Pharmacological modulation of the contractile response to toluene diisocyanate in the rat isolated urinary bladder.

1. Toluene diisocyanate produced concentration-dependent contractions of the rat isolated urinary bladder. 2. The contractions were tetrodotoxin-resistant and were abolished by previous exposure of the strips to capsaicin. 3. Indomethacin (5 microM) and ruthenium red (30 microM) inhibited toluene diisocyanate-induced contractions. Responses expressed as a percentage of the response obtained with substance P, 30 nM, were respectively 141.6 +/- 24.8% and 20.1 +/- 5.1% in control and indomethacin-treated strips (P less than 0.005); 123.0 +/- 30.2% and 14.0 +/- 6.5% in control and ruthenium red-treated strips (0.01 less than P less than 0.05). 4. These results suggest that toluene diisocyanate-induced contractions of the rat isolated bladder are the result of the release of cyclo-oxygenase products which may act by activating the capsaicin receptor.

Relevance of classification by size to topographical differences in bronchial smooth muscle response.

To investigate heterogeneity of airway smooth muscle response, we studied strips of large and small branches from third- to sixth-generation bronchi obtained from ragweed antigen-sensitized and control dogs. The response to electrical field stimulation and carbamylcholine chloride was greater in strips from larger branches of the same generation when expressed as "tissue stress" (force per unit cross-sectional area of the whole tissue), whereas no difference emerged with use of the more appropriate "smooth muscle stress" (force per unit cross-sectional area of the muscle tissue). The response to histamine was significantly higher in small branches than in large ones, and histamine sensitivity [mean effective concentration (EC50)] was 7.79 x 10(-6) [geometric standard error of the mean (GSEM) 1.20] and 1.49 x 10(-5) M (GSEM 1.14), respectively (P < 0.01). Strips from control and sensitized animals at each site and strips from different generations did not show any significant difference. When we clustered our preparations according to dimensions, the response to histamine was significantly higher in small bronchi than in large ones and histamine EC50 was 8.95 x 10(-6) (GSEM 1.17) and 1.57 x 10(-5) M (GSEM 1.18), respectively (P < 0.05). We conclude that evaluation of muscle response in different tissues requires appropriate normalization. Furthermore, classification into generations is inadequate to study bronchial responsiveness, inasmuch as major differences originate from airway size.

Different ontogeny of rate of force generation and shortening velocity in guinea pig trachealis.

Juveniles of many species, including humans, display greater airway responsiveness than do adults. This may involve changes in airway smooth muscle function. In the present work we studied force production and shortening velocity in trachealis from 1-wk-old (1 wk), 3-wk-old (3 wk), and 3-mo-old (adult) guinea pigs. Strips were electrically stimulated (60 Hz, 18 V) at their optimal length (l(o)) to obtain maximum active stress (P(o)) and rate of stress generation. Then, force-velocity curves were elicited at 2.5 s from the onset of the stimulus. By applying a recently developed modification of Hill's equation for airway smooth muscle, the maximum shortening velocity at zero load (V(o)) and the value alpha. gamma/beta, an index of internal resistance to shortening (Rsi), were calculated (alpha, beta, and gamma are the constants of the equation). P(o) increased little with maturation, whereas the rate of stress generation increased significantly (0.40 +/- 0.03, 0.45 +/- 0.03, 0. 51 +/- 0.03 P(o)/s for 1 wk, 3 wk, and adult animals). V(o) slightly increased early with maturation to decrease significantly later (1. 79 +/- 0.67, 2.45 +/- 0.92, and 0.55 +/- 0.09 l(o)/s for 1 wk, 3 wk, and adult animals), whereas the Rsi showed an opposite trend (14.98 +/- 5.19, 8.99 +/- 3.01, and 32.07 +/- 5.54 mN. mm(-2). l(o)(-1). s for 1 wk, 3 wk, and adult animals). This early increase of force generation in combination with late increase of Rsi may explain the changes of V(o) with age. An elevated V(o) may contribute to the incidence of airway hyperresponsiveness in healthy juveniles.

Response to acetylcholine and myosin content of isolated canine airways.

