hyporesponsive in vitro. The decreased narrowing in bronchi from hyperresponsive pigs may be related to decreased smooth muscle contractility. These data suggest that mechanisms external to the airway wall may be important to the hyperresponsive nature of sensitized lungs.     

Cyclical elongation regulates contractile responses of isolated airways.

Bronchoconstrictor responses are quantitatively different when they are evoked under static conditions and during or after periods of deep inspiration. In vivo, deep inspirations produce bronchodilation and protect the lung from subsequent bronchoconstriction (termed bronchoprotection). These effects may be due in part to dynamic stretch on airways produced by cyclical expansion of airway diameter. However, airways also lengthen cyclically during breathing. The effects of cyclical airway elongation on evoked bronchoconstriction have not been examined. This study recorded evoked contractions of pig bronchial segments 1) at different airway lengths, 2) after a period of cyclical lengthening in relaxed airways, and 3) during cyclical lengthening in pretoned airways. Airway segments were mounted in organ baths and bathed in Krebs solution luminally and on the adventitia. Airways were cyclically lengthened by 5-30% of their deflated length at 0.5-2 Hz for 5 min. Contractions were evoked by electrical field stimulation or carbachol and were recorded under isovolumic conditions. Under static conditions, there was a blunt relationship between length and response to electrical field stimulation. After a period of airway length cycling, electrical field stimulation-induced contractions were increased. In airways pretoned with carbachol, cyclical lengthening produced a transient bronchodilation and a sustained increase in contraction. Contractile responses were not blocked by indomethacin. The results show that isolated airways respond actively to dynamic changes in length. Our results indicate that cyclical lengthening of airways could contribute to lung function in vivo but does not appear to account for the phenomenon of bronchoprotection.     

Maintenance of airway caliber in isolated airways by deep inspiration and tidal strains.

Deep inspirations (DIs) are large periodic breathing maneuvers that regulate airway caliber and prevent airway obstruction in vivo. This study characterized the intrinsic response of the intact airway to DI, isolated from parenchymal attachments and other in vivo interactions. Porcine isolated bronchial segments were constricted with carbachol and subjected to transmural pressures of 5-10 cmH2O at 0.25 Hz (tidal breathing) interspersed with single DIs of amplitude 5-20 cmH2O, 5-30 cmH2O, or 5-40 cmH2O (6-s duration) or DI of amplitude 5-30 cmH2O (30-s duration). Tidal breathing was ceased after DI in a subset of airways and in control airways in which no DI was performed. Luminal cross-sectional area was measured using a fiber-optic endoscope. Bronchodilation by DI was amplitude dependent; 5-20 cmH2O DIs produced less dilation than 5-30 cmH2O and 5-40 cmH2O DIs (P=0.003 and 0.012, respectively). Effects of DI duration were not significant (P=0.182). Renarrowing after DI followed a monoexponential decay function to pre-DI airway caliber with time constants between 27.4+/-4.3 and 36.3+/-6.9 s. However, when tidal breathing was ceased after DI, further bronchoconstriction occurred within 30s. This response was identical in both the presence and absence of DI (P=0.919). We conclude that the normal bronchodilatory response to DI occurs as a result of the direct mechanical effects of DI on activated ASM in the airway wall. Further bronchoconstriction occurs by altering the airway wall stress following DI, demonstrating the importance of continual transient strains in maintaining airway caliber.     

Parenchymal interdependence and airway response to methacholine in excised dog lobes

The objective of this investigation was to determine the minimum transpulmonary pressure (PL) at which the forces of interdependence between the airways and the lung parenchyma can prevent airway closure in response to maximal stimulation of the airways in excised canine lobes. We first present an analysis of the relationship between PL and the transmural pressure (Ptm) that airway smooth muscle must generate to close the airways. This analysis predicts that airway closure can occur at PL less than or equal to 10 cmH2O with maximal airway stimulation. We tested this prediction in eight excised canine lobes by nebulizing 50% methacholine into the airways while the lobe was held at constant PL values ranging from 25 to 5 cmH2O. Airway closure was assessed by comparing changes in alveolar pressure (measured by an alveolar capsule technique) and pressure at the airway opening during low-amplitude oscillations in lobar volume. Airway closure occurred in two of the eight lobes at PL = 10 cmH2O; in an additional five it occurred at PL = 7.5 cmH2O. We conclude that the forces of parenchymal interdependence per se are not sufficient to prevent airway closure at PL less than or equal to 7.5 cmH2O in excised canine lobes.     

