Respiratory Tract Of Guinea Pigs Research Articles

Influenza C and D Viruses Demonstrated a Differential Respiratory Tissue Tropism in a Comparative Pathogenesis Study in Guinea Pigs.

Influenza C virus (ICV) is increasingly associated with community-acquired pneumonia (CAP) in children and its disease severity is worse than the influenza B virus, but similar to influenza A virus associated CAP. Despite the ubiquitous infection landscape of ICV in humans, little is known about its replication and pathobiology in animals. The goal of this study was to understand the replication kinetics, tissue tropism, and pathogenesis of human ICV (huICV) in comparison to the swine influenza D virus (swIDV) in guinea pigs. Intranasal inoculation of both viruses did not cause clinical signs, however, the infected animals shed virus in nasal washes. The huICV replicated in the nasal turbinates, soft palate, and trachea but not in the lungs while swIDV replicated in all four tissues. A comparative analysis of tropism and pathogenesis of these two related seven-segmented influenza viruses revealed that swIDV-infected animals exhibited broad tissue tropism with an increased rate of shedding on 3, 5, and 7 dpi and high viral loads in the lungs compared to huICV. Seroconversion occurred late in the huICV group at 14 dpi, while swIDV-infected animals seroconverted at 7 dpi. Guinea pigs infected with huICV exhibited mild to moderate inflammatory changes in the epithelium of the soft palate and trachea, along with mucosal damage and multifocal alveolitis in the lungs. In summary, the replication kinetics and pathobiological characteristics of ICV in guinea pigs agree with the clinical manifestation of ICV infection in humans, and hence guinea pigs could be used to study these distantly related influenza viruses. IMPORTANCE Similar to influenza A and B, ICV infections are seen associated with bacterial and viral co-infections which complicates the assessment of its real clinical significance. Further, the antivirals against influenza A and B viruses are ineffective against ICV which mandates the need to study the pathobiological aspects of this virus. Here we demonstrated that the respiratory tract of guinea pigs possesses specific viral receptors for ICV. We also compared the replication kinetics and pathogenesis of huICV and swIDV, as these viruses share 50% sequence identity. The tissue tropism and pathology associated with huICV in guinea pigs are analogous to the mild respiratory disease caused by ICV in humans, thereby demonstrating the suitability of guinea pigs to study ICV. Our comparative analysis revealed that huICV and swIDV replicated differentially in the guinea pigs suggesting that the type-specific genetic differences can result in the disparity of the viral shedding and tissue tropism.

Lowest observed adverse effect level of pulmonary pathological alterations due to nitrous acid exposure in guinea pigs

BackgroundWe previously demonstrated that continuous exposure to nitrous acid gas (HONO) for 4 weeks, at a concentration of 3.6 parts per million (ppm), induced pulmonary emphysema-like alterations in guinea pigs. In addition, we found that HONO affected asthma symptoms, based on the measurement of respiratory function in rats exposed to 5.8 ppm HONO. This study aimed to investigate the dose-response effects of HONO exposure on the histopathological alterations in the respiratory tract of guinea pigs to determine the lowest observed adverse effect level (LOAEL) of HONO.MethodsWe continuously exposed male Hartley guinea pigs (n = 5) to four different concentrations of HONO (0.0, 0.1, 0.4, and 1.7 ppm) for 4 weeks (24 h/day). We performed histopathological analysis by observing lung tissue samples. We examined samples from three guinea pigs in each group under a light microscope and measured the alveolar mean linear intercept (Lm) and the thickness of the bronchial smooth muscle layer. We further examined samples from two guinea pigs in each group under a scanning electron microscope (SEM) and a transmission electron microscope (TEM).ResultsWe observed the following dose-dependent changes: pulmonary emphysema-like alterations in the centriacinar regions of alveolar ducts, significant increase in Lm in the 1.7 ppm HONO-exposure group, tendency for hyperplasia and pseudostratification of bronchial epithelial cells, and extension of the bronchial epithelial cells and smooth muscle cells in the alveolar duct regions.ConclusionsThese histopathological findings suggest that the LOAEL of HONO is < 0.1 ppm.

Replication and Transmission of the Novel Bovine Influenza D Virus in a Guinea Pig Model.