Contractility of tracheal smooth muscle strips and spiral strips of fourth to fifth generation bronchi was studied in organ baths. The relationship among contractility, airway smooth muscle myosin, and smooth muscle thickness was also examined. The trachea was divided into three segments, each consisting of 12-14 rings. Smooth muscle strips from each of the three regions (top, middle, and bottom of the trachea) and from fourth to fifth generation bronchi were studied. Acetylcholine (ACh) sensitivity (-log EC50) was 8.1, 7.1, 7.9, and 6.1 for the top, middle, and bottom of the trachea and the bronchi, respectively. At P = 0.01, the EC50 ACh value of the top of the trachea differed from the EC50 value of the bronchi. Maximal tension (Tmax) generated in bronchi (3.2 g) was lower (P less than 0.01) than in the top (10.4 g), middle (7.1 g), and bottom of the trachea (5.1 g). Differences between trachea and bronchi disappeared when Tmax was corrected for smooth muscle myosin content. Thickness of smooth muscle in bronchi was less (P less than 0.01) than in the three regions of trachea. Tmax was significantly correlated with airway smooth muscle thickness (r = 0.56; P less than 0.05). These results suggest that in mongrel dogs sensitivity to ACh shows a gradient from the top of the trachea to the bronchi and that Tmax is greater in the trachea than in the bronchi and is significantly correlated with thickness of smooth muscle.

Bronchial smooth muscle responses evoked by toluene diisocyanate are inhibited by ruthenium red and by indomethacin.

We have investigated the ability of ruthenium red, an inorganic dye with Ca2+ entry-blocking properties and a selective antagonist of capsaicin, and of indomethacin, a cyclooxygenase inhibitor, to inhibit bronchial smooth muscle responses evoked by toluene diisocyanate in guinea pigs. Previous exposure of isolated guinea pig bronchi to ruthenium red significantly decreased the response produced by toluene diisocyanate. Further, the response to toluene diisocyanate was significantly decreased by pretreatment with indomethacin. These findings provide evidence that toluene diisocyanate-induced contractions of guinea pig bronchi are produced indirectly by generation of a prostanoid that activates capsaicin-sensitive afferents via a ruthenium red-sensitive mechanism.

The products of the reaction between toluene diisocyanate and water contract isolated guinea pig bronchi.

We have investigated the ability of the products of the reaction between toluene diisocyanate (TDI) and water to contract bronchial smooth muscle. The experiments were performed in isolated guinea pig bronchi. TDI, both 2,4- and 2,6-toluenediamine (TDA) and mixtures of 2,4- and 2,6-TDA (ratio 80:20 and 20:80) caused concentration-dependent contraction in the isolated bronchi. The mixture of disubstituted urea and biuret also contracted the bronchi, but not in a concentration-dependent fashion. Our results provide evidence that all products of the reaction between toluene diisocyanate and water have the ability to contract isolated bronchial smooth muscle in guinea pigs. Whatever the role of toluenediamine in the adverse respiratory effects induced by exposure to isocyanates, our findings reveal the necessity of in vivo studies on the metabolism of inhaled toluene diisocyanate in humans to improve our understanding of the mechanism of action of isocyanates.

The effects of toluene diisocyanate and of capsaicin on human bronchial smooth muscle in vitro.

Toluene diisocyanate contracts guinea-pig bronchial smooth muscle through a mechanism involving capsaicin-sensitive sensory nerves. In the present study, we investigated the effects of toluene diisocyanate, capsaicin and tachykinins on isolated human bronchi. In 44 rings, toluene diisocyanate (0.3 mM) produced a relaxation which averaged 16.9 +/- 1.1%, in ten rings it produced a shortening that was 15.1 +/- 3.3% and in ten preparations it gave no response. A second administration of toluene diisocyanate (0.3 mM) always produced a relaxation (n = 13, 18.1 +/- 3.9%). Capsaicin (0.03 mM) produced shortening in 15 (35 +/- 6.6%) and relaxation in 11 preparations (41 +/- 6.8%), whereas a second administration caused shortening in nine (25.1 +/- 6.1%) and relaxation in 16 rings (36.4 +/- 4.9%). When toluene diisocyanate was given after two consecutive capsaicin administrations, we observed shortening in two rings (10.0 +/- 3.6%), relaxation in ten rings (15.9 +/- 3.6%), and no response in four preparations. To test the role of NK1 and NK2 receptors in these conflicting responses, we performed concentration-response curves to different tachykinins. Substance P, neurokinin A and neurokinin A-(4-10), a specific NK2 receptor agonist, gave a concentration-dependent shortening, with neurokinin A being the most effective and neurokinin A-(4-10) the least. The specific NK1 receptor agonist, [Sar9, Met(O2)11]substance P, produced both shortening and relaxation. We conclude that toluene diisocyanate and capsaicin may produce both shortening and relaxation in isolated human bronchi through NK1 receptors.

