Pulmonary Inflation Pressure Research Articles

Effect of sustained inflation with different degrees of negative pressure for sputum aspiration in patients with acute respiratory distress syndrome on lung recruitment

To investigate the effect of different degrees of pressure of sustained inflation (SI) in patients with acute respiratory distress syndrome (ARDS) after lung recruitment as the result of different negative pressure for sputum aspiration. A prospective single-blind randomized controlled trial was conducted. The factorial analysis of variance was adopted. 150 patients with ARDS admitted to the emergency intensive care unit (ICU) of Chongqing Three Gorges Central Hospital from January 2012 to December 2014 were enrolled, and they were randomly divided into S1, S2, S3 group, with 50 patients in each group, suction pressure varying from 150, 175, to 200 mmHg (1 mmHg=0.133 kPa) was respectively used in each group. Then the patients of each group were randomly subdivided into five subgroups of P0, P1, P2, P3, P4, with 10 patients in each group, and 0, 30, 35, 40, and 45 cmH2O (1 cmH2O=0.098 kPa) were used for control pulmonary inflation pressure, respectively. The respiratory mechanics and the hemodynamic parameters were recorded, and they were compared before and after the sputum aspiration as well as lung recruitment with sustained inflation. The lung recruitment volume (mL: 87.56±28.47 vs. 109.38±34.63, t=3.573, P=0.001) and lung static compliance [Cst (mL/cmH2O): 27.69±13.25 vs. 35.87±17.47, t=2.814, P=0.004] after sputum aspiration in the 150 patients were significantly lower than those before the sputum aspiration, and peak airway pressure [PIP (cmH2O): 24.16±8.28 vs. 18.63±6.67, t=2.957, P=0.005], airway plateau pressure [Pplat (cmH2O): 21.28±9.14 vs. 17.47±7.26, t=2.089, P=0.032], and mean airway pressure [Pm (cmH2O): 13.26±4.65 vs. 10.41±3.54, t=3.271, P=0.001] were significantly higher than those before the treatment. There were no significant differences in the lung recruitment volume, Cst, PIP, Pplat and Pm between groups with different negative pressure for sputum aspiration (F value was 0.809, 0.986, 1.121, 0.910, 1.043, and P value was 0.452, 0.381, 0.335, 0.410, 0.361), but statistical significance was found among different groups of different lung recruitment pressures (F value was 3.581, 5.028, 3.064, 3.036, 4.050, and P value was 0.013, 0.002, 0.026, 0.027, 0.007). There was no interaction between the two factors. After pairwise comparison, under the same negative pressure for sputum aspiration, lung recruitment volume and Cst in different lung recruitment pressures subgroups (P1, P2, P3, P4) were significantly higher than those of P0 subgroup, and PIP, Pplat, and Pm were significantly lower than those of P0 subgroup. There was no significant difference among P1, P2, P3 and P4 groups. There were no significant differences in mean arterial pressure (MAP) and pulmonary arterial pressure (PAP) among different groups with negative pressures for sputum aspiration and different lung recruitment pressures (negative pressure for sputum aspiration: F=0.586, P=0.561, F=1.373, P=0.264; lung recruitment pressure: F=1.313, P=0.280, F=1.621, P=0.186), there was no interaction between the two factors (F=0.936, P=0.497, F=1.391, P=0.227). The difference of heart rate (HR) in different negative pressure for sputum aspiration groups was not significant (F=1.144, P=0.328), and there were significant differences in different lung recruitment pressure groups (F=3.297, P=0.019), there was no interaction between the two factors (F=1.277, P=0.280). After pairwise comparison, under the same negative pressure for sputum aspiration, HR in P3 and P4 subgroups was significantly higher than that in P0, P1, and P2 subgroups (all P<0.05). 30 cmH2O and 35 cmH2O were the suitable pressure for SI in ARDS patients, and they were not affected by different negative pressure for sputum aspiration.

Low Voltage Vagal Nerve Stimulation Reduces Bronchoconstriction in Guinea Pigs Through Catecholamine Release

Electrical stimulation of the vagus nerve at relatively high voltages (e.g., >10 V) can induce bronchoconstriction. However, low voltage (≤2 V) vagus nerve stimulation (VNS) can attenuate histamine-invoked bronchoconstriction. Here, we identify the mechanism for this inhibition. In urethanea-nesthetized guinea pigs, bipolar electrodes were attached to both vagus nerves and changes in pulmonary inflation pressure were recorded in response to i.v. histamine and during VNS. The attenuation of the histamine response by low-voltage VNS was then examined in the presence of pharmacologic inhibitors or nerve ligation. Low-voltage VNS attenuated histamine-induced bronchoconstriction (4.4 ± 0.3 vs. 3.2 ± 0.2 cm H(2) O, p < 0.01) and remained effective following administration of a nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester, and after sympathetic nerve depletion with guanethidine, but not after the β-adrenoceptor antagonist propranolol. Nerve ligation caudal to the electrodes did not block the inhibition but cephalic nerve ligation did. Low-voltage VNS increased circulating epinephrine and norepinephrine without but not with cephalic nerve ligation. These results indicate that low-voltage VNS attenuates histamine-induced bronchoconstriction via activation of afferent nerves, resulting in a systemic increase in catecholamines likely arising from the adrenal medulla.