Influenza D virus (FLUDV) is a novel influenza virus that infects cattle and swine. The goal of this study was to investigate the replication and transmission of bovine FLUDV in guinea pigs. Following direct intranasal inoculation of animals, the virus was detected in nasal washes of infected animals during the first 7 days postinfection. High viral titers were obtained from nasal turbinates and lung tissues of directly inoculated animals. Further, bovine FLUDV was able to transmit from the infected guinea pigs to sentinel animals by means of contact and not by aerosol dissemination under the experimental conditions tested in this study. Despite exhibiting no clinical signs, infected guinea pigs developed seroconversion and the viral antigen was detected in lungs of animals by immunohistochemistry. The observation that bovine FLUDV replicated in the respiratory tract of guinea pigs was similar to observations described previously in studies of gnotobiotic calves and pigs experimentally infected with bovine FLUDV but different from those described previously in experimental infections in ferrets and swine with a swine FLUDV, which supported virus replication only in the upper respiratory tract and not in the lower respiratory tract, including lung. Our study established that guinea pigs could be used as an animal model for studying this newly emerging influenza virus. Influenza D virus (FLUDV) is a novel emerging pathogen with bovine as its primary host. The epidemiology and pathogenicity of the virus are not yet known. FLUDV also spreads to swine, and the presence of FLUDV-specific antibodies in humans could indicate that there is a potential for zoonosis. Our results showed that bovine FLUDV replicated in the nasal turbinate and lungs of guinea pigs at high titers and was also able to transmit from an infected animal to sentinel animals by contact. The fact that bovine FLUDV replicated productively in both the upper and lower respiratory tracts of guinea pigs, similarly to virus infection in its native host, demonstrates that guinea pigs would be a suitable model host to study the replication and transmission potential of bovine FLUDV.

Novel H7N9 influenza virus shows low infectious dose, high growth rate, and efficient contact transmission in the guinea pig model.

The zoonotic outbreak of H7N9 subtype avian influenza virus that occurred in eastern China in the spring of 2013 resulted in 135 confirmed human cases, 44 of which were lethal. Sequencing of the viral genome revealed a number of molecular signatures associated with virulence or transmission in mammals. We report here that, in the guinea pig model, a human isolate of novel H7N9 influenza virus, A/Anhui/1/2013 (An/13), is highly dissimilar to an H7N1 avian isolate and instead behaves similarly to a human seasonal strain in several respects. An/13 was found to have a low 50% infectious dose, grow to high titers in the upper respiratory tract, and transmit efficiently among cocaged guinea pigs. The pH of fusion of the hemagglutinin (HA) and the binding of virus to fixed guinea pig tissues were also examined. The An/13 HA displayed a relatively elevated pH of fusion characteristic of many avian strains, and An/13 resembled avian viruses in terms of attachment to tissues. One important difference was seen between An/13 and both the H3N2 human and the H7N1 avian viruses: when inoculated intranasally at a high dose, only the An/13 virus led to productive infection of the lower respiratory tract of guinea pigs. In sum, An/13 was found to retain fusion and attachment properties of an avian influenza virus but displayed robust growth and contact transmission in the guinea pig model atypical of avian strains and indicative of mammalian adaptation.

Neurotrophin and GDNF family ligand receptor expression in vagal sensory nerve subtypes innervating the adult guinea pig respiratory tract

We combined retrograde tracing techniques with single-neuron RT-PCR to compare the expression of neurotrophic factor receptors in nodose vs. jugular vagal sensory neurons. The neurons were further categorized based on location of their terminals (tracheal or lungs) and based on expression of the ionotropic capsaicin receptor TRPV1. Consistent with functional studies, nearly all jugular neurons innervating the trachea and lungs expressed TRPV1. With respect to the neurotrophin receptors, the TRPV1-expressing jugular C-fiber neurons innervating both the trachea and lung compartments preferentially expressed tropomyosin-receptor kinase A (TrkA), with only a minority of neurons expressing TrkB or TrkC. The nodose neurons that express TRPV1 (presumed nodose C-fibers) innervate mainly intrapulmonary structures. These neurons preferentially expressed TrkB, with only a minority expressing TrkA or TrkC. The expression pattern in tracheal TRPV1-negative neurons, nodose tracheal presumed Aδ-fiber neurons as well as the intrapulmonary TRPV1-negative presumed Aβ-fiber neurons, was similar to that observed in the nodose C-fiber neurons. We also evaluated the expression of GFRα receptors and RET (receptors for the GDNF family ligands). Virtually all vagal sensory neurons innervating the respiratory tract expressed RET and GFRα1. The jugular neurons also categorically expressed GFRα3, as well as ∼50% of the nodose neurons. GFRα2 was expressed in ∼50% of the neurons irrespective of subtype. The results reveal that Trk receptor expression in vagal afferent neurons innervating the adult respiratory tract depends more on the location of the cell bodies (jugular vs. nodose ganglion) than either the location of the terminals or the functional phenotype of the nerve. The data also reveal that in addition to neurotrophins, the GDNF family ligands may be important neuromodulators of vagal afferent nerves innervating the adult respiratory tract.