Bronchopulmonary inflammation and airway smooth muscle hyperresponsiveness induced by nitrogen dioxide in guinea pigs.

We investigated whether acute exposure to nitrogen dioxide (NO2) causes major inflammatory responses (inflammatory cell recruitment, oedema and smooth muscle hyperresponsiveness) in guinea pig airways. Anaesthetised guinea pigs were exposed to 18 ppm NO2 or air for 4 h through a tracheal cannula. Bronchoalveolar lavage was performed and airway microvascular permeability and in vitro bronchial smooth muscle responsiveness were measured. Exposure to NO2 induced a significant increase in eosinophils and neutrophils in bronchoalveolar lavage fluid, microvascular leakage in the trachea and main bronchi (but not in peripheral airways), and a significant in vitro hyperresponsiveness to acetylcholine, electrical field stimulation, and neurokinin A, but not to histamine. Thus, this study shows that in vivo exposure to high concentrations of NO2 induces major inflammatory responses in guinea pig airways that mimic acute bronchitis induced by exposure to irritant gases in man.

Evidence that toluene diisocyanate activates the efferent function of capsaicin-sensitive primary afferents.

Isocyanates are an important cause of occupational asthma. The mechanism of isocyanate-induced asthma is still unknown. To determine whether toluene diisocyanate stimulates the 'efferent' function of peripheral endings of capsaicin-sensitive sensory nerves, we investigated the effect of toluene diisocyanate in the rat isolated urinary bladder, a preparation in which the action of capsaicin has been well characterized. Toluene diisocyanate (0.03-3 mM) produced a concentration-dependent contraction of the bladder strips. Its maximal effect was about 50% of the response to capsaicin (1 microM). Previous exposure of the strips to capsaicin followed by washing out produced complete unresponsiveness, both to the first exposure to toluene diisocyanate and to a second exposure of capsaicin. Further, the response to both toluene diisocyanate and capsaicin was completely prevented by extrinsic bladder denervation, achieved by bilateral removal of pelvic ganglia (72 h before). Repeated exposure of the rat bladder to toluene diisocyanate reduced the capsaicin-evoked release of calcitonin gene-related peptide-like immunoreactivity (CGRP-LI), taken as biochemical marker of activation of these sensory nerves. These experiments provide the first evidence that toluene diisocyanate activates directly or indirectly the efferent function of capsaicin-sensitive primary sensory nerves.

The effect of compound 48/80 on contractions induced by toluene diisocyanate in isolated guinea-pig bronchus.

We have investigated the ability of compound 48/80 and of histamine H1 and H2 receptor antagonists to inhibit toluene diisocyanate-induced contractions in isolated guinea-pig bronchi. Compound 48/80 (100 micrograms/ml) significantly inhibited toluene diisocyanate-induced contractions. By contrast, the two histamine H1 and H2 receptor antagonists, chlorpheniramine (10 microM) and cimetidine, (10 microM) did not affect toluene diisocyanate-induced contractions, but significantly inhibited contractions induced by exogenously applied histamine (100 microM) and by 48/80. We investigated which mechanisms 48/80 used to inhibit toluene diisocyanate-induced contractions, paying particular attention to the possible involvement of capsaicin-sensitive primary afferents. In vitro capsaicin desensitization (10 microM for 30 min followed by washing) significantly reduced compound 48/80-induced contractions. A capsaicin-resistant component of contraction was also evident. Ruthenium red (3 microM), an inorganic dye which acts as a selective functional antagonist of capsaicin, did not affect 48/80-induced contraction. MEN 10,207 (Tyr5,D-Trp6,8,9,Arg10)-neurokinin A (4-10) (3 microM) a selective antagonist of NK2-tachykinin receptors significantly reduced 48/80-induced contractions. These results show that compound 48/80 inhibits toluene diisocyanate-induced contractions in isolated guinea-pig bronchi. It is likely that two mechanisms are involved in the inhibition: (1) the release of mediators other than histamine by mast cells, (2) an effect of 48/80 on sensory nerves.

Maturational changes in airway smooth muscle shortening and relaxation. Implications for asthma.

Greater airway responsiveness in healthy juveniles is considered a factor in the higher asthma prevalence at a young age compared with adults. Several studies on the contractile response of airway smooth muscle (ASM) from birth to adulthood have addressed the hypothesis that a maturation of ASM plays a role in juvenile airway hyperresponsiveness. Maturation of distinct ASM properties, i.e. force generation, shortening, and relaxation, has been reported, although the majority of the studies have focused on maturation of maximum force and/or sensitivity to contractile agonists. However, in most animal species maturation of the ability to generate force does not correlate with maturation of airway responsiveness. Ontogenesis of ASM shortening has been less extensively studied and the existing reports emphasize an increase during maturation of tissue passive forces opposing shortening. ASM spontaneous relaxation has been very minimally investigated. We have recently demonstrated that the ability of ASM to spontaneously relax during stimulation is sharply reduced in juvenile airway tissue. It remains to be determined the role of these ASM properties in the onset of childhood asthma and whether specific alterations are induced by the occurrence of obstructive airway diseases in young individuals.

In-vitro exposure of guinea pig main bronchi to 2.5 ppm of nitrogen dioxide does not alter airway smooth muscle response.

In order to investigate whether the oxidant airborne pollutant nitrogen dioxide (NO2) affects airway smooth muscle responsiveness, the contractile response of guinea pig main bronchi after in vitro exposure to 2.5 ppm of nitrogen dioxide was studied. Main bronchi were cannulated and exposed for 2 or 4 h to a constant flow of either NO2 or air. After exposure, bronchial rings were obtained and placed in a 37 degrees C jacketed organ bath filled with Krebs-Henseleit solution. Concentration-response curves were performed for acetylcholine (10(-9)-10(-3) M), substance P (10(-9)-10(-4) M), and neurokinin A (10(-10)-10(-5) M), and voltage-response curves (12-28 V) were performed for electrical field stimulation. There was no significant difference in either the smooth muscle maximal contractile response, or sensitivity between the bronchi exposed to NO2 and those exposed to air. We conclude that in vitro exposure to 2.5 ppm of NO2 does not alter airway smooth muscle responsiveness in guinea pigs.

Serum-mediated relaxant response to toluene diisocyanate (TDI) in isolated guinea-pig bronchi.

The present study was designed to evaluate whether pre-incubation with serum, obtained from both control and toluene diisocyanate (TDI)-immunized guinea-pigs, modified the contractile response to TDI in isolated guinea-pig bronchial rings. Guinea-pigs were anaesthetized and the main bronchi dissected in two rings. Bronchial rings were incubated with normal or immune serum (100 microl ml(-1) for 2 h) and dose-response curves to TDI (0.03-1000 microM) were studied isometrically. Before serum incubation, in eight bronchial rings, epithelium was removed by rubbing the luminal surface gently with a gauze. In control rings, TDI produced a concentration-dependent contraction, whereas in rings pre-incubated with either normal or TDI-immune serum, it produced a concentration-dependent relaxation. Relaxation was 101.4 (SEM 17.4)% and 94.9 (SEM 21)% of the relaxation induced by isoproterenol (1 mM) respectively with normal and TDI-immune serum. Similarly to the pre-incubation with serum, pre-incubation with albumin produced a concentration-dependent relaxation to TDI. Serum-induced relaxant response to TDI was not affected by capsaicin desensitization, it was only partially inhibited by an NK1-tachykinin antagonist, whereas it was blocked by indomethacin. In bronchial rings without epithelium, pre-incubated with serum, TDI caused contraction at highest doses, while it still induced relaxation at the lowest doses. This study shows that one or more components of the serum modify the contractile response to TDI in isolated guinea-pig bronchi. In bronchial rings without epithelium serum was able to inhibit the contration induced by low doses of TDI.

Airway smooth muscle dynamics: a common pathway of airway obstruction in asthma.

Excessive airway obstruction is the cause of symptoms and abnormal lung function in asthma. As airway smooth muscle (ASM) is the effecter controlling airway calibre, it is suspected that dysfunction of ASM contributes to the pathophysiology of asthma. However, the precise role of ASM in the series of events leading to asthmatic symptoms is not clear. It is not certain whether, in asthma, there is a change in the intrinsic properties of ASM, a change in the structure and mechanical properties of the noncontractile components of the airway wall, or a change in the interdependence of the airway wall with the surrounding lung parenchyma. All these potential changes could result from acute or chronic airway inflammation and associated tissue repair and remodelling. Anti-inflammatory therapy, however, does not "cure" asthma, and airway hyperresponsiveness can persist in asthmatics, even in the absence of airway inflammation. This is perhaps because the therapy does not directly address a fundamental abnormality of asthma, that of exaggerated airway narrowing due to excessive shortening of ASM. In the present study, a central role for airway smooth muscle in the pathogenesis of airway hyperresponsiveness in asthma is explored.

