Interleukin-1beta and tumour necrosis factor-alpha increase microvascular leakage in the guinea pig trachea

OBJECTIVE: Airway microvascular leakage is considered to be an important component of airway inflammation in asthma. In the present study we examined the effect of interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNFalpha) on airway microvascular leakage in vivo. METHODOLOGY: Tracheal Evans blue extravasation was examined in an isolated tracheal segment, in anaesthetized mechanically ventilated guinea pigs. Baseline tracheal microvascular leakage was measured in five animals. As a control group for aerosol challenge, the isolated tracheal segment (n = 5) underwent saline aerosol challenge. To test whether a combination of IL-1beta (10 ng/mL) and TNFalpha (100 ng/mL) induced Evans blue extravasation, the trachea was exposed to an aerosol of these cytokines (n = 5). As a positive control the tracheal segment was challenged with histamine aerosol (5 x 10(-2) mol) (n = 3). All aerosol challenges were for 1 min. RESULTS: TNFalpha and IL-1beta aerosol challenge significantly increased Evans blue extravasation (28.9 +/- 1.6 microg/g wet tissue, mean +/- SE) compared to saline challenge (13.8 +/- 3.0 microg/g; P < 0.05). Tracheal dye extravasation without aerosol challenge, was not significantly different from saline-challenged animals (17.5 +/- 2.9 and 13.8 +/- 3.0 microg/g, respectively). Histamine significantly increased Evans blue extravasation (50.1 +/- 4.8 microg/g; P < 0.05) compared to saline challenge. CONCLUSION: Pro-inflammatory cytokines, TNFalpha and IL-1beta are able to induce significant microvascular leakage in the guinea pig trachea.     

Effect of antiasthma drugs on microvascular leakage in guinea pig airways

We have studied the effect of intravenous epinephrine, albuterol, verapamil, and aminophylline on airway microvascular leakage in guinea pigs. Microvascular leakage was induced by platelet-activating factor (PAF; 50 ng/kg intravenously), which acts directly on venular endothelial cells, and measured by quantifying extravasation of Evans blue (EB) dye. Epinephrine (20 micrograms/kg) inhibited PAF-induced changes in dye leakage in larynx and main bronchi; at 80 and 160 micrograms/kg, significant inhibition was observed in all airways studied. This effect was reversed by phentolamine (2.5 mg/kg) or prazosin (100 micrograms/kg). By contrast, albuterol (20 to 320 micrograms/kg) and aminophylline (12.5 to 50 mg/kg) failed to inhibit dye leakage at any dose studied. Verapamil inhibited PAF-increased leakage in larynx, main bronchi, and intrapulmonary airways at the lowest dose tested (125 micrograms/kg), although inhibition was not dose dependent. These results suggest that the antiedema effect of epinephrine may be due to vasoconstriction rather than to a direct effect on endothelial cell contractility and that neither beta-agonists nor theophylline have an inhibitory effect. The inhibitory effect of epinephrine on airway microvascular leakage may have therapeutic implications for asthma.     

热、酸刺激豚鼠食管诱导气道血浆渗出及其机制探讨

目的观察热及酸刺激豚鼠食管对气道的影响,探讨瞬时受体电位香草酸亚型1（TRPV1）及神经肽在发生机制中的作用。方法构建豚鼠食管灌流模型,进行持续（20min）热生理盐水（45℃）和盐酸（0.1mol/L,37℃）灌流刺激食管,通过伊文思蓝染色法检测其气管、支气管及细支气管的血浆渗出,观察支气管肺泡灌洗液中炎症细胞总数及分类,并观察TRPV1特异性受体拮抗剂（capsazepine,CPZ）和神经内肽酶抑制剂磷阿米酮对刺激效应的影响。结果与对照组相比（P值均〈0.05）,热刺激和酸刺激豚鼠食管均可增加其气管、支气管和细支气管血浆渗出 CPZ能抑制热和盐酸刺激引起的气管、支气管血浆渗出增多（P值均〈0.05） 磷阿米酮能明显增强盐酸和热刺激的效果（P值均〈0.05）。结论热、酸刺激豚鼠食管会引起气道血浆渗出增多,TRPV1及神经肽可能参与其发生机制。     

Platelet-activating factor-induced functional changes in the guinea pig trachea in vitro