Effects of nitrous acid exposure on pulmonary tissues in guinea pigs

Many epidemiological studies on the effects of nitrogen dioxide (NO2) on respiratory function may have included nitrous acid (HONO) exposures in their measures, because conventional NO2 assays detect HONO as NO2. A few epidemiological studies and human HONO inhalation experiments have associated HONO with decrements in lung functions. However, there have been few HONO exposure experiments in animals. This study aims to develop a HONO generation system for the animal exposure experiments, and to assess the association of HONO exposure with histopathologic alterations in the respiratory tract of guinea pigs. We exposed the guinea pigs to 3.6 ppm HONO with secondary products of 0.3 ppm NO2 and 1.6 ppm nitric oxide (NO) for 4 weeks (24 h/day). We conducted histopathologic analyses and measured specific airway resistance (sRaw) from 7 h 40 min to 8 h 30 min after the end of HONO exposure. We found pulmonary emphysema-like alterations in the alveolar duct centriacinar regions, distortion of the centriacinar regions of alveolar ducts with extension of the bronchial epithelial cells and smooth muscle cells, and expansion of bronchial epithelial cells, in the HONO exposure. These histopathologic results suggest that a high concentration of HONO with some NO2 and NO may associate with decrements in lung functions and some respiratory symptoms. Although the increased tendency of the sRaw value was observed in the HONO exposure group, no statistically significant difference was found between the sRaw values from the HONO exposure group and the filtered air group (p = 0.06, student’s t-test).

Effects of some Malian medicinal plants on the respiratory tract of guinea-pigs.

Crossopteryx febrifuga, Pteleopsis suberosa and Entada africana are used in Mali traditional medicine for fever and various respiratory diseases. We have investigated the effects of these three drugs in the form of a decoction on the respiratory tract using different experimental models. On citric acid-induced cough in guinea-pigs, the three drugs significantly decreased the number of coughs at the doses of 250 (P < 0.01), 500 (P < 0.05; P < 0.01) and 1000 (P < 0.01) mg kg(-1). The percent inhibition was respectively 62.86, 69.03 and 77.44% for C. febrifuga, 57.80, 53.90 and 61.40% for E. africana, and 37.13, 42.44 and 73.72% for P. suberosa. Codeine phosphate (10 mg kg(-1)) used as reference drug showed an inhibition of 76.32%. E. africana (1000 mg kg(-1)) reduced (65% inhibition) significantly (P < 0.05) bronchoconstriction induced by histamine (99.25% and 34.00% for control and extract, respectively). Furthermore, E. africana (1000 mg kg(-1)) provoked a bronchodilatation response when administered under basal conditions. On antigen-induced bronchospasm, C. febrifuga protected (54% inhibition) sensitized guinea-pigs with a pulmonary ventilation pressure (PVP) of 24.87% (control value < 55.00%). P. suberosa was inactive in both experimental models. The reference drug, disodium cromoglycate (10 mg kg(-1), i.v.) protected significantly (P < 0.05) with a PVP of 12.00% (78% of inhibition). This study confirmed the traditional use of these plants in the treatment of cough and other respiratory disorders.

Histopathology and growth kinetics of influenza viruses (H1N1 and H3N2) in the upper and lower airways of guinea pigs

Recent investigations have shown that guinea pigs are important for the study of influenza A virus (IAV) transmission. However, very little is known about IAV replication and histopathology in the guinea pig respiratory tract. Here, we describe viral growth kinetics, target cells and histopathology in the nasosinus, trachea and lungs of IAV-infected guinea pigs. We found that guinea pigs infected with either A/Puerto Rico/8/34 (H1N1) or A/Hong Kong/8/68 (H3N2) developed a predominantly upper airway infection with high nasal viral titres. IAV grew to moderate titres in the lungs but induced marked inflammatory responses, resulting in severe bronchopneumonia and alveolitis. Although non-lethal at the high dose of 2x10(6) p.f.u., infections with these IAV strains were associated with reduced weight gain. IAV infection in guinea pigs is characterized by extensive viral replication in the ciliated nasal epithelial cells followed by heavy nasal mucus secretion.

Differential Phenotype of Placodal vs Neural Crest Derived Afferent C‐fibers in Airways and Esophagus.