Role of TNF‐α in virus‐induced airway hyperresponsiveness and neuronal M2 muscarinic receptor dysfunction

Infections with respiratory viruses induce exacerbations of asthma, increase acetylcholine release and potentiate vagally mediated bronchoconstriction by blocking inhibitory M₂ muscarinic receptors on parasympathetic neurons. Here we test whether virus-induced M₂ receptor dysfunction and airway hyperresponsiveness are tumour necrosis factor-alpha (TNF-α) dependent. Guinea pigs were pretreated with etanercept or phosphate-buffered saline 24 h before intranasal infection with parainfluenza. Four days later, pulmonary inflation pressure, heart rate and blood pressure were measured. M₂ receptor function was assessed by the potentiation by gallamine (an M₂ receptor antagonist) of bronchoconstriction caused by electrical stimulation of the vagus nerves and measured as increased pulmonary inflation pressure. Human airway epithelial cells were infected with influenza and TNF-α concentration in supernatant was measured before supernatant was applied to human neuroblastoma cells. M₂ receptor expression in these neuroblastoma cells was measured by qRT-PCR. Influenza-infected animals were hyperresponsive to vagal stimulation but not to intravenous ACh. Gallamine did not potentiate vagally induced bronchoconstriction in virus-infected animals, indicating M₂ receptor dysfunction. Etanercept prevented virus-induced airway hyperresponsiveness and M₂ receptor dysfunction, without changing lung viral titres. Etanercept caused a non-significant decrease in total cells, macrophages and neutrophils in bronchoalveolar lavage. Influenza infection significantly increased TNF-α release from isolated epithelial cells, sufficient to decrease M₂ receptors in neuroblastoma cells. This ability of supernatants from infected epithelial cells to inhibit M₂ receptor expression was blocked by etanercept. TNF-α is a key mediator of virus-induced M₂ muscarinic receptor dysfunction and airway hyperresponsiveness.

Airway reactivity to inhaled spasmogens 18-24 h after antigen-challenge in sensitized anaesthetized guinea-pigs.

The anaesthetized allergic guinea-pig was used to assess changes in airway reactivity to four different inhaled spasmogens: methacholine, 5-hydroxytryptamine (5-HT), histamine and the thromboxane A2 mimetic, 9,11-dideoxy-9 alpha,11 alpha-methano-epoxy-PGF2 alpha (U-46619). Reactivity was determined 18 to 24 h after challenge of ovalbumin-sensitized guinea-pigs with inhaled ovalbumin. This time coincides with the appearance of a late-phase bronchoconstriction in these animals. Sensitivity to the spasmogen was assessed from the concentration-response curve for the increase in pulmonary inflation pressure (PIP) in ovalbumin- and saline-challenged sensitized animals. When methacholine, 5-HT or histamine were the spasmogens there was no hyper-reactivity. The geometric mean EC50 values (i.e. the concentrations inducing half the maximum effect) obtained from the dose-response curves for methacholine (73 (42-129) and 94 (66-134) micrograms mL-1), 5-HT (1.5 (0.81-3.03) and 1.1 (0.51-2.24 micrograms mL-1) and histamine (39 (21-75) and 72 (32-162) micrograms mL-1) did not differ significantly (P > 0.05) between saline- and ovalbumin-challenged animals, respectively. However, when U-46619 was the spasmogen, ovalbumin-induced airway hyper-reactivity was observed as a leftwards shift of the concentration-response curve and the EC50 value for ovalbumin-challenged animals (8.1 (5.1-13) ng mL-1) was significantly (P < 0.05) less than the value for control animals (39 (21-75) ng mL-1). Our findings suggest that airway hyper-reactivity is not 'non-specific', but instead depends on the chosen spasmogen. The absence of hyper-reactivity with certain spasmogens was not a result of poor delivery, because all spasmogens caused a bronchoconstriction by the inhaled route. It was also not associated with the model because ozone has been shown to induce hyper-reactivity to inhaled methacholine and 5-HT. Because airway hyper-reactivity to both inhaled histamine and agonists at muscarinic receptors is regularly seen in man, the anaesthetized guinea-pig might not be the ideal model for assessing airway hyper-reactivity after antigen challenge and its modification by anti-asthma drugs.

Gantacurium and CW002 Do Not Potentiate Muscarinic Receptor-mediated Airway Smooth Muscle Constriction in Guinea Pigs

Neuromuscular blocking agents are an integral component of general anesthesia. In addition to their intended pharmacologic target on skeletal muscle nicotinic receptors, undesirable airway effects (i.e., bronchoconstriction) can result from neuromuscular blocking agents' affinity for airway muscarinic receptors. We questioned whether two new members of a bisquaternary nondepolarizing muscle relaxant family, gantacurium and CW002, demonstrated detrimental effects of airway muscarinic receptors using an in vivo model in guinea pig airways. Urethane-anesthetized male guinea pigs were ventilated through a tracheostomy with continuous digital recordings of pulmonary inflation pressure and heart rate. The dose for 95% twitch suppression for gantacurium, CW002, cisatracurium, and rapacuronium was defined in the guinea pig. Transient and reproducible changes in pulmonary inflation pressure and heart rate were recorded after vagal nerve stimulation or intravenous injection of acetylcholine before and after pretreatment with cumulatively increasing concentrations of gantacurium, CW002, cisatracurium or a single concentration of rapacuronium. The doses for 95% twitch suppression for gantacurium, CW002, cisatracurium, and rapacuronium were 0.064 +/- 0.006, 0.012 +/- 0.0006, 0.10 +/- 0.003, and 0.31 +/- 0.05 mg/kg, respectively. Gantacurium, CW002, and cisatracurium were without effects on baseline pulmonary inflation pressures and were devoid of significant interactions with M2 and M3 muscarinic receptors in vivo. These findings suggest that gantacurium and CW002 are devoid of significant effects at airway muscarinic receptors particularly M3 receptors on bronchial smooth musculature at doses several fold higher than those required for functional muscle paralysis.

Inhibition of Histamine-Induced Bronchoconstriction in Guinea Pig and Swine by Pulsed Electrical Vagus Nerve Stimulation

Objective. Smooth muscle help regulate the diameter of the airways and their constriction can contribute to the pathology of acute asthma attacks. This study sought to determine if applying a specific electrical signal to the vagus nerve (VN) could minimize histamine-induced bronchoconstriction. Methods. Sixteen guinea pigs and three swine were anesthetized and had bipolar electrodes positioned on the cervical VNs. After the animals stabilized, i.v. histamine was titrated to elicit a moderate 2-4 cm H(2) O increase in pulmonary inflation pressure (Ppi). Histamine was then dosed with or without concurrent low voltage VN stimulation. Results. The peak change in Ppi following a histamine challenge was reduced in the guinea pig by VN stimulation (3.4 ± 0.4 vs. 2.1 ± 0.2 cm H(2) O, p < 0.001). The results were confirmed in a limited study in swine and indicate VN treatment is applicable to larger animals. Conclusion. This study suggests that VN stimulation can reduce bronchoconstriction and may prove useful as a rescue therapy in the treatment of acute asthma.

The GABAA agonist muscimol attenuates induced airway constriction in guinea pigs in vivo

GABA(A) channels are ubiquitously expressed on neuronal cells and act via an inward chloride current to hyperpolarize the cell membrane of mature neurons. Expression and function of GABA(A) channels on airway smooth muscle cells has been demonstrated in vitro. Airway smooth muscle cell membrane hyperpolarization contributes to relaxation. We hypothesized that muscimol, a selective GABA(A) agonist, could act on endogenous GABA(A) channels expressed on airway smooth muscle to attenuate induced increases in airway pressures in anesthetized guinea pigs in vivo. In an effort to localize muscimol's effect to GABA(A) channels expressed on airway smooth muscle, we pretreated guinea pigs with a selective GABA(A) antagonist (gabazine) or eliminated lung neural control from central parasympathetic, sympathetic, and nonadrenergic, noncholinergic (NANC) nerves before muscimol treatment. Pretreatment with intravenous muscimol alone attenuated intravenous histamine-, intravenous acetylcholine-, or vagal nerve-stimulated increases in peak pulmonary inflation pressure. Pretreatment with the GABA(A) antagonist gabazine blocked muscimol's effect. After the elimination of neural input to airway tone by central parasympathetic nerves, peripheral sympathetic nerves, and NANC nerves, intravenous muscimol retained its ability to block intravenous acetylcholine-induced increases in peak pulmonary inflation pressures. These findings demonstrate that the GABA(A) agonist muscimol acting specifically via GABA(A) channel activation attenuates airway constriction independently of neural contributions. These findings suggest that therapeutics directed at the airway smooth muscle GABA(A) channel may be a novel therapy for airway constriction following airway irritation and possibly more broadly in diseases such as asthma and chronic obstructive pulmonary disease.

Differential responses to various classes of drugs in a model of allergic rhinitis in guinea pigs

Different drugs from various pharmacological classes were compared for their ability to protect against the nasal effects of acute allergen challenge in a guinea pig model. In the model, sneezing and nose rubbing were recorded after an initial allergen challenge in guinea pigs previously sensitized to egg albumin. Four days later the same guinea pigs were re-challenged a second time when anesthetised. In these anaesthetized animals, nasal airway pressure, pulmonary inflation pressure and cellular infiltration into nasal lavage fluid were measured. The drug tested were autacoid antagonists (mepyramine—3 mg/kg, cetirizine—3 mg/kg and montelukast—10 mg/kg), L-NAME (10 or 20 mg/kg), heparin (20 mg/kg) and dexamethasone (20 mg/kg) given either intraperitoneally or intravenously; all were given shortly before challenge. Sneezing induced by allergen challenge was statistically significantly reduced by mepyramine, cetirizine and dexamethasone whereas only cetirizine reduced nose rubbing. Changes in nasal airway pressure due to allergen exposure were reduced by cetirizine, montelukast, L-NAME, and heparin, but not by mepyramine, nor dexamethasone. In the presence of L-NAME, nasal airway pressure actually changed in the opposite direction. Cellular infiltration, as assessed by cytometry in nasal lavage fluid 60 min after acute allergen challenge, was reduced by montelukast and heparin but not by antihistamines, L-NAME nor dexamethasone. This pattern of effects of the drugs, given by doses and routes previously described in the literature as being effective was not completely consistent with expected responses. The lack of effect of dexamethasone probably reflects the fact that it was given acutely whereas in the clinic chronic administration is used. The two antihistamines were not identical in their actions, presumably reflecting the fact that cetirizine has therapeutic actions not entirely confined to blockade of H1 receptors. Montelukast has not been reported to have major effects on sneezing and itching in the clinic but reduces nasal obstruction (lower nasal airway pressure or nasal patency). Montelukast's effects on cellular infiltration indicate the possible involvement of leukotrienes. Heparin has actions on inflammatory cell infiltration. This could explain its profile of reducing both cellular infiltration, and increased nasal airway pressure.