The role of platelet-activating factor (PAF) in airway hyperresponsiveness was investigated in guinea pigs challenged in vivo for 7 days, 10 min per day, with aerosolized PAF. Tracheal smooth muscle strips set up in vitro for measuring isometric force exhibited increased sensitivity to histamine as compared to tissues from saline-challenged control animals. No significant differences in the concentration-response relationships in tissues from control and PAF-treated animals were seen for acetylcholine or KCl. The threshold concentration of histamine to elicit contraction was 10(-7) M for control and 10(-10) M for the PAF-treated tissues, and these changes persisted for several days following the last exposure to PAF. The histamine H2 receptor antagonist cimetidine, and the arachidonate cyclooxygenase inhibitor indomethacin potentiated histamine-induced contractions in tissues from both the control and the PAF-treated animals to similar magnitudes. Intracellular microelectrode studies showed similar resting membrane potentials (-51 mV) and maximum depolarizations to the three agonists in the cells from control and PAF-treated tissues. However, histamine depolarized the membrane greater in the range of 10(-7)-10(-5) M in the PAF-treated tissues than in the controls. Contractions to threshold concentrations of histamine were unaccompanied by any detectable changes in membrane potential. Histological studies showed eosinophilic infiltration of the submucosa of the trachea. The results suggest hypersensitivity to histamine in vitro following in vivo PAF challenge in guinea pig tracheal smooth muscle, with characteristic changes in the excitation-contraction coupling. These changes may be related to inflammation and cellular infiltration into the airways.     

Induction of acetylcholinesterase inhibition in the guinea pig trachea by kerosene

The effect of kerosene on in vivo and in vitro tracheal acetylcholinesterase (CHE) activity of guinea pigs has been investigated. Kerosene aerosol administered to guinea pigs during 20 min at a mean concentration of 20.4 mg/l elicited immediately, 1 h and 24 h later tracheal CHE inhibition by 30, 48.8, and 32.3% of control values (p less than 0.05). Kerosene at 1% concentration inhibited significantly, after 1 h of incubation in vitro, the CHE activity by 44.9% of control values. The tracheal CHE inhibition induced by kerosene could actually increase the acetylcholine concentration acting on smooth muscle of airways, and this partially explains the respiratory symptoms which are frequently observed after kerosene intoxication.     

Effects of prostaglandin E2 on ganglionic transmission in the guinea pig trachea.

We studied the effects of prostaglandin E2 (PGE2) on the contractile responses of in vitro guinea pig tracheal preparations with intact vagal innervation. The preparation was stimulated either through the vagal nerves (NS) or with an electrical field (EFS) and trachealis response was assessed from the pressure change inside the tracheal tube. Ganglionic blockade by hexamethonium inhibited responses to NS but did not affect EFS while both responses to NS and EFS were abolished by atropine or tetrodotoxin. This indicates that responses to both stimulation modalities were mediated by cholinergic nerves but that NS involved a ganglionic relay whereas EFS did not. Within the frequency range of 0.1-20 Hz, there was a gradual increase in the pressure generated by the trachealis muscle with increasing frequency of stimulation. The frequency-response relationship was similar for NS and EFS. Thus, the ganglion does not appear to play an important filtering or amplifying role under those conditions. PGE2 (1-50 mM) produced a concentration-dependent inhibition of NS and EFS without affecting responses to exogenous acetylcholine (ACh). This suggests that the main action of PGE2 is to reduce ACh release from post-ganglionic nerve terminals. PGE2 inhibited EFS to a larger extent than NS; we postulate a possible excitatory effect of PGE2 on neurotransmission in the airway ganglia.     

Hyperresponsiveness to tobacco dust extract in sensitized guinea pig trachea

The role of pre-existing airway inflammation in the pathogenesis of occupational airway disease is poorly understood. Previously we studied an extract of tobacco dust (TDE) and determined that it causes concentration dependent contractions of nonsensitized guinea pig trachea (GPT). In the present study animals were sensitized using Ovalbumin (OA) and subsequently challenged with an aerosol of 2.5% OA on day 21. A control group of nonsensitized GPs were divided into rings in which the epithelium was retained (EPI+) or removed (EPI-). Concentration related contractions of sensitized and nonsensitized GPTs were elicited with TDE. Sensitized GPTs demonstrated a greater contractile response to TDE than did nonsensitized GPTs. In nonsensitized animals the EPI- GPTs demonstrated a lesser response to TDE than did the EPI+. Similar findings were demonstrated in sensitized GPTs with and without epithelium. When epithelium was removed, sensitized and nonsensitized GPTs behaved similarly. Moreover, sensitized GPTs without epithelium and nonsensitized with epithelium responded similarly. These findings suggest that presensitization with an unrelated antigen enhances the response to an occupational agent and that in sensitized animals at least part of the enhanced response is mediated by the epithelial layer.     