C‐fibers in the guinea pig respiratory tract comprise two distinct phenotypes, depending on whether the cell body is situated in the nodose (placodal origin) ganglion or jugular (neural crest origin) ganglion (Undem et al. J.Physiol, 2004). Both types of C‐fibers are responsive to capsaicin but only the nodose C‐fibers are responsive to purinergic P2X receptor, 5‐HT‐3 receptor, and adenosine A1 receptor agonists. The vast majority of jugular, but not nodose ganglia express neurokinins. We speculate that the C‐fiber phenotype may be determined more by their embryonic source than from molecular cues arising from the tissue they innervate. To further evaluate this, we compared lung and esophageal C‐fiber neurons arising from dorsal root ganglion (DRG), nodose, and jugular ganglia, based on the knowledge that the DRG neurons innervating these tissues are derived from the same embryonic source as the jugular neurons. Neurons labeled from the lungs and studied with single cell RT‐PCR, histology, and whole cell patch clamp recordings, revealed that DRG and Jugular C‐fiber phenotype are equivalent, but quite distinct from the nodose C‐fibers. A similar distinction was observed in esophageal‐specific afferent neurons. The results indicate that the C‐fiber phenotypes innervating the lungs and esophagus are determined more by their embryonic environment than by the environment of the tissue they ultimately innervate.

Formulation and Evaluation of Insulin Dry Powder for Inhalation

Purpose. Dry powder formulation of insulin for pulmonary administration was prepared to obtain increased drug deposition in the alveolar absorptive region. The deposition was studied by investigating the dispersion and deaggregation of insulin from the carrier lactose using an Andersen cascade impactor and twin stage impinger. The subsequent absorption following the deposition was studied by in vivo method. Methods. Insulin in solution with absorption promoters was lyophilized. The powder was incorporated with lactose of different grades and their combinations as carriers to deliver using an inhaler device. Solid-state characteristics of the carrier as well as the drug powder were assessed by particle size and distribution measurement. The flow properties such as moisture content, powder density, angle of repose, and carr's compressibility index of the powder mixture were determined. The aerosol behavior of the powder was studied by dispersion using rotahaler© connected to a twin-stage impinger (TSI) and an eight-stage Andersen cascade impactor (ACI) operating at different flow rates of 30–90 l/min. The in vivo performance was studied by deliverance to the respiratory tract of guinea pigs. The intratracheal bioavailability with respective to intravenous route was calculated by measuring the blood glucose reduction. Results. The coarser particles of lactose in fractions of carrier containing a wide particle size distribution impacted in the preseperator of cascade impactor, and only the particle less than 10 µm size entered stage 0–stage 7. Formulation containing 1:1 mixture of Respitose ML006 (62%<50 µm) and Respitose ML003 (37.8%<50 µm) as carrier imparts well deaggregation of insulin, and higher deposition leads to 52.3% of fine particle fraction at 60 Lit/min and in vivo bioavailability of 82%. Conclusions. Insulin formulations containing 1:1 mixture of Respitose ML006 and Respitose ML003 as carrier can impart deeper deposition of drug particles and cause higher bioavailability. This suggests that carrier used in the formulation influenced the amount of insulin deposition in the alveolar region of the lung. Hence, it was concluded that the availability of insulin for systemic absorption depends on the particle size of the drug as well as the carrier lactose.

Cross-complementation between the products of the genes P1 and ORF6 of Mycoplasma pneumoniae subtypes 1 and 2

The genes P1 (MPN141) and ORF6 (MPN142) are essential for the successful colonization of the human respiratory tract by Mycoplasma pneumoniae. Both genes are located in the P1 operon, which consists of three genes. The P1 gene is the second gene in the operon, followed by the ORF6 gene. The P1 gene contains two (RepMP2/3, RepMP4) and the ORF6 gene one (RepMP5) specific repetitive DNA sequence, of which seven to nine similar but not identical copies are dispersed on the genome. Despite this large potential pool for genetic variation, M. pneumoniae isolates from patients contain only one of two distinct combinations of the genes P1 and ORF6. To analyse the functions of the repetitive DNA sequences, two 'new' combinations of the genes P1 and ORF6 were constructed, keeping the P1 gene constant but exchanging RepMP5 copies of the ORF6 gene. M. pneumoniae was transformed with these constructs and the transformants were tested for their ability to grow and survive under in vitro conditions and in guinea pigs. The two transformants colonized the respiratory tract of guinea pigs and showed no obvious differences in their growth behaviour compared to M. pneumoniae isolates from patients. The results indicate that the subtype-specific combinations of the repetitive elements in the P1 and ORF6 genes are not essential for the successful adherence of M. