Differences in fish communities on natural versus artificial temperate reefs, groundfish conservation applications in British Columbia.

Human-made marine habitats such as artificial reefs are used to mitigate marine habitat degradation and aid conservation of species at risk. We used ROV and sonar to survey threatened rockfish (Sebastes spp.) and other groundfish species associated with 18 artificial and natural reefs along the south coast of BC, Canada. Using an information-theoretic approach, we found that community composition significantly differed between natural and artificial reefs. Artificial reefs had high variability in rockfish abundance, some supporting very high or low relative abundance. Natural reefs consistently supported intermediate rockfish abundances. Groundfish diversity was significantly greater on natural reefs than artificial reefs. Depth and relief were significant predictors for both abundance and species richness. Interestingly, rockfish abundance was negatively associated with proximity to nearest rockfish conservation area. This research is a first step in understanding causal mechanisms leading to differences between fish communities on artificial reefs in our study system, and which reef attributes may facilitate successful contributions to conservation. Though artificial reefs show promise in the conservation of some threatened species, the maintenance of diverse fish communities depends on protection of heterogenous natural reef communities.

CO(2) rebreathing: an undergraduate laboratory to study the chemical control of breathing.

The Read CO2 rebreathing method (Read DJ. A clinical method for assessing the ventilatory response to carbon dioxide. Australas Ann Med 16: 20-32, 1967) provides a simple and reproducible approach for studying the chemical control of breathing. It has been widely used since the modifications made by Duffin and coworkers. Our use of a rebreathing laboratory to challenge undergraduate science students to investigate the control of breathing provided 8 yr of student-generated data for comparison with the literature. Students (age: 19-22 yr, Research Ethics Board approval) rebreathed from a bag containing 5% CO2 and 95% O2 (to suppress the peripheral chemoreflex to hypoxia). Rebreathing was performed, and ventilation measured, after hyperventilation to deplete tissue CO2 stores and enable the detection of the central chemoreflex threshold. We analyzed 43 data sets, of which 10 were rejected for technical reasons. The mean threshold and ventilatory sensitivity to CO2 were 43.3 ± 3.8 mmHg and 4.60 ± 3.04 l·min(-1)·mmHg(-1) (means ± SD), respectively. Threshold values were normally distributed, whereas sensitivity was skewed to the left. Both mean values agreed well with those in the literature. We conclude that the modified rebreathing protocol is a robust method for undergraduate investigation of the chemical control of breathing.

OVA-induced airway hyperresponsiveness alters murine heart rate variability and body temperature.

Altered autonomic (ANS) tone in chronic respiratory disease is implicated as a factor in cardiovascular co-morbidities, yet no studies address its impact on cardiovascular function in the presence of murine allergic airway (AW) hyperresponsiveness (AHR). Since antigen (Ag)-induced AHR is used to model allergic asthma (in which ANS alterations have been reported), we performed a pilot study to assess measurement feasibility of, as well as the impact of allergic sensitization to ovalbumin (OVA) on, heart rate variability (HRV) in a murine model. Heart rate (HR), body temperature (T(B)), and time- and frequency-domain HRV analyses, a reflection of ANS control, were obtained in chronically instrumented mice (telemetry) before, during and for 22 h after OVA or saline aerosolization in sensitized (OVA) or Alum adjuvant control exposed animals. OVA mice diverged significantly from Alum mice with respect to change in HR during aerosol challenge (P < 0.001, Two-Way ANOVA; HR max change Ctrl = +80 ± 10 bpm vs. OVA = +1 ± 23 bpm, mean ± SEM), and displayed elevated HR during the subsequent dark cycle (P = 0.006). Sensitization decreased the T(B) during aerosol challenge (P < 0.001). Sensitized mice had decreased HRV prior to challenge (SDNN: P = 0.038; Low frequency (LF) power: P = 0.021; Low/high Frequency (HF) power: P = 0.042), and increased HRV during Ag challenge (RMSSD: P = 0.047; pNN6: P = 0.039). Sensitized mice displayed decreased HRV subsequent to OVA challenge, primarily in the dark cycle (RMSSD: P = 0.018; pNN6: P ≤ 0.001; LF: P ≤ 0.001; HF: P = 0.040; LF/HF: P ≤ 0.001). We conclude that implanted telemetry technology is an effective method to assess the ANS impact of allergic sensitization. Preliminary results show mild sensitization is associated with reduced HRV and a suppression of the acute T(B)-response to OVA challenge. This approach to assess altered ANS control in the acute OVA model may also be beneficial in chronic AHR models.

A dp/dt method to assess dynamic properties of lung mechanoreceptors.

The activity of airway slowly adapting mechanoreceptors (SARs) reflects the presence of both a static and dynamic component. The dynamic response is typically assessed by the adaptation index; however, this is an indirect reflection of the more appropriate physiological stimulus, the rate of change of inflation pressure (dp/dt). We describe a method in which measurement of receptor discharge exceeding the SAR static response is used to measure dynamic discharge and dynamic sensitivity of lung mechanoreceptors. Repeat inflations with varying dp/dt illustrate the method for a SAR in which the dynamic sensitivity is inversely related to dp/dt and the initial "onset" discharge is highly dp/dt sensitive. The method may provide new insight into the classification and behaviour of lung mechanoreceptors.

Vascular defects in gain-of-function fps/fes transgenic mice correlate with PDGF- and VEGF-induced activation of mutant Fps/Fes kinase in endothelial cells.

BACKGROUND: Fps/Fes is a cytoplasmic tyrosine kinase that is abundantly expressed in the myeloid, endothelial, epithelial, neuronal and platelet lineages. Genetic manipulation in mice has uncovered potential roles for this kinase in hematopoiesis, innate immunity, inflammation and angiogenesis. OBJECTIVE: We have utilized a genetic approach to explore the role of Fps/Fes in angiogenesis. METHODS: A hypervascular line of mice generated by expression of a 'gain-of-function' human fps/fes transgene (fps(MF)) encoding a myristoylated variant of Fps (MFps) was used in these studies. The hypervascular phenotype of this line was extensively characterized by intravital microscopy and biochemical approaches. RESULTS: fps(MF) mice exhibited 1.6-1.7-fold increases in vascularity which was attributable to increases in the number of secondary vessels. Vessels were larger, exhibited varicosities and disorganized patterning, and were found to have defects in histamine-induced permeability. Biochemical characterization of endothelial cell (EC) lines derived from fps(MF) mice revealed that MFps was hypersensitive to activation by vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF). CONCLUSIONS: MFps mediates enhanced sensitization to VEGF and PDGF signaling in ECs. We propose that this hypersensitization contributes to excessive angiogenic signaling and that this underlies the observed hypervascular phenotype of fps(MF) mice. These phenotypes recapitulate important aspects of the vascular defects observed in both VEGF and angiopoietin-1 transgenic mice. The fps/fes proto-oncogene product therefore represents a novel player in the regulation of angiogenesis, and the fps(MF) line of mice constitutes a unique new murine model for the study of this process.

G protein-coupled receptor kinase 5 regulates airway responses induced by muscarinic receptor activation.

G protein-coupled receptors (GPCRs) transduce extracellular signals into intracellular events. The waning responsiveness of GPCRs in the face of persistent agonist stimulation, or desensitization, is a necessary event that ensures physiological homeostasis. GPCR kinases (GRKs) are important regulators of GPCR desensitization. GRK5, one member of the GRK family, desensitizes central M(2) muscarinic receptors in mice. We questioned whether GRK5 might also be an important regulator of peripheral muscarinic receptor responsiveness in the cardiopulmonary system. Specifically, we wanted to determine the role of GRK5 in regulating muscarinic receptor-mediated control of airway smooth muscle tone or regulation of cholinergic-induced bradycardia. Tracheal pressure, heart rate, and tracheal smooth muscle tension were measured in mice having a targeted deletion of the GRK5 gene (GRK5(-/-)) and littermate wild-type (WT) control mice. Both in vivo and in vitro results showed that the airway contractile response to a muscarinic receptor agonist was not different between GRK5(-/-) and WT mice. However, the relaxation component of bilateral vagal stimulation and the airway smooth muscle relaxation resulting from beta(2)-adrenergic receptor activation were diminished in GRK5(-/-) mice. These data suggest that M(2) muscarinic receptor-mediated opposition of airway smooth muscle relaxation is regulated by GRK5 and is, therefore, excessive in GRK5(-/-) mice. In addition, this study shows that GRK5 regulates pulmonary responses in a tissue- and receptor-specific manner but does not regulate peripheral cardiac muscarinic receptors. GRK5 regulation of airway responses may have implications in obstructive airway diseases such as asthma or chronic obstructive pulmonary disease.

Calcium-independent contraction and sensitization of airway smooth muscle by p21-activated protein kinase.

In Triton-skinned phasic ileal smooth muscle, constitutively active recombinant p21-activated kinase (PAK3) has been shown to induce Ca(2+)-independent contraction, which is accompanied by phosphorylation of caldesmon and desmin (Van Eyk JE, Arrell DK, Foster DB, Strauss JD, Heinonen TY, Furmaniak-Kazmierczak E, Cote GP, and Mak AS. J Biol Chem 273: 23433-23439, 1998). In the present study, we investigated whether PAK has a broad impact on smooth muscle in general by testing the hypothesis that PAK induces Ca(2+)-independent contractions and/or Ca(2+) sensitization in tonic airway smooth muscle and that the process is mediated via phosphorylation of caldesmon. In the absence of Ca(2+) (pCa > 9), constitutively active glutathione-S-transferase-murine PAK3 (GST-mPAK3) caused force generation of Triton-skinned canine tracheal smooth muscle (TSM) fibers to approximately 40% of the maximal force generated by Ca(2+) at pCa 4.4. In addition, GST-mPAK3 enhanced Ca(2+) sensitivity of contraction by increasing force generation by 80% at intermediate Ca(2+) concentrations (pCa 6.2), whereas it had no effect at pCa 4.4. Catalytically inactive GST-mPAK3(K297R) had no effect on force production. Using antibody against one of the PAK-phosphorylated sites (Ser(657)) on caldesmon, we showed that a basal level of phosphorylation of caldesmon occurs at this site in skinned TSM and that PAK-induced contraction was accompanied by a significant increase in the level of phosphorylation. Western blot analyses show that PAK1 is the predominant PAK isoform expressed in murine, rat, canine, and porcine TSM. We conclude that PAK causes Ca(2+)-independent contractions and produces Ca(2+) sensitization of skinned phasic and tonic smooth muscle, which involves an incremental increase in caldesmon phosphorylation.

Role of vagal C-fiber afferents in the bronchomotor response to lactic acid in the newborn dog.

We addressed the hypothesis that vagal C-fiber afferents and cyclooxygenase products are the mechanisms responsible for lactic acid (LA)-induced bronchoconstriction in the newborn dog. Perineural capsaicin and indomethacin were used to block conduction of vagal C fibers and production of cyclooxygenase products, respectively. Perineural capsaicin eliminated (85%) the increase in lung resistance (RL; 45 +/- 5.6%) due to capsaicin (25 microg/kg), whereas the increase in RL (54 +/- 6.9%) due to LA (0.4 mmol/kg) was only inhibited by 37 +/- 4.7% (P < 0.05). Atropine reduced LA-induced bronchoconstriction (42 +/- 2.1%) by an amount similar to that obtained with perineural capsaicin. However, inhibition was significantly increased when atropine was combined with indomethacin (61 +/- 2.7%; P < 0.05), implicating cyclooxygenase products in the LA-induced bronchoconstrictor response. We conclude that the mechanisms responsible for LA-induced bronchoconstriction in the newborn are 1) activation of vagal C-fibers, which, through projections to medullary respiratory centers, leads to activation of vagal cholinergic efferents; 2) production of cyclooxygenase products, which cause bronchoconstriction independent of medullary involvement; and 3) an unknown bronchoconstrictor mechanism, putatively tachykinin mediated. On the basis of our data, pharmaceutical targeting of pulmonary afferents would prevent multiple downstream mechanisms that lead to airway narrowing due to inflammatory lung disease.

A new method to perform quantitative measurement of bronchoscopic images.

Bronchoscopy is a commonly used clinical tool that provides a direct image of the bronchial lumen. However, bronchoscopy has seen little use as a quantitative measurement tool, mainly because of the wide-angle lens which distorts the image. The present authors have tested the ability of numerical algorithms and commercial software to correct for this distortion. Test objects of known size were imaged with four different bronchoscopes. Commercial image analysis software was used to measure the size of features in the images before and after applying distortion correction algorithms. The technique was then applied by measuring airway narrowing in anaesthetized pigs during vagal stimulation. Without correction, object size near the edge of the field of view is underestimated by approximately 40%. The error in measured diameter of concentric circles was dependent on the radius of the circle, increasing to 25% for circles occupying 90% of the field. Third order polynomial functions were required to correct these errors. After correction, errors were independent of object size or location in the image. Correction for lens distortion was independent of the distance between bronchoscope and object. The authors conclude that modern image processing software can correct for the distortion produced by wide-angle bronchoscope lenses.