Release of substance P from guinea pig trachea leukotriene D4.

Coordinated studies of leukotriene D4 (LTD4)-mediated contractile responses and LTD4-evoked release of the tachykinin substance P (SP) in both intact and epithelium abraded guinea pig tracheal smooth muscle preparations were performed. A partial contribution by axon reflex mechanisms to the magnitude of LTD4-induced tracheal contractions was suggested by a maximum inhibition of 21% and 28% by 5 x 10(-6) M tetrodotoxin (TTX) in abraded and intact preparations, respectively. SP-induced contractions were antagonized by the SP analog [DPro4DTrp7,9]-SP 4-11 in both types of preparation. The SP analog produced 58% and 72% inhibition of contractile responses to 10(-8) M LTD4 in abraded and intact preparations, respectively. Direct measurement of SP release by radioimmunoassay of the bathing medium showed TTX-sensitive LTD4-evoked release of SP. Inhibition by 5 x 10(-6) M TTX of LTD4-evoked SP release was 77%. The SP antagonist produced greater inhibition of LTD4-evoked contractions (58% in abraded, and 72% in intact preparations) than maximum TTX inhibition of LTD4-evoked contractions (21% in abraded, and 28% in intact). However, LTD4 (10(-8) M)-evoked SP release was at least 77% blocked by maximum doses of TTX. We therefore suggest that an additional agent, released by TTX-insensitive mechanisms, but whose contractile effects are also antagonized by [DPro4DTrp7,9]-SP 4-11, may participate in the LTD4 response.     

Inflammatory mediators involved in antigen-induced airway microvascular leakage in guinea pigs

Antigen challenge of ovalbumin (OA)-sensitized guinea pigs results in significant (p less than 0.05) increases in vascular permeability to Evans blue (EB) dye in the airways, esophagus, and bladder. Mean values +/- SEM in ng EB/mg wet weight tissue for unsensitized versus sensitized animals were: trachea, 23.6 +/- 6.6 versus 92.5 +/- 11.1; main bronchi, 31.1 +/- 12.2 versus 153.1 +/- 14.9; "central" intrapulmonary airways (ipa), 34.6 +/- 11.2 versus 101.3 +/- 6.2; and "peripheral" ipa, 26.2 +/- 6.8 versus 93.5 +/- 13.6. We investigated the involvement of several mediators of inflammation in this process. FPL 55712, a sulfidopeptide leukotriene receptor antagonist, caused significant inhibition of leakage in trachea (to 55.1 +/- 9.8) and main bronchi (91.7 +/- 15.8). Blockade of the cyclooxygenase and lipoxygenase pathways with BW 755C, but not of the cyclooxygenase pathway alone with indomethacin, also significantly reduced EB dye extravasation in trachea (55.1 +/- 18.0), main bronchi (71.7 +/- 23.0), and "central" ipa (62.7 +/- 16.4). The histamine antagonists, chlorpheniramine and cimetidine, only inhibited microvascular leakage in main bronchi (94.4 +/- 20.0). PAF-receptor blockade with the ginkgolide mixture BN 52063 had no effect. Nedocromil sodium, a mast cell stabilizer and an inhibitor of inflammatory cell activation, caused significant inhibition throughout the airways: trachea, 50.4 +/- 10.6; main bronchi, 72.0 +/- 15.3; "central" ipa 61.0 +/- 8.6; "peripheral" ipa 41.9 +/- 12.2. Thus, histamine and lipoxygenase products (in particular, leukotrienes), but not PAF, may mediate the antigen-induced increase in vascular permeability to different degrees in differing regions of the respiratory tract in guinea pigs.     

Relaxation kinetics of formoterol and salmeterol in the guinea pig trachea in vitro

The mechanisms producing long duration of action for formoterol and salmeterol are not fully understood. The aim of the current study was to examine how the concentration of long and short acting ₂-adrenoceptor agonists affects their relaxation kinetics in airway smooth muscle. Onset (time to peak relaxation) and offset of action (reassertion of reversible relaxation following repeated -adrenoceptor blockade and washout) were measured in the guinea pig trachea precontracted postjunctionally by carbachol 0.3 M in vitro. At 10–1,000% (C _1O–C _1,000) of the maximally effective concentration (C ₁₀₀: 150 nM formoterol, 10 M salbutamol, 30 M salmeterol), salbutamol had a shorter time to peak relaxation than did salmeterol. Formoterol and salmeterol had a similar time to peak relaxation at C ₁₀, but, in contrast to salmeterol, formoterol's time to peak relaxation became markedly shorter and similar to that of salbutamol as the concentration was increased up to C _1,000. Significant reversible reasserted relaxation was demonstrated for salmeterol alone at C ₁₀. At C ₃₀–C _1,000, however, salmeterol produced irreversible relaxation only, in spite of repeated -adrenoceptor blockade by sotalol 10 M followed by washout. In contrast, formoterol produced an increasing reversible reasserted relaxation at C ₃₀–C _1,000. Salbutamol produced significant, reversible reasserted relaxation at C _1,000 only. In conclusion, the concentration determines the onset and offset of action for formoterol and to a lesser extent for salbutamol, but not for salmeterol. To cause sustained action, a submaximally effective concentration is sufficient for salmeterol, whereas formoterol requires a maximally effective concentration. The rank order of concentration dependence for the relaxation kinetics is not paralleled by the rank order of lipophilicity for formoterol, salbutamol, and salmeterol. Therefore, factors other than lipophilicity may also play a role in determining the relationship between concentration and relaxation kinetics for the investigated ₂-agonists. Offprint requests to: Anders Linden, MD, PhD     

盐酸灌注豚鼠食管诱导气道血管微渗漏和神经源性炎症

目的 胃食管反流(GER)是常见的临床病症,并与多种呼吸系统疾病有关,包括慢性咳嗽和支气管哮喘等,其发病机制尚未明确.通过观察盐酸灌注食管是否可以引起气道速激肽释放和气道血浆渗出,旨在探讨GER性咳嗽的发病机制.方法 动物预先给予阿托品和心得安处理,给麻醉豚鼠食管灌注1 mol/L盐酸,分别观察气管、主支气管、细支气管的气道微血管血浆渗出的变化,气道血浆渗出测定采用伊文思蓝法;并观察应用神经内肽酶抑制剂磷阿米酮、神经肽1 受体拮抗剂FK888、SR140333对盐酸灌注豚鼠食管气管、主支气管、细支气管的气道微血管血浆渗出的影响.结果 灌注盐酸入食管可以显著增加气道血浆渗出.磷阿米酮可以显著增加盐酸灌注食管引起的气管、主支气管、细支气管血浆渗出,FK888、SR140333可以显著抑制气道血浆渗出.在双侧迷走神经切断组,神经内肽酶增强的气道血浆渗出能被明显抑制.结论 ①食管灌注盐酸通过诱导气道速激肽释放引起气道神经源性炎症;②食管和气管之间有神经通路相联系,RARs和C神经纤维参与盐酸灌注食管所致的气道血浆渗出;③该食管-支气管反射可能是GER诱发咳嗽的重要机制.     

盐酸灌注豚鼠食管诱导气道血管微渗漏和神经源性炎症

目的胃食管反流（GER）是常见的临床病症,并与多种呼吸系统疾病有关,包括慢性咳嗽和支气管哮喘等,其发病机制尚未明确。通过观察盐酸灌注食管是否可以引起气道速激肽释放和气道血浆渗出,旨在探讨GER性咳嗽的发病机制。方法。动物预先给予阿托品和心得安处理,给麻醉豚鼠食管灌注1mol／L盐酸,分别观察气管、主支气管、细支气管的气道微血管血浆渗出的变化,气道血浆渗出测定采用伊文思蓝法;并观察应用神经内肽酶抑制剂磷阿米酮、神经肽1受体拮抗剂FK888、SR140333对盐酸灌注豚鼠食管气管、主支气管、细支气管的气道微血管血浆渗出的影响。结果灌注盐酸入食管可以显著增加气道血浆渗出。磷阿米酮可以显著增加盐酸灌注食管引起的气管、主支气管、细支气管血浆渗出,FK888、SR140333可以显著抑制气道血浆渗出。在双侧迷走神经切断组,神经内肽酶增强的气道血浆渗出能被明显抑制。结论①食管灌注盐酸通过诱导气道速激肽释放引起气道神经源性炎症。②食管和气管之间有神经通路相联系,RARs和C神经纤维参与盐酸灌注食管所致的气道血浆渗出。③该食管-支气管反射可能是GER诱发咳嗽的重要机制。     

Effect of artificial surfactant on ciliary beat frequency in guinea pig trachea

This study defines the effect of artificial surfactant on ciliary beat frequency. We employed guinea pig tracheal rings and a photomultiplier which allows in vitro measurement of ciliary beat frequency. The beat frequency without surfactant decreased continuously while surfactant caused a relative increase in beat frequency. The difference between the relative beat frequency in the presence and absence of surfactant was significant. The effect of surfactant was dose dependent. Fifteen minute treatments with 2 mM hydrogen peroxide reduced the beat frequency. The reduction partially recovered in the absence of surfactant. In contrast, surfactant markedly accelerated the recovery of the beat frequency. Surfactant did not influence the effect of terbutaline (a beta 2-adrenergic agonist) on beat frequency. These results indicate that artificial surfactant can, in some circumstances, promote ciliary beat frequency. This concept is important to the clearance mechanism of the airways, and its application to some pathophysiological states such as chronic bronchitis and bronchial asthma should be considered.     

重组干扰素—γ雾化吸入抑制嗜酸细胞肺内募集作用的实验研究

观察干扰素－γ降低气道高反应性与抑制嗜酸性粒细胞在肺组织内募集的作用。     

Effects of leukotriene B4 inhalation. Airway sensitization and lung granulocyte infiltration in the guinea pig

Male Hartley guinea pigs were exposed by inhalation to leukotriene B4 (LTB4) and challenged 5 min or 4 h later with bronchoconstrictive aerosols of histamine or the divalent cationic ionophore A23187. Pulmonary gas trapping measured in excised lungs indicated the severity of post-challenge airway obstruction. Airway granulocyte infiltration was scored by an observer who was unaware of animal assignments. Treatment with LTB4 produced a marked influx of eosinophils and neutrophils into tracheal and bronchial airways. Granulocyte scores for LTB4-treated groups were 1.9 to 3.3 times higher than those for vehicle-treated groups at 5 min after exposure and 3.3 to 10.7 times higher at 4 h after exposure. Leukotriene B4 itself did not produce hyperinflation. However, histamine-induced gas trapping was increased 5 min after LTB4 exposure. Histamine responsiveness was unaffected 4 h after LTB4 treatment. In contrast, A23187-induced gas trapping was unaffected at 5 min, but diminished at 4 h after LTB4. Nonchemotactic stereoisomers of LTB4 did not produce granulocyte influx, but did produce altered airway responses similar to those seen for LTB4. We conclude that inhaled LTB4 produces airway granulocyte infiltration in the guinea pig and alterations in airway responsiveness that vary with the challenge stimulus and time after exposure. Alterations in airway responses may result from granulocyte-independent effects of LTB4 and its stereoisomers.     

Airway eosinophil accumulation on sensory neuropeptide release in a guinea pig model of distilled-water-induced bronchoconstriction.

BACKGROUND: Increased numbers of eosinophils in the airways is characteristic of asthma. However, it remains unclear whether airway eosinophils enhance or reduce the release of neuropeptides in the airways in vivo. This study was conducted to elucidate the influence of airway eosinophil accumulation on the ultrasonically nebulized distilled water (UNDW)-induced bronchoconstriction in our newly developed animal model, which is mediated by sensory neuropeptides. METHODS: Guinea pigs were transnasally treated with 100 mg/kg of platelet activating factor (PAF), or vehicle, twice a week for 3 weeks. We then conducted three experiments. In the first, UNDW was inhaled 20 min after aerosolized antigen challenge, and bronchoalveolar lavage (BAL) was performed in PAF-treated and passively sensitized animals. In the second, PAF-treated animals were exposed for 20 s to ascending doses of methacholine at intervals of 5 min In the third, passively sensitized animals were administered selective NK1 antagonist, SR 140333, selective NK2 antagonist, SR 48968, or vehicle, intravenously 5 min before UNDW-induced bronchoconstriction. RESULTS: The proportion of eosinophils in BAL fluid was significantly increased in guinea pigs treated with PAF, compared with the vehicle. The PAF treatment did not affect antigen-induced immediate asthmatic response, UNDW-induced bronchoconstriction, or bronchial responsiveness to inhaled methacholine. SR 140333, but not SR 48968, inhibited the UNDW-induced bronchoconstriction. CONCLUSION: We conclude that eosinophils accumulated in the airways, caused by repeated intranasal administration of PAF, does not affect the release of substance P induced by UNDW inhalation, or the action of released substance P in vivo.     

Identification and substance P content of vagal afferent neurons innervating the epithelium of the guinea pig trachea.

Both the nodose and jugular vagal ganglia provide sensory innervation to the airways. The purpose of this study was to localize and characterize the substance P (SP) content of vagal afferent neurons that project specifically to the tracheal epithelium. A retrograde neuronal tracer, fast blue dye or rhodamine-labeled latex microspheres, was instilled into the guinea pig trachea. After 7 d, the nodose and jugular ganglia were removed, sectioned, and prepared for immunocytochemistry. Sections of tracheal mucosa demonstrated that fast blue dye diffused throughout the airway wall, whereas the rhodamine-labeled microspheres, as expected, did not penetrate the basement membrane and were thus localized to the epithelium. When the diffusible fast blue dye was used, approximately 60% of the labeled neurons were found in the nodose ganglia and 40% in the jugular ganglia. By contrast, when the beads were used to label only epithelial nerve fibers, 97 +/- 1% of the tracheal neurons taking up the dye were derived from jugular neurons, 60 +/- 6% of which contained SP immunoreactivity. These studies demonstrate that, in contrast to the submucosa, nerve fibers innervating the epithelium of the trachea are derived nearly exclusively from neurons with cell bodies in the jugular ganglia.     

Immunohistochemical localization of histamine-stimulated increases in cyclic GMP in guinea pig lung

A significant number of asthmatic subjects are provoked by allergic reactions. The underlying pathophysiologic event is mast cell degranulation with the release and generation of the mediators of anaphylaxis. Histamine, one of the major mast cell mediators, causes 10- to 50-fold increases in guinea pig lung cyclic 3',5'-guanosine monophosphate (cyclic GMP) through H1 receptor stimulation. Employing monoclonal antibodies directed at cyclic GMP, immunocytochemical techniques were used to identify those specific cells in lung responding to histamine stimulation with increases in cyclic GMP. The most responsive cells were alveolar and parenchymal macrophages, pleural lining cells, and endothelial and epithelial cells. Little or no increases in bronchial or vascular smooth muscle cyclic GMP was noted. At the height of the reaction, a generalized increase in cyclic GMP staining of all alveolar cells was observed. These findings suggest that the lining cells of the lung including macrophages, mesothelial, endothelial, and epithelial cells may be the most responsive cells to histamine released during allergic responses. The absence of muscular staining suggests that cyclic GMP does not participate in histamine-stimulated muscle contraction.     

Airway responsiveness to intravenous and inhaled acetylcholine in the guinea pig after cigarette smoke exposure

Exposure of conscious guinea pigs to cigarette smoke results in bronchial hyperresponsiveness. To examine the mechanisms involved, we measured airway responses to increasing doses of intravenous or inhaled acetylcholine in guinea pigs exposed to cigarette smoke (n = 20) or to air (n = 20). After exposure the guinea pigs were anesthetized, paralyzed, and studied in a pressure-sensitive body plethysmograph while ventilated through a tracheostomy. Two and 6 puffs of an aerosol of increasing concentrations (0.05 to 500 micrograms/ml) of acetylcholine were delivered via the tracheostomy. Intravenous acetylcholine was delivered in boluses of 0.1 ml of increasing concentrations (0.5 to 50,000 micrograms/ml) via a catheter in an external jugular vein. Pulmonary resistance (RL), dynamic compliance (Cdyn), and heart rate (HR) were measured at baseline (after aerosolized or intravenous saline) and after each dose of acetylcholine. The peak responses to both inhaled and intravenous acetylcholine were rapid in onset (less than 15 s), short-lived (3 to 4 breaths), and were noncumulative. The baseline RL, Cdyn, and HR were not different in the smoke and air exposure groups. In the intravenous acetylcholine group, there were no differences in RL, Cdyn, and HR responses between the air and smoke exposure groups. In the inhaled acetylcholine group, the dose-response curve was shifted to the left (p less than 0.05) and reached a higher maximal response (p less than 0.01) after smoke exposure.(ABSTRACT TRUNCATED AT 250 WORDS)