In vivo adenosine A2B receptor desensitization in guinea-pig airway smooth muscle: Implications for asthma

This study was aimed at characterizing the role of adenosine receptor subtypes in the contractility modulation of guinea-pig airway smooth muscle in normal and pathological settings. In vitro and in vivo experiments were performed by testing selective agonists and antagonists on isolated tracheal smooth muscle preparations and pulmonary inflation pressure, respectively, under normal conditions or following ovalbumin-induced allergic sensitization. In normal and sensitized animals, the adenosine A(2A)/A(2B) receptor agonist, NECA, evoked relaxing responses of isolated tracheal preparations precontracted with histamine, and such an effect was reversed by the adenosine A(2B) antagonist, MRS 1706, in the presence or in the absence of epithelium. The expression of mRNA coding for adenosine A(2B) receptors was demonstrated in tracheal specimens. In vitro desensitization with 100 microM NECA markedly reduced the relaxing effect of the agonist. In vivo NECA or adenosine administration to normal animals inhibited histamine-mediated bronchoconstriction, while these inhibitory effects no longer occurred in sensitized guinea-pigs. Adenosine plasma levels were significantly higher in sensitized than normal animals. In conclusion, our data demonstrate that: (i) adenosine A(2B) receptors are responsible for the relaxing effects of adenosine on guinea-pig airways; (ii) these receptors can undergo rapid adaptive changes that may affect airway smooth muscle responsiveness to adenosine; (iii) ovalbumin-induced sensitization promotes a reversible inactivation of adenosine A(2B) receptors which can be ascribed to homologous desensitization. These findings can be relevant to better understand adenosine functions in airways as well as mechanisms of action of asthma therapies targeting the adenosine system.

Adrenal Influences on the Inhibitory Effects of Procaterol, a Selective Beta-Two-Adrenoceptor Agonist, on Antigen-Induced Airway Microvascular Leakage and Bronchoconstriction in Guinea Pigs

While the guinea pig has been the preferred choice for use as a model of allergic bronchial asthma in the evaluation of anti-asthmatic drugs, it has been shown that antigen-induced bronchoconstriction in guinea pigs is attenuated by epinephrine released from the adrenal gland. In order to investigate the possible influence of the adrenal gland on the effects of antiexudative and bronchodilative drugs on antigen-induced airway responses, we examined the inhibitory effects of procaterol, a selective β₂-adrenoceptor agonist, on antigen-induced airway microvascular leakage and bronchoconstriction in adrenalectomized guinea pigs and compared them with the drug’s effects in sham-operated animals. Guinea pigs sensitized passively with anti-ovalbumin (OA) guinea-pig serum were adrenalectomized or sham-operated under urethane anesthesia and examined 30 min after surgery in the following experiments. (1) Animals were intravenously administered Evans blue dye to quantify airway plasma exudation, and then OA was inhaled for 10 min while measuring pulmonary inflation pressure, a parameter of bronchoconstriction. Procaterol (1, 3, 10, or 30 µg/kg) or saline (control) was administered into the airways 10 min prior to OA inhalation. The amount of extravasated Evans blue dye in the airways was calculated. (2) Venous blood samples were collected during OA or saline inhalation and plasma catecholamine levels were compared. In control animals, OA-induced increases in both the amount of Evans blue dye and in pulmonary inflation pressure were markedly greater in adrenalectomized animals than in sham-operated animals. Procaterol dose-dependently inhibited OA-induced airway microvascular leakage and bronchoconstriction, and its effects were more potent in adrenalectomized animals (significant at 1 µg/kg and higher) than in sham-operated animals (significant at 10 µg/kg and higher). Although the plasma concentration of epinephrine during OA inhalation was approximately 3 times higher than that during saline inhalation in sham-operated animals, no difference was seen in adrenalectomized animals. In conclusion, while procaterol essentially possesses pronounced inhibitory effects on antigen-induced airway microvascular leakage and bronchoconstriction in guinea pigs, the effects are considerably masked by epinephrine released from the adrenal gland.

Nociceptin/orphanin FQ inhibits capsaicin-induced guinea-pig airway contraction through an inward-rectifier potassium channel.

Nociceptin/orphanin FQ (N/OFQ), an endogenous opioid-like orphan receptor (NOP receptor, previously termed ORL1 receptor) agonist, has been found to inhibit capsaicin-induced bronchoconstriction in isolated guinea-pig lungs and in vivo. The underlying mechanisms are not clear. In the present studies, we tested the effect of N/OFQ on VR1 channel function in isolated guinea-pig nodose ganglia cells. Capsaicin increased intracellular Ca(2+) concentration in these cells through activation of vanilloid receptors. Capsaicin-induced Ca(2+) responses were attenuated by pretreatment of nodose neurons with N/OFQ (1 microM). N/OFQ inhibitory effect on the Ca(2+) response in nodose ganglia cells was antagonized by tertiapin (0.5 microM), an inhibitor of inward-rectifier K(+) channels, but not by verapamil, a voltage gated Ca(2+) channel blocker, indicating that an inward-rectifier K(+) channel is involved in N/OFQ inhibitory effect. In isolated guinea-pig bronchus, N/OFQ (1 microM) inhibited capsaicin-induced airway contraction. Tertiapin (0.5 microM) abolished the N/OFQ inhibition of capsaicin-induced bronchial contraction. Capsaicin (10 microg) increased pulmonary inflation pressure in the isolated perfused guinea-pig lungs. This response was significantly attenuated by pretreatment with N/OFQ (1 microM). Tertiapin also abolished the N/OFQ inhibitory effect on capsaicin-induced bronchoconstriction in perfused lungs. Capsaicin increased the release of substance P and neurokinin A from isolated lungs. N/OFQ (1 microM) blocked the capsaicin-induced tachykinin release. These results indicate that N/OFQ-induced hyperpolarization of tachykinin containing airway sensory nerves, through an inward-rectifier K(+) channel activation, accounts for the inhibition of capsaicin-evoked broncoconstriction.

Pharmaceutical dry powder aerosols: correlation of powder properties with dose delivery and implications for pharmacodynamic effect.

Efficient dispersion of bulk solids is critical for dry powder aerosol production which can be viewed as a sequence of events from stationary through dilated, flowing and finally dispersed particulates. The purpose of this study was to test the hypothesis that numerical descriptors of powder flow properties predict aerosol dispersion and pharmacodynamic effect. Drug and excipient particles were prepared in size ranges suitable for inhalation drug delivery, and their physico-chemical properties were evaluated. Novel techniques (chaos analysis of dynamic angle of repose and impact force separation) were developed and utilized to measure and characterize powder flow and particle detachment from solid surfaces, respectively. Dry powder aerosol dispersion was evaluated using inertial impaction. Pharmacodynamic evaluations of bronchodilation were performed in guinea pigs, for selected formulations. We observed a direct correlation of powder flow with ease of particle separation (r2=0.9912) and aerosol dispersion (r2= 0.9741). In vivo evaluations indicated that formulations exhibiting a higher in vitro dose delivery resulted in a greater reduction in pulmonary inflation pressure. These results integrate powder behavior at various levels and indicate that numerical descriptors of powder flow accurately predict dry powder aerosol dispersion. A proportionality between aerosol dispersion and pharmacodynamic effect was observed in preliminary in vivo evaluations, which demonstrates the potential of these techniques for correlation studies between in vitro powder properties and in vivo effect.

The effect of treatment with a new antidepressant: INN 00835 on platelet serotonin uptake in depressed patients

The effect of chronic administration of platelet-activating factor (PAF) on airway reactivity, cell recruitment and lung morphology in the guinea-pig has been investigated. Alzet osmotic minipumps delivering either PAF (7.2 mg/kg/14 days) in 0.25% (w/v) bovine serum albumin in saline (saline-BSA), acetylcholine or saline-BSA alone were implanted s.c. in the neck region of guinea-pigs and connected to the jugular vein. In some experiments, implanted and non-implanted animals were treated daily with the PAF antagonist, BN 52021 (15 mg/kg, twice a day, p.o.). On day 15 after minipump implantation, the animals were anesthetized with urethane (1.2 g/kg, i.p.) and tracheal cannula was inserted for mechanical ventilation. Pulmonary inflation pressure (PIP) was monitored and airway responsiveness was assessed by administration of increasing doses of histamine (0.2–100 μg/kg, i.v.). As compared to saline-BSA-treated or non-implanted guinea-pigs, chronic treatment of the animals with PAF induced a significant (p<0.01) increase in airway response. No significant change in airway responsiveness was observed following chronic acetylcholine administration. In contrast, regardless of the treatment of the animals, no change in the threshold dose of histamine inducing alteration in PIP was noted, suggesting that PAF induces bronchopulmonary hyperreactivity rather than hyperresponsiveness. In addition, no significant difference was observed in the in vitro responsiveness to histamine of lung parenchymal strips from animals having received PAF or saline-BSA. Histological evaluation of the lung from guinea-pigs receiving PAF exhibited moderate hyperplasia of airway smooth muscle cells, marked eosinophil infiltration and increase in mast cell number in the peribronchial zone as compared to those from control, acetylcholine- or solvent-treated animals. These alterations were markedly reduced or absent in guinea-pigs implanted with PAF-containing minipumps and concomitantly treated with BN 52021. In conclusion, chronic administration of PAF to guinea-pigs results in airway hyperreactivity to histamine in vivo, associated with structural changes in airway smooth muscle cell mass, but which is unlikely to be secondary to protracted bronchoconstriction.