The Schultz-Dale response of sensitized canine bronchial smooth muscle.

We wished to determine why in vitro agonist dose-response curves show reduced leftward shift (1/2-1 log dose units) in sensitized canine airway smooth muscle compared to curves elicited in vivo (2-3 log dose unit). The Schultz-Dale response was studied in sensitized dog tracheal (TSM) and bronchial (BSM) smooth muscle. Sensitized TSM challenged with specific antigen showed greater mechanical response, but only on exposure to 300 micrograms/ml; BSM responded to concentrations of ragweed as low as 0.001 microgram/ml. This result resolved the problem cited at the outset. Control TSM and BSM showed no response. The response in BSM is mediated through histamine release, and to a smaller extent by acetylcholine. With challenge release of histamine and acetylcholine increased significantly in sensitized airway smooth muscle. Integrated contractile responses obtained with high and low concentrations of antigen showed a dose-response relationship. Increased sensitivity of BSM to antigen compared to TSM indicates the former is the preparation of choice for study of allergic bronchoconstriction.

Effect of oxidant air pollutants on the respiratory system: insights from experimental animal research.

In the present paper, we have reviewed experimental animal studies on the effects of the two most important oxidant airborne pollutants, nitrogen dioxide and ozone, on the respiratory system. The toxic effects depend on concentration and length of exposure, and are generally similar for both oxidants, with ozone operative at lower concentrations. High doses of both oxidants cause death due to lung oedema. Exposure to sublethal levels causes functional alterations such as airflow limitation and airway hyperresponsiveness to bronchoconstrictor stimuli. These effects, which are generally reversible, are associated with epithelial injury, oedema and airway and parenchymal infiltration by inflammatory cells. Loss of cilia of airway epithelium and necrosis of type I alveolar epithelial cells are the most prominent consequences at the epithelial level. Inflammation is characterized by early neutrophilic infiltration, followed by an increased number of mononuclear cells, predominantly alveolar macrophages. After long-term exposure, whilst nitrogen dioxide causes predominantly emphysema, ozone produces mainly pulmonary fibrosis. Biochemical effects include lipid peroxidation, increased antioxidant metabolism, and alteration of enzyme activity. Nitrogen dioxide and ozone may also alter the immunological response and reduce the defence against infections, increasing the susceptibility of exposed animals to infections.

Reducing agents inhibit the contractile response of isolated guinea-pig main bronchi.

BACKGROUND: Oxidants are involved in many respiratory disorders, including asthma and chronic obstructive pulmonary diseases. Reduced glutathione (GSH), one of the most important antioxidant compounds against oxidant free radicals, is particularly abundant in the respiratory epithelial lining fluid, where its concentration is increased in inflammatory disorders. OBJECTIVE: We hypothesized that reducing agents may have a direct effect on airway smooth muscle. Therefore, we studied the effects of GSH on airway smooth muscle contractility in guinea-pig main bronchi. In parallel, we evaluated superoxide anion generation associated with in vitro bronchial smooth muscle contraction. METHODS: Guinea-pig main bronchi were mounted in organ baths filled with Krebs-Henseleit solution. Concentration-response curves to acetylcholine (Ach) (10(-9)-10(-3) M), carbachol (10(-9)-10(-4) M), or histamine (10(-9)-10(-3) M) were performed in the presence or absence of either reduced or oxidized glutathione (GSSG) (10(-5)-10(-3) M). We also evaluated the effects of GSH and GSSG on allergen-induced contraction in main bronchi obtained from ovalbumin-sensitized guinea-pig. Superoxide dismutase (SOD)-inhibited cytochrome c reduction kinetics was performed to evaluate superoxide anion (O2-) production during Ach-induced contraction. RESULTS: Reduced but not oxidized glutathione significantly decreased smooth muscle contraction induced by Ach, carbachol, and histamine. Similarly, only the reduced form of glutathione attenuated the bronchoconstriction induced by allergen exposure in bronchi from sensitized animals. Finally, SOD-inhibited cytochrome c reduction kinetics demonstrated increased O2- production following bronchial smooth muscle contraction. This production was not affected by epithelium removal. CONCLUSION: Our findings show that GSH decreases bronchial smooth muscle contraction to different stimuli and that oxidant free radicals are produced during bronchial smooth muscle contraction. We suggest that oxidants are involved in the mechanisms of bronchoconstriction and that reducing agents could be a possible therapeutic option for airway obstruction sustained by bronchospasm.

Dehydroepiandrosterone and analogs inhibit DNA binding of AP-1 and airway smooth muscle proliferation.

The adrenal steroid dehydroepiandrosterone (DHEA) and its analogs reduce growth of immortalized and malignant cell lines. We therefore explored their effects on the growth of airway smooth muscle, whose hyperplasia may lead to fixed airways obstruction and enhanced airways hyperresponsiveness in severe chronic asthma. DHEA and its potent analog 16 alpha-bromoepiandrosterone dramatically reduced proliferation in primary cultures of rat tracheal smooth muscle stimulated with fetal bovine serum or platelet-derived growth factor. Growth inhibition was dose-dependent and could not be attributed to interference with glucose-6-phosphate dehydrogenase activity or cholesterol metabolism, as reported for immortalized or malignant cell lines, respectively. Expression of the early response gene c-fos remained intact, but DHEA and 16 alpha-bromoepiandrosterone decreased DNA binding of the transcription factor activator protein-1, a later response important for expression of genes that mediate DNA synthesis and cell cycle progression. These results suggest that the nonglucocorticoid steroid DHEA and its analogs may impair activation of secondary growth response genes in a fashion analogous to that reported for glucocorticoids and that they may prove useful for treatment of asthmatic airway remodeling in the human.

In vivo exposure to nitrogen dioxide (NO2) induces a decrease in calcitonin gene-related peptide (CGRP) and tachykinin immunoreactivity in guinea-pig peripheral airways.

The mammalian respiratory tract is densely innervated by sensory and autonomic fibres. Subsets of the nerves contain bioactive regulatory peptides, such as substance P, calcitonin gene-related peptide (CGRP), and neurokinins. The sensory nervous system responds to inhaled irritants, resulting in a release of neuropeptides and, thus, a decrease in the peptide immunoreactivity of the fibres. We examined the effects of inhaled nitrogen dioxide (NO2), a well-known indoor and outdoor air pollutant, on pulmonary sensory neuropeptides. Guinea-pigs were exposed for 4 h to 18 parts per million (ppm) NO2 or to air (n = 5 each). At the end of the exposure, they were killed with urethane and their lungs were fixed in 1% paraformaldehyde in phosphate-buffered saline. Cryostat sections were stained with antisera to an anatomical nerve marker, protein gene product (PGP) 9.5, and to CGRP and tachykinins, utilizing the avidin-biotinylated peroxidase method. In the noncartilaginous airways (diameter < 250 microns) of NO2-exposed animals, less tachykinin- and CGRP-immunoreactive nerve fibres were found compared with controls. No change was seen in the total nerve fibre distribution (PGP 9.5). It is concluded that the peptidergic nerves of guinea-pig peripheral airways are a sensitive indicator of exposure to nitrogen dioxide.

Isotonic smooth muscle response in human bronchi exposed in vitro to nitrogen dioxide.

Exposure to nitrogen dioxide (NO2), a common oxidant airborne pollutant, has been shown to cause reversible effects on lung function and airway responsiveness, in addition to airways inflammation. However, there have been conflicting reports concerning NO2-induced airway hyperresponsiveness. In the present study, we investigated the isotonic smooth muscle response in isolated human bronchi previously exposed in vitro to NO2. Bronchial segments were obtained from 12 patients who had undergone thoracotomy for lung cancer. Bronchial segments from each patient were exposed to air and to 2.5 parts per million (ppm) NO2 for 4 h. The contractile response of bronchial rings to acetylcholine, neurokinin A (NKA), and substance P was then studied under isotonic conditions. The response to NKA was also studied in rings, with or without epithelium, exposed either to air or 7 ppm NO2. No NO2-induced alteration of the bronchial smooth muscle isotonic response was found under any of the experimental conditions. We conclude that in vitro exposure to up to 7 ppm nitrogen dioxide does not cause alterations of the human bronchial smooth muscle shortening capacity.