Respiratory system responsiveness in rabbits in vivo is reduced by prolonged continuous positive airway pressure.

Active, nonanesthetized, tracheotomized rabbits were subjected to continuous positive airway pressure (CPAP) for 4 days to determine the effects of chronic mechanical strain on lung and airway function. Rabbits were maintained for 4 days at a CPAP of 6 cmH(2)O (high CPAP), at a CPAP of 0 cmH(2)O (low CPAP), or without tracheostomy (no CPAP). After treatment with CPAP, changes in respiratory resistance in response to increasing concentrations of inhaled ACh were measured during mechanical ventilation to evaluate respiratory system responsiveness in vivo. Intraparenchymal bronchial segments were isolated from the lungs of all animals to evaluate airway smooth muscle responsiveness and bronchial compliance in vitro. Rabbits maintained for 4 days at high CPAP demonstrated significantly lower responsiveness to ACh compared with rabbits that were maintained at low CPAP or with no CPAP. Airways isolated from the lungs of animals subjected to the chronic application of high CPAP were also less responsive to ACh in vitro than the airways isolated from animals subjected to low CPAP or no CPAP. The persistence of the decreased responsiveness in the excised airway tissues suggests that the decreased respiratory system responsiveness observed in vivo results primarily from direct effects on the airways. The results demonstrate that the application of prolonged mechanical strain in vivo can reduce airway reactivity.     

Effect of deep inspiration avoidance on ventilation heterogeneity and airway responsiveness in healthy adults

The mechanisms by which deep inspiration (DI) avoidance increases airway responsiveness in healthy subjects are not known. DI avoidance does not alter respiratory mechanics directly; however, computational modeling has predicted that DI avoidance would increase baseline ventilation heterogeneity. The aim was to determine if DI avoidance increased baseline ventilation heterogeneity and whether this correlated with the increase in airway responsiveness. Twelve healthy subjects had ventilation heterogeneity measured by multiple-breath nitrogen washout (MBNW) before and after 20 min of DI avoidance. This was followed by another 20-min period of DI avoidance before the inhalation of a single methacholine dose. The protocol was repeated on a separate day with the addition of five DIs at the end of each of the two periods of DI avoidance. Baseline ventilation heterogeneity in convection-dependent and diffusion-convection-dependent airways was calculated from MBNW. The response to methacholine was measured by the percent fall in forced expiratory volume in 1 s/forced vital capacity (FVC) (airway narrowing) and percent fall in FVC (airway closure). DI avoidance increased baseline diffusion-convection-dependent airways (P = 0.02) but did not affect convection-dependent airways (P = 0.9). DI avoidance increased both airway closure (P = 0.002) and airway narrowing (P = 0.02) during bronchial challenge. The increase in diffusion-convection-dependent airways due to DI avoidance did not correlate with the increase in either airway narrowing (r(s) = 0.14) or airway closure (r(s) = 0.12). These findings suggest that DI avoidance increases diffusion-convection-dependent ventilation heterogeneity that is not associated with the increase in airway responsiveness. We speculate that DI avoidance reduces surfactant release, which increases peripheral ventilation heterogeneity and also predisposes to peripheral airway closure.     

In vivo dimensional response of airways of different size to transpulmonary pressure.

We studied four supine dogs that were anesthetized with pentobarbital, intubated, and ventilated with a piston pump. The dimensional response of central (CAW) (greater than 2 mm diam) and peripheral airways (PAW) (smaller than 2 mm diam) to changes in transpulmonary pressure (Ptp) was determined by progressive increments in tidal volume (VT). A specially designed electronics relay circuit permitted this relationship to be obtained for points of no flow during tidal volume breathing: i.e., preinspiration (FRC); end inspiration (FRC + VT). The airways were dusted with powdered tantalum. Six airway divisions were identified: four CAW: trachea, main stem, lobar, segmental; and two PAW: subsegmental, and lobular. AP and lateral roentgenograms were obtained by standard technics and primary magnification (mag factor 2). Airway diameters were plotted as a function of transpulmonary pressure between 3 and 26 cmH2O with the diameter at total lung capacity expressed as 100%. The data show that: 1) there is significant distensibility above 5 cmH2O for all airways from the trachea to the lobular airways; 2) that the pressure-diameter plot is a linear plot for each airway from 3 to 26 cmH2O with R values between 0.846 and 0.957; 3) the peripheral lobular airways are more distensible than the central airways (P smaller than 0.05). We attribute the difference in distensibility of the peripheral lobular airways to their lack of cartilaginous support, and their decreased muscular support when compared to the CAW.     

Activation of smooth muscle in the airway wall, force production, and airway narrowing.

Airway narrowing depends on smooth muscle force production and muscle shortening, but the structural and geometric properties exhibited by individual generations of the bronchial tree largely determine the extent and characteristics of airway narrowing. Properties of major importance include the nature and integrity of the epithelium, the structural and mechanical properties of the airway wall, as well as airway diameter. The influence of these properties on airway narrowing measured as flow or flow resistance in large and small diameter segments of airways from pig lung is described using a novel preparation, the perfused bronchial segment.     

Reduction in airway hyperresponsiveness to methacholine by the application of RF energy in dogs.

We delivered controlled radio frequency energy to the airways of anesthetized, ventilated dogs to examine the effect of this treatment on reducing airway narrowing caused by a known airway constrictor. The airways of 11 dogs were treated with a specially designed bronchial catheter in three of four lung regions. Treatments in each of the three treated lung regions were controlled to a different temperature (55, 65, and 75 degrees C); the untreated lung region served as a control. We measured airway responsiveness to local methacholine chloride (MCh) challenge before and after treatment and examined posttreatment histology to 3 yr. Treatments controlled to 65 degrees C as well as 75 degrees C persistently and significantly reduced airway responsiveness to local MCh challenge (P < or = 0.022). Airway responsiveness (mean percent decrease in airway diameter after MCh challenge) averaged from 6 mo to 3 yr posttreatment was 79 +/- 2.2% in control airways vs. 39 +/- 2.6% (P < or = 0.001) for airways treated at 65 degrees C, and 26 +/- 2.7% (P < or = 0.001) for airways treated at 75 degrees C. Treatment effects were confined to the airway wall and the immediate peribronchial region on histological examination. Airway responsiveness to local MCh challenge was inversely correlated to the extent of altered airway smooth muscle observed in histology (r = -0.54, P < 0.001). We conclude that the temperature-controlled application of radio frequency energy to the airways can reduce airway responsiveness to MCh for at least 3 yr in dogs by reducing airway smooth muscle contractility.     

Intraluminal pressure oscillation enhances subsequent airway contraction in isolated bronchial segments.

A period of deep inspiration in humans has been shown to attenuate subsequent bronchoconstriction, a phenomenon termed bronchoprotection. The bronchoprotective effect of deep inspiration may be caused though a depression in the force production of airway smooth muscle (ASM). We determined the response of whole airway segments and isolated ASM to a period of cyclic stretches. Isovolumetric contraction to electrical field stimulation (EFS) was assessed in porcine bronchial segments before and after intraluminal pressure oscillation from 5 to 25 cmH(2)O for 10 min at 0.5 Hz. Morphometry showed that this pressure oscillation stretched ASM length by 21%. After pressure oscillation, the response to EFS was not reduced but instead was modestly enhanced (P < 0.01). Airway responses to EFS returned to preoscillation levels 10 min after the end of oscillation. The increase in EFS response after pressure oscillation was not altered by the addition of indomethacin. In a separate experiment, we assessed isometric force in isolated ASM strips before and after length oscillation. The amplitude, frequency, and duration of length oscillation were similar to those induced in bronchial segments. In contrast to bronchial segments, length oscillation of ASM produced a significant depression in isometric force induced by EFS (P < 0.01). These results suggest that the response of ASM to length oscillation is modified by the airway wall. They also suggest that the phenomenon of bronchoprotection reported in some in vivo studies may not be an intrinsic property of the airway.     

Effects of tidal volume stretch on airway constriction in vivo.

Tidal stresses are thought to be involved in maintaining airway patency in vivo. The present study examined the effects of normal stresses exerted by the lung parenchyma during tidal ventilation on recovery from agonist-induced airway constriction. In seven anesthetized dogs, one lung was selectively ventilated with a Univent endotracheal tube (Vitaid, Lewiston, NY). Airway tone was increased either transiently (intravenous bolus) or continuously (intravenous infusion) with methacholine (MCh). During one-lung ventilation, changes in the airway size of both lungs were measured for up to 40 min during recovery from constriction by using high-resolution computed tomography. After recovery to baseline, the alternate lung was ventilated, and the protocol was repeated. The absence of tidal stresses led to an attenuated recovery from either transient or steady-state airway constriction. The effectiveness or lack thereof of normal tidal stress in stabilizing airway size may be one factor that contributes to the lack of reversal with tidal breathing and deep inspiration seen in asthmatic subjects.     

Prostaglandin F2 alpha increases responsiveness of pulmonary airways in dogs

We studied the effect of prostaglandin F2 alpha (PGF2 alpha) on the responsiveness of pulmonary airways in dogs. Airway responsiveness was assessed by determining the bronchoconstrictor response to increasing concentrations of acetylcholine aerosol delivered to the airways. In each of five dogs, we determined responsiveness during treatment with physiologic saline, histamine, or PGF2 alpha aerosols. The doses of histamine and PGF2 alpha were determined by establishing the largest dose of each which could be given to the dog without causing bronchoconstriction (subthreshold doses). We found that airway responsiveness was not significantly different during histamine treatment than after saline, however, responsiveness increased during treatment with PGF2 alpha. In addition, the hyperresponsiveness induced by PGF2 alpha was prevented by pretreatment with the ganglion blocking drug hexamethonium (5 mg/kg given intravenously). The results show that PGF2 alpha specifically increases the responsiveness of pulmonary airways in doses that do not cause bronchoconstriction, and suggest that the hyperresponsiveness involves a neural mechanism such as increased responsiveness of airway sensory nerves.     

In vivo estimation of tracheal distensibility and hysteresis in normal adults

We used the acoustic reflection technique to measure the cross-sectional area of tracheal and bronchial airway segments of eight healthy adults. We measured airway area during a slow continuous expiration from total lung capacity (TLC) to residual volume (RV) and during inspiration back to TLC. Lung volume and esophageal pressure were monitored continuously during this quasi-static, double vital capacity maneuver. We found that 1) the area of tracheal and bronchial segments increases with increasing lung volume and transpulmonary pressure, 2) the trachea and bronchi exhibit a variable degree of hysteresis, which may be greater or less than that of the lung parenchyma, 3) extrathoracic and intrathoracic tracheal segments behaved as if they were subjected to similar transmural pressure and had similar elastic properties, and 4) specific compliance (means +/- SE) for the intrathoracic and bronchial segments, calculated with the assumption that transmural pressure is equal to the transpulmonary pressure, was significantly (P less than 0.05) smaller for the intrathoracic segment than for the bronchial segment: (2.1 +/- 2.0) X 10(-3) cmH2O-1 vs. (9.1 +/- 2.1) X 10(-3) cmH2O-1. Direct measurements of airway area using acoustic reflections are in good agreement with previous estimates of airway distensibility in vivo, obtained by radiography or endoscopy.     

Significance of thromboxane generation in ozone-induced airway hyperresponsiveness in dogs

To determine whether thromboxane A2 may be involved in ozone (O3)-induced airway hyperresponsiveness, we studied the effect of a thromboxane synthase inhibitor (OKY-046, 100 micrograms X kg-1 X min-1 iv) in five dogs exposed to O3. Airway responsiveness was assessed by determining the provocative concentration of acetylcholine aerosol that increased total pulmonary resistance by 5 cmH2O X l-1 X s. O3 (3 ppm) increased airway responsiveness as demonstrated by a decrease in acetylcholine provocative concentration from 2.42 (geometric SEM = 1.64) to 0.14 mg/ml (geometric SEM = 1.30). OKY-046 significantly inhibited this effect without altering pre-O3 responsiveness or the O3-induced increase in neutrophils and airway epithelial cells in bronchoalveolar lavage fluid. To further examine the role of thromboxane A2, we studied the effect of a thromboxane A2 mimetic, U-46619, on airway responsiveness in five additional dogs. U-46619 in subthreshold doses (i.e., insufficient to increase base-line pulmonary resistance) caused a fourfold increase in airway responsiveness to acetylcholine. Subthreshold doses of histamine had no effect. These results suggest that thromboxane A2 may be an important mediator of O3-induced airway hyperresponsiveness.     

Glutamate receptors and the airways hyperreactivity

It is proposed the link between the hyperactivity of NMDA receptors and airway hyperresponsiveness. We investigated the effect of agents modulating the activity of NMDA receptors in the ovalbumin-induced airway hyperreactivity in guinea pigs. The airways hyperreactivity was influenced by the agonist (NMDA) and selective antagonist - competitive (AP-5) and non-competitive (MK-801) of NMDA receptors. Airway responsiveness to histamine or acetylcholine was evaluated in in vitro conditions. NMDA administration caused the increase of tracheal smooth muscle response in ovalbumin-induced hyperreactivity to acetylcholine. MK 801 as well as AP-5 provoked the decrease of reactivity mainly to acetylcholine in tracheal smooth muscle, while the former, non-competitive antagonist was more effective. We recorded more pronounced response in tracheal than in lung tissue smooth muscle with more considerable response to acetylcholine than to histamine. The results of experiments show the modification of airway smooth muscles responses by agents modulating the activity of NMDA receptors. They confirm the possibility of NMDA receptors participation in experimental airway hyperreactivity. The results enlarge information regarding the link of the inflammatory diseases and glutamatergic system.     

Association between reduced bronchodilatory effect of deep inspiration and loss of alveolar attachments

Background

We have previously shown that the bronchodilatory effect of deep inspiration is attenuated in individuals with COPD. This study was designed to investigate whether the impairment in this effect is associated with loss of alveolar attachments.

Methods

We measured deep inspiration (DI)-induced bronchodilation in 15 individuals with and without COPD (67 ± 2.2 yrs of age, mean ± SEM) undergoing lobar resection for peripheral pulmonary nodule. Prior to surgery, we measured TLCO and determined the bronchodilatory effect of deep inspiration after constricting the airways with methacholine. The number of destroyed alveolar attachments, as well as airway wall area and airway smooth muscle area, were determined in tumor-free, peripheral lung tissue.

Results

The bronchodilatory effect of deep inspiration correlated inversely with the % destroyed attachments (r = -0.51, p = 0.05) and directly with the airway smooth muscle area (r = 0.59, p = 0.03), but not with the total wall area (r = 0.39, p = 0.15).

Conclusion

We postulate that attenuation of airway stretch due to loss of alveolar attachments contributes to the loss of the bronchodilatory effect of lung inflation in COPD.     

Effects of pulmonary congestion on airway reactivity to histamine aerosol in dogs

We examined the effect of acute pulmonary vascular congestion on bronchial reactivity in dogs in a standard challenge protocol. Airway responsiveness to histamine whose concentration was varied in a stepwise incremental fashion was assessed from changes in pulmonary resistance (RL) and dynamic compliance (Cdyn) in 10 anesthetized dogs. Brief acute pulmonary congestion was created by inflating a balloon placed in the left atrium to raise left atrial pressure to 20-30 cmH2O for 1 min. Pulmonary congestion did not change RL in the control condition. However, after histamine inhalation, RL was further increased by pulmonary congestion, making the two effects synergistic. This phenomenon could not be observed with vagi cut. Pulmonary congestion decreased Cdyn in all dogs regardless of histamine concentration, with or without vagotomy. We conclude that pulmonary vascular congestion makes the bronchi hyperreactive through vagal reflexes. The reduction in Cdyn caused by pulmonary congestion appears to stem mainly from the narrowing of peripheral airways by adjacent vascular engorgement.     

Selected contribution: airway caliber in healthy and asthmatic subjects: effects of bronchial challenge and deep inspirations.

In 9 healthy and 14 asthmatic subjects before and after a standard bronchial challenge and a modified [deep inspiration (DI), inhibited] bronchial challenge and after albuterol, we tracked airway caliber by synthesizing a method to measure airway resistance (Raw; i.e., lung resistance at 8 Hz) in real time. We determined the minimum Raw achievable during a DI to total lung capacity and the subsequent dynamics of Raw after exhalation and resumption of tidal breathing. Results showed that even after a bronchial challenge healthy subjects can dilate airways maximally, and the dilation caused by a single DI takes several breaths to return to baseline. In contrast, at baseline, asthmatic subjects cannot maximally dilate their airways, and this worsens considerably postconstriction. Moreover, after a DI, the dilation that does occur in airway caliber in asthmatic subjects constricts back to baseline much faster (often after a single breath). After albuterol, asthmatic subjects could dilate airways much closer to levels of those of healthy subjects. These data suggest that the asthmatic smooth muscle resides in a stiffer biological state compared with the stimulated healthy smooth muscle, and inhibiting a DI in healthy subjects cannot mimic this.