Effect of vagotomy on airway hyperreactivity to endogenously released neurotransmitters at 18–24 h after inhaled antigen

Airway reactivity was examined in anaesthetized guinea-pigs 18–24 h after inhalation challenge of ovalbumin-sensitized animals with ovalbumin. Bronchoconstrictor responses were measured from the increases in pulmonary inflation pressure. The study was undertaken to examine whether ovalbumin challenge induced airway hyperreactivity to neurotransmitters released endogenously by vagal nerve stimulation. Stimulation parameters were selected to cause release of either acetylcholine (0.3 ms pulse width for 3 s, 20 V, 2–40 Hz), both acetylcholine and neuropeptide (5 ms pulse width for 15 s, 20 V, 0.5–8 Hz) or neuropeptide only, using the latter parameters in the presence of atropine. The vagi were paired for stimulation and in some experiments were cut central to the stimulation point. Frequency–response curves for acetylcholine- and neuropeptide-mediated bronchoconstrictor responses to vagal stimulation when the nerves were intact revealed no airway hyperreactivity after ovalbumin challenge. The presence of atropine failed to reveal airway hyperreactivity. However, when the vagi were cut, the frequency–response curves were displaced to the left after ovalbumin challenge compared with saline challenged animals, indicating airway hyperreactivity. This airway hyperreactivity was significant after atropine and suggests an increase in sensitivity to endogenously released neuropeptides rather than acetylcholine. It also indicates that the airway hyperreactivity is dependent on removal of the afferent vagal pathways. Frequency–response curves for cholinergic stimulation (0.3 ms) with intact vagi revealed no airway hyperreactivity after antigen challenge. Comparisons of exogenously administered 5-hydroxytryptamine (5-HT, 300 ng/100 g i.v.) and a single vagal stimulation of 0.3 ms pulse width (cholinergic) revealed no airway hyperreactivity to either stimulus after ovalbumin challenge. However if the vagi were cut, airway hyperreactivity was observed, again suggesting that removal of afferent pathways is important for revealing airway hyperreactivity in the anaesthetized guinea-pig. Ovalbumin challenge caused significant increases in the bronchoconstrictor responses to a single dose of capsaicin (50 μg/100 g i.v.) or dose–response curves to bradykinin. Since these agents release neuropeptides from sensory C-fibres, this is further support for a raised sensitivity to endogenously released neuropeptides.

Prevention of ozone-induced airway hyperresponsiveness and epithelial injury by phosphodiesterase inhibitors in guinea pigs

We investigated the effects of phosphodiesterase (PDE) inhibitors on ozone-induced airway hyperresponsiveness (AHR) in guinea pigs. Theophylline (10–100 mg/kg), rolipram (1–100 mg/kg) and T-440 (1–100 mg/kg) were orally administered 30 min before ozone exposure (3 ppm, for 30 min). Ozone exposure caused an increase in airway responsiveness to methacholine aerosol, and log PC 10 (log-transformed methacholine concentration causing a 10 cm H 2O increase in pulmonary inflation pressure) in the control and ozone-exposed group was 4.43±0.05 ( n=6) and 3.26±0.15 ( n=12), respectively. Theophylline at 100 mg/kg, 10 mg/kg rolipram and 10 mg/kg T-440 significantly inhibited AHR with log PC 10 value of 4.73±0.16 ( n=7), 3.74±0.11 ( n=7), 3.82±0.15 ( n=6), respectively. On histological examination, epithelial damage in the trachea and peripheral airways was recognized after ozone exposure. At 100 mg/kg, rolipram, T-440 and theophylline caused complete inhibition of AHR, and prevented epithelial damage of the trachea and peripheral airways. These results indicate that PDE inhibitors prevent not only ozone-induced AHR but also airway epithelial injury by ozone.

Effects of FR173657, a non‐peptide B2 antagonist, on kinin‐induced hypotension, visceral and peripheral oedema formation and bronchoconstriction

1. Kinins are believed to play a key role in many inflammatory conditions. Therefore, bradykinin antagonists are being developed for potential therapeutic applications. In the present investigation we describe the pharmacology, in vivo, of (E)-3-(6-acetamido-3-pyridyl)-N-[N-[2,4-dichloro-3-[2-methyl-8-quinoliny l) oxymethyl]phenyl]-N-methylaminocarbonylmethyl]acrylamide (FR173657), a novel, non-peptide bradykinin antagonist. 2. The hypotensive effects of i.v. injections of bradykinin (50 pmol) in captopril-pre-treated anaesthetized rats were significantly inhibited by 100 nmol kg-1 FR173657 s.c., and completely abolished by 300 nmol kg-1. The full inhibitory effect developed within 60 min and remained unchanged for at least 4 h. However, the effect was reversible, since 24 h after an injection of 300 nmol kg-1 FR173657 no inhibitory effect could be observed. 3. The plasma protein extravasation into the pancreas and duodenum induced by an i.v. infusion of bradykinin (11 nmol kg-1 within 20 min) in captopril-treated anaesthetized rats was completely abolished by FR173657 at doses of 30 nmol kg-1 s.c. and above, given 60 min before bradykinin. FR173657 3 nmol kg-1 was ineffective, while a dose of 10 nmol kg-1 produced an intermediate effect. 4. The paw oedema induced by the subplantar injection of bradykinin (30 nmol) in anaesthetized rats was inhibited slightly by s.c. injection of FR173657 0.3 mumol kg-1, whereas 1 and 3 mumol kg-1 produced significant inhibition of the bradykinin-induced oedema. The maximum inhibition amounted to about 50% and could not be increased even when the dose of FR173657 was increased to 30 mumol kg-1. FR173657 did not effect the oedema caused by histamine or 5-hydroxytryptamine. 5. Bradykinin (20 nmol kg-1, i.v.) caused increases in pulmonary inflation pressure by 300-600 Pa in anaesthetized, respirated guinea-pigs. The effect was reduced to 58 +/- 9% of the initial value 60 min after the s.c. injection of FR173657 1 mumol kg-1, whereas only 9 +/- 7% remained after 10 mumol kg-1. The bronchoconstrictor actions of histamine remained unaffected by FR173657. 6. In summary, FR173657 is a highly potent and selective bradykinin antagonist. The inhibitory action in vivo lasts for longer than 4 h but is fully reversible. FR173657, or similar compounds, will be a useful tool for the pharmacological investigation of pathophysiological states and may possess a therapeutic potential in diseases involving the endogenous release of kinins.

Investigation of the contributions of nitric oxide and prostaglandins to the actions of endothelins and sarafotoxin 6c in rat isolated perfused lungs.

1. The aims of the study were to assess the contribution of prostaglandins and nitric oxide (NO) to the effects of endothelin (ETs) and sarafotoxin 6c (SX6c) in perfused rat lungs. This was carried out by using indomethacin, a cyclo-oxygenase inhibitor and NG-nitro-L-arginine (L-NOARG), a NO synthase inhibitor. Responses were studied under basal perfusion conditions and in other experiments after the elevation of vascular tone with the thromboxane-mimetic, U46619. The sub-types of ET receptors involved were characterized by use of ET receptor antagonists and cross-tachyphylaxis. 2. Pulmonary perfusion pressure (PPP), lung weight and pulmonary inflation pressure (PIP), were continuously recorded. Although L-NOARG (100 microM) did not alter basal parameters it markedly augmented the vasoconstriction and lung weight increases induced by ET-1 (50-400 pmol) or SX6C (25-200 pmol) while vasoconstrictor responses to phenylephrine were not affected by L-NOARG. 3. L-NOARG markedly potentiated the bronchoconstriction induced by ET-1 or SX6C whereas it had no effect on responses to carbachol. 4. When vascular tone was elevated, low doses (1.25-40 pmol) of ET-1, ET-3 and SX6C produced falls in PPP. The vasodilator potencies were SX6C > ET-1 = ET-3. The ETA receptor antagonist, BQ123, did not affect these depressor responses whereas the mixed ETA/ETB antagonist, bosentan, blocked them. 5. Indomethacin (10 microM) partially inhibited vasodilator response to ET-1, whereas it had no effect on SX6C-induced vasodilation. 6. L-NOARG plus indomethacin completely blocked ET-1 induced vasodilation, whereas responses to SX6C were blocked by L-NOARG alone. 7. Repeated injections of submaximal doses of ET-1 or SX6C caused tachyphylaxis to vasodilator responses. Subsequent injections of SX6C or ET-1 did not elicit depressor responses showing cross tachyphylaxis had occurred. 8. These findings indicate that under basal conditions the pulmonary vasoconstrictor, lung weight and bronchoconstrictor responses to ET-1 and SX6C are attenuated by evoked release of nitric oxide (NO). When vascular tone was elevated, lower doses of ETs and SX6C produced vasodilatation. These vasodilator responses are indirect, those to SX6C being mediated via NO production, whereas those to ET-1 involve both NO and prostanoid(s). Tachyphylaxis and ET antagonist experiments indicate that the same receptor subtype is involved in mediating the vasodilatation and that this is of the ETB type located on the endothelium. However the post-receptor vasodilator events triggered by ET-1 or SX6C appear to be different.

The inhibition of antigen-induced eosinophilia and bronchoconstriction by CDP840, a novel stereo-selective inhibitor of phosphodiesterase type 4.