Altered airway and cardiac responses in mice lacking G protein-coupled receptor kinase 3.

Contraction and relaxation of airway smooth muscles is mediated, in part, by G protein-coupled receptors (GPCRs) and dysfunction of these receptors has been implicated in asthma. Phosphorylation of GPCRs, by G protein-coupled receptor kinase (GRK), is an important mechanism involved in the dampening of GPCR signaling. To determine whether this mechanism might play a role in airway smooth muscle physiology, we examined the airway pressure time index and heart rate (HR) responses to intravenous administration of the cholinergic agonist methacholine (MCh) in genetically altered mice lacking one copy of GRK2 (GRK2 +/-), homozygous GRK3 knockout (GRK3 -/-), and wild-type littermates. (GRK2 -/- mice die in utero.) GRK3 -/- mice demonstrated a significant enhancement in the airway response to 100 and 250 microgram/kg doses of MCh compared with wild-type and GRK2 +/- mice. GRK3 -/- mice also displayed an enhanced sensitivity of the airway smooth muscle response to MCh. In addition, GRK3 -/- mice displayed an altered HR recovery from MCh-induced bradycardia. Although direct stimulation of cardiac muscarinic receptors measured as vagal stimulation-induced bradycardia was similar in GRK3 -/- and wild-type mice, the baroreflex increase in HR associated with sodium nitroprusside-induced hypotension was significantly greater in GRK3 -/- than wild-type mice. Therefore, these data demonstrate that in the mouse, GRK3 may be involved in modulating the cholinergic response of airway smooth muscle and in regulating the chronotropic component of the baroreceptor reflex.

Mechanisms of capsaicin- and lactic acid-induced bronchoconstriction in the newborn dog.

1. Capsaicin activation of the pulmonary C fibre vanilloid receptor (VR1) evokes the pulmonary chemoreflex and reflex bronchoconstriction. Among potential endogenous ligands of C fibre afferents, lactic acid has been suggested as a promising candidate. We tested the hypotheses that (a) lactic acid behaves as a stimulant of C fibre receptors in the newborn dog to cause reflex bronchoconstriction, and (b) lactic acid causes reflex bronchoconstriction via the same pulmonary C fibre receptor mechanism as capsaicin using the competitive capsaicin/VR1 receptor antagonist capsazepine. 2. Right heart injection of lactic acid caused a significant increase (47 +/- 8.0 %) in lung resistance (RL) that was atropine sensitive (reduced by 75 %; P < 0.05), consistent with reflex activation of muscarinic efferents by stimulation of C fibre afferents. 3. Infusion of the competitive capsaicin antagonist capsazepine caused an 80 % reduction (P < 0.01) in the control bronchoconstrictor response (41 +/- 8.5 % increase in RL) to right heart injections of capsaicin. The effects of capsazepine are consistent with reversible blockade of the VR1 receptor to abolish C fibre-mediated reflex bronchoconstriction. 4. Lactic acid-evoked increases in RL were unaffected by VR1 blockade with capsazepine, consistent with a separate lactic acid-induced reflex mechanism. 5. We conclude that (a) putative stimulation of C fibres with lactic acid causes reflex bronchoconstriction in the newborn dog, (b) capsazepine reversibly antagonizes reflex bronchoconstriction elicited by right heart injection of capsaicin, presumably by attenuating capsaicin-induced activation of the C fibre 'capsaicin' receptor (VR1), and (c) capsazepine resistance of lactic acid-induced bronchoconstriction indicates that lactic acid evokes reflex bronchoconstriction by a separate mechanism, possibly via the acid-sensing ionic channel.

Quantitative densitometry of proteins stained with coomassie blue using a Hewlett Packard scanjet scanner and Scanplot software.

In the present study we evaluated the performance of a software/scanner system that employed the Hewlett Packard (HP) ScanJet Plus and Scanplot Software for densitometric quantification of protein loads stained with Coomassie Brilliant Blue following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Gels with bovine serum albumin (BSA) standards, ranging from 0.125 to 10 micrograms, were scanned using reflectance densitometry with 127 microns step size in both the x and y directions and a resolution of 200 dots per inch. Densitometric volume was calculated for each protein band from scanner output in the tagged image file format (TIFF) by a customized software package, Scanplot V. 4.05 (Cunningham Engineering). Protein loads between 0.125 and 10.0 micrograms vs. volume were fit by a second-order regression: Volume = -0.58 x protein load2 + 16.82 x protein load + 7.87 (r = 0.991, p < 0.01). The same gels were scanned and quantified using a transmittance laser densitometer; densitometric volumes measured by both systems were highly correlated (r2 = 0.981, p < 0.01). Additional gels of BSA, smooth muscle myosin heavy chain (myosin), and actin displayed linear relationships between protein loads up to 4.0 micrograms and densitometric volume reflecting unique dye binding properties. We conclude that accurate and reproducible quantitative densitometry of SDS-PAGE can be performed using the HP ScanJet Plus scanner and Scanplot software.

Quantitative videobronchoscopy: a new technique to assess airway caliber.

Quantitative assessment of airway caliber is generally confined to indirect physiologic methods or to radiographic techniques. Fiberoptic bronchoscopy provides a direct view of airways, permitting quantification of airway caliber by image analysis. We investigated the characteristics of a bronchoscopic imaging system, determined its limitations in quantification and the corrections necessary for accurate assessment of image dimension, validated the methodology with airway models, and applied the technique to airways in vivo. The system comprised a bronchoscope, videocamera, videocassette recorder (VCR), computer with a frame grabber, and image-analysis program. Image quantification was affected by two sources of distortion: (1) Distance distortion: a loss of image resolution with increasing distance between the object and bronchoscope, requiring determination of the operational distance range. (2) Radial distortion: a progressive reduction in image size from the center to the periphery of the bronchoscopic field of view (FOV), requiring correction of airway dimension according to airway size and location in FOV. Validation of the methodology with different sized airway models indicated an underestimation of measured diameters, which normalized with distortion correction. We provide an example of quantitative videobronchoscopy with measurements of in vivo airway narrowing due to vagal stimulation in the anesthetized dog. Measurements of airway narrowing made with videobronchoscopy were also compared with those made with high-resolution computer-assisted tomography (HRCT) which suggested that the two technologies provide unique but complementary perspectives on airway dimensions. We conclude that videobronchoscopy and image analysis provide a novel and accurate method for the quantification of airway caliber.

Cardiopulmonary actions of muscarinic receptor subtypes in the newborn dog.

We addressed the role of muscarinic receptor subtypes in neurally mediated bronchoconstriction in vivo and airway smooth muscle contraction in vitro in the newborn dog. The in vivo dose-response effects of "selective" muscarinic antagonists on changes in lung resistance (RL) and heart rate (HR) in response to electrical stimulation of the vagus nerves were obtained in four groups of newborns. Each group was exposed to a different muscarinic antagonist: M1-selective pirenzepine (pir), M2-selective AF-DX 116 (11-[2-[(diethylamino)methyl]-1-piperidinyl]acetyl-5,11-dihydro-6H-pyrid o- [2,3-b]-[1,4]-benzodiazepine-6-one), M3-selective p-F-HHSiD (p-fluoro-hexahydro-sila-difenidol), and nonselective atropine (atr). In vitro concentration-response effects of pir and AF-DX 116 were obtained for neurally induced contractions of tracheal smooth muscle, elicited by electrical field stimulation. In a separate series of experiments we measured the bronchoconstrictor response to the muscarinic agonist acetylcholine delivered by right heart injection. Muscarinic antagonists reduced RL and HR responses to vagal stimulation in a dose-dependant fashion; however, ED50 values and selectivity for airway and cardiac responses (HR/RL ED50 ratio) were significantly different between antagonists. The rank order of potencies for inhibition of the increase in RL was atr > pir, M1 > p-F-HHSiD, M3 > AF-DX 116, M2, while that for HR was atr > AF-DX 116 > pir > p-F-HHSiD. AF-DX 116 preferentially inhibited the HR response, as reflected by the lowest HR/RL ED50 ratio (p < 0.001). The remaining antagonists preferentially inhibited RL, with the highest HR/RL ED50 ratio seen for p-F-HHSiD. These data suggest that muscarinic receptor subtypes are differentiated at birth and mediate cardiac and airway responses to vagal stimulation. We did not find autoinhibitory actions of airway M2 receptors on either the in vivo bronchoconstrictor response or the in vitro contractile response to electrical field stimulation. This suggests that neonatal airway M2 receptors, but not myocardial M2 receptors, are reduced in number or weakly coupled to muscarinic signal transduction mechanisms. Direct activation of airway smooth muscle by acetylcholine caused dose-dependent increases in RL that reached a plateau at approximately 200% at 100 micrograms, similar to values reported for vagal stimulation.

Hering-Breüer reflexes in high-altitude infants.

1. Pulmonary ventilation was found to be similar in high-altitude and low-altitude newborn infants, but the breathing pattern was deeper and slower at high altitude (Mortola, J.P., Frappell, P.B., Frappell, D.E., Villena-Cabrera, N., Villena-Cabrera, M., Peña, F., Am Rev Respir Dis 1992, 46: 1206-9). We questioned the contribution of vagal reflexes to these differences in breathing pattern. 2. Measurements were performed on high-altitude (La Paz, Bolivia, 3600-4050 m, inspired O2 pressure approximately 92 mmHg, n = 34) and low-altitude infants (Santa Cruz, Bolivia, 400 m, PIO2 approximately 141 mmHg, n = 26). The strength of the Hering-Breüer inspiratory inhibitory reflex was estimated from the inspiratory time during a respiratory effort against airways closed at end-expiration (Tloccl). The strength of the Hering-Breüer expiratory facilitatory reflex was estimated from the expiratory duration when airways were occluded during expiration (TEoccl). 3. Tloccl was significantly longer than the open-airways TI at both low and high altitude, but significantly more so (approximately 14%) at high altitude. TEoccl was longer than open-airways TE in both groups of infants, but significantly less so at high altitude, whether TEoccl was compared between occlusions of similar tidal volume (on average, TEoccl at high altitude was 79% of that at low altitude) or similar airway pressure (87%). 4. The results suggest that at high altitude the contribution of the phasic volume-dependent vagal input to the inspiratory off-switch threshold is higher, and that the tonic vagal expiratory facilitation is lower, than at low altitude, presumably because of hypoxia.(ABSTRACT TRUNCATED AT 250 WORDS)

Bronchomotor responses to hypoxia and hypercapnia in decerebrate cats.

Decerebrate animals are often used in investigations of the control of breathing because anesthesia-induced depression of respiratory reflexes is absent. We therefore investigated the level of tone and responsiveness of airway smooth muscle in seven decerebrate, paralyzed, and ventilated cats. Specifically, we measured the changes in pulmonary resistance (RL) and dynamic pulmonary compliance (CLdyn) in response to hypoxia and hypercapnia. All cats responded to hypoxia (approximately 10% O2 in N2) with significant increases (mean 49%, range 5-156%) in RL from a mean control value of 0.0197 +/- 0.0081 (SD) cmH2O.ml-1.s. During inhalation of 5% CO2 in O2, RL increased significantly (mean 59%, range 16-135%) from a mean control value of 0.0190 +/- 0.0056 cmH2O.ml-1.s. Decreases in CLdyn during hypoxia and hypercapnia were much smaller, averaging -9 and -11%, respectively. After atropine was administered, average control RL fell 50%, from 0.0269 to 0.0134 cmH2O.ml-1.s (P < 0.05; n = 4). Hypoxic and hypercapnic gas mixtures did not affect pulmonary mechanics after atropine was administered. In three cats, oscillations of RL were synchronized to phrenic activity but only at low respiratory frequencies (approximately 12 cycles/min), indicating that airway smooth muscle responded slowly to vagal input. Pentobarbital sodium, like atropine, reduced control RL in three cats. These cats lost their bronchoconstrictor response to hypercapnia but had augmented responses to hypoxia compared with preanesthetic responses. We conclude that decerebrate cats possess resting bronchomotor tone and retain their responsiveness to hypoxia and hypercapnia. Thus the decerebrate cat is a useful model for studying the control of tracheobronchial smooth muscle.

[Physiological basis for the use of muscarine antagonists in bronchopulmonary dysplasia]. / Bases physiologiques de l'utilisation des antagonistes muscariniques dans les dysplasies bronchopulmonaires.

The rationale for the use of muscarinic antagonists in bronchopulmonary dysplasia (BPD) is based on the physiology and pharmacology of airway smooth muscle, the pathology of BPD, and the response of infants with BPD to bronchodilators, in vivo and in vitro studies of airway smooth muscle of newborn animals and humans indicate that vagal efferent airway innervation and/or muscarinic receptors are functional at birth, as well as early in gestation. Current concepts regarding muscarinic receptor subtypes suggest that M3 receptors mediate airway smooth muscle contraction, M2 receptors are autoinhibitory and limit vagally-mediated bronchoconstriction, and M1 receptors may play a facilitatory role in ganglionic transmission. Muscarinic receptor subtypes appear to be functionally expressed at birth but may undergo developmental regulation. Infants with BPD have an elevated pulmonary resistance that is accompanied by hypertrophy of airway smooth muscle, b2-agonists cause bronchodilation in BPD as does atropine in infants recovering from severe BPD. The synthetic congener of atropine, ipratropium bromide (IPB) causes bronchodilation in ventilator-dependent infants with BPD in a dose-dependent fashion. Nebulized IPB causes a decrease in respiratory resistance that reaches a maximum of 20% at 175 mg. The bronchodilation seen with muscarinic antagonists suggests that part of the elevated resistance associated with BPD is due to increased muscarinic tone, presumably vagal in origin. When IPB is combined with salbutamol (0.04 mg) the response is increased in magnitude and duration; reaching a slightly larger decreases in resistance (26%) that is now accompanied by an increase in compliance (20%).(ABSTRACT TRUNCATED AT 250 WORDS)

Epithelial modulation of neonatal and fetal porcine bronchial contractile responses.

We tested the hypothesis that bronchial epithelium of newborn and fetal pigs exerts an inhibitory effect on bronchial contractile function. In vitro studies were performed on 13 isolated bronchial segments from seven neonatal pigs (12 h to 4 d old) and on 9 bronchial segments from five late-term pig fetuses (105 d gestation, term = 115 d). This preparation keeps the lumen of the airway physiologically intact and separate from the serosal aspect. Stem bronchi of the right and left lung were mounted horizontally at 6 cm H2O in a chamber containing oxygenated Kreb's solution. One bronchus was intact while the other had the epithelium removed with a cotton-tipped applicator soaked in Kreb's solution. Removal of epithelium was confirmed histologically. The lumen pressure of the mounted bronchi was measured at constant volume after the luminal application of acetylcholine (ACh) or K2SO4 (80 mM/L in place of NaCl), after serosal applications of these two substances at the peak of the luminal response, and after serosal application of ACh alone (newborn only). ACh (1, 10, and 100 microM) in the lumen of intact airways of neonatal pigs caused contractions of 0.1 +/- 0.07 (SEM), 0.4 +/- 0.14, and 0.9 +/- 0.18 cm H2O, respectively; in airways denuded of epithelium, ACh caused contractions of 3.9 +/- 1.1, 15.5 +/- 2.94, and 2.95 +/- 4.97 cm H2O (p < 0.001 versus intact airways). Luminal contractions were 2.9 +/- 2.2%, 2.6 +/- 0.9%, and 2.8 +/- 0.7% of the luminal+serosal values for intact segments, as compared with 65.5 +/- 4.6%, 58.8 +/- 7.7%, and 75.1 +/- 6.2% for denuded segments (p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

The integrity of the epithelium is a major determinant of the responsiveness of the dog bronchial segment to mucosal provocation.

The intact bronchial segment of the dog was used to investigate the role of the epithelium in modulating bronchial responsiveness: firstly, because it retains many of the properties of the airway wall including the separation of the mucosal and adventitial surfaces; and secondly, because airway tissue from dogs has been principally used in advancing evidence for epithelial relaxant substances. The development of tone by the airway smooth muscle was measured as an increase in luminal pressure of the isovolumic segment. Cholinergic excitatory and sympathetic inhibitory responses were obtained to electrical field stimulation. Responses to acetylcholine (ACh), carbachol, depolarizing K+ solution and vanadate (VO3-) applied to the mucosal surface were greatly attenuated both in sensitivity and reactivity compared with their effect on the adventitial surface. Mechanical removal of the epithelium greatly augmented these responses to agents in the lumen (ACh sensitivity increased 100-fold) to equal those by adventitial stimulation. The latter were unaffected by epithelial removal. No functional evidence for epithelial-derived inhibitory factors could be demonstrated. It is concluded that the impermeable nature of the epithelium acts as a barrier to diffusion of hydrophilic and charged molecules thereby producing large concentration gradients across the airway wall. A breach in the integrity of the epithelium enables molecules to gain access to the smooth muscle.