1. The novel tri-aryl ethane CDP840, is a potent and selective inhibitor of cyclic AMP phosphodiesterase type 4 (PDE 4) extracted from tissues or recombinant PDE 4 isoforms expressed in yeast (IC50S: 4-45 nM). CDP840 is stereo-selective since its S enantiomer (CT 1731) is 10-50 times less active against all forms of PDE 4 tested while both enantiomers are inactive (IC50S: > 100 microM) against PDE types 1, 2, 3 and 5. 2. Oral administration of CDP840 caused a dose-dependent reduction of interleukin-5 (IL-5)-induced pleural eosinophilia in rats (ED50 = 0.03 mg kg-1). The eosinophils in pleural exudates from CDP840-treated animals contained higher levels of eosinophil peroxidase (EPO) than cells from control animals, suggesting a stabilizing effect on eosinophil degranulation. CDP840 was approximately equi-active with the steroid dexamethasone in this model and was 10-100 times more potent than the known PDE 4-selective inhibitors rolipram and RP73401. The activity of CDP840 was not influenced by adrenalectomy, beta-sympathomimetics or beta-sympatholytics. 3. Antigen-induced pulmonary eosinophilia in sensitized guinea-pigs was reduced dose-dependently by CDP840 (0.01-1 mg kg-1, i.p.) and intracellular EPO levels were significantly higher. CDP840 was more potent in these activities than CT1731 or rolipram and comparable in potency to RP73401. 4. Rolipram or CDP840 were less active than dexamethasone in preventing neutrophil accumulation, or exudate formation in carrageenan-induced pleurisy in rats and thus do not exhibit general anti-inflammatory activity. 5. In sensitized guinea-pigs, aerosols of the antigen ovalbumin caused a dose-dependent bronchoconstriction demonstrated by an increase in pulmonary inflation pressure. Administration of CDP840 (0.001-1.0 mg kg-1, i.p.), 1 h before antigen challenge, resulted in dose-dependent reduction in response to antigen. This activity was not due to bronchodilatation since higher doses of CDP840 (3 mg kg-1) did not significantly change the bronchoconstrictor response to histamine. Rolipram was approximately 10 times less active than CDP840 in preventing antigen-induced bronchoconstriction. 6. These results confirm the observations that selective PDE 4 inhibitors reduce antigen-induced bronchoconstriction and pulmonary eosinophilic inflammation. CDP840 is more potent than rolipram in inhibiting native or recombinant PDE 4. Unlike the recently described potent PDE 4 inhibitor RP73401, CDP840 is more active than rolipram in the rat IL-5 model following oral administration. The novel series of tri-aryl ethanes, of which CDP840 is the lead compound, could be the basis of an orally active prophylactic treatment for human asthma.

Involvement of tachykinin NK1 and NK2 receptors in substance P-induced microvascular leakage hypersensitivity and airway hyperresponsiveness in guinea-pigs.

Tachykinins, such as substance P, might be involved in the development of airway hyperresponsiveness (AHR) and airway inflammation. However, it is unknown which tachykinin receptors mediate these biological activities. The effects of two antagonists of tachykinin neurokinin-1 (NK1) and tachykinin neurokinin-2 (NK2) receptors, SR 140333 and SR 48968, respectively, were investigated on substance P (SP)-induced airway hyperresponsiveness and potentiation of the histamine-induced increase in microvascular leakage, in phosphoramidon-pretreated guinea-pigs. Guinea-pigs were pretreated with phosphoramidon (0.1 mM aerosol for 15 min) and exposed 15 min later to saline solution alone or to saline solution containing SP (0.1 mg.mL-1 for 30 min). Twenty four hours later, the animals were anaesthetized and prepared for the recording of the pulmonary inflation pressure (PIP) to acetylcholine or for the investigation of microvascular leakage to histamine. Pretreatment of the guinea-pigs with a single dose of SR 48968 (1 mg.kg-1, i.p.) 30 min before SP exposure, significantly prevented the development of AHR, whereas SR 140333 (1 mg.kg-1, i.p.) did not. In a second set of experiments, phosphoramidon-pretreated guinea-pigs exposed to SP presented a significant potentiation of the histamine-induced increase in microvascular leakage in pulmonary airways. When the guinea-pigs were pretreated with SR 140333, an inhibition of the increased microvascular leakage to histamine was observed. In contrast, no significant inhibitory activity was noted when the guinea-pigs were pretreated with SR 48968. The present data demonstrate the importance of tachykinin NK2 receptor stimulation in the development of airway hyperresponsiveness and that of tachykinin NK1 receptor stimulation in microvascular leakage hypersensitivity in phosphoramidon-pretreated and substance P-exposed guinea-pigs. The results also suggest a dissociation between the presence of microvascular leakage and the occurrence of airway hyperresponsiveness.

Importance of impairment of the airway epithelium for ozone-induced airway hyperresponsiveness in guinea pigs.

We examined the relationship between ozone (O3)-induced airway hyperresponsiveness (AHR) and inflammation in guinea pigs. Inhalation of methacholine (MCh) was adopted in the time course study of AHR that was assessed by measuring pulmonary inflation pressure after O3 exposure (3 ppm, for 2 hr) because the degree of AHR detected by inhalation of MCh was greater than that detected by i.v. administration. AHR was detected up to 5 hr after O3 exposure and was not observed at 24 and 48 hr. In the bronchoalveolar lavage (BAL) study, the numbers of neutrophils, eosinophils, lymphocytes and macrophages in BAL fluid (BALF) reached maximum at 24 hr or later. On the other hand, the number of airway epithelial cells in the BALF significantly increased at 2 and 5 hr. In the histological study, disorder and impairment of the airway epithelium in the trachea and lung were observed at 2 and 5 hr. Changes in the airway epithelium were recovered at 48 hr, although an increase in leukocytes was observed in the lung. These results indicate that O3-induced AHR in guinea pigs is most probably associated with impairment of the epithelium rather than with infiltration of inflammatory cells in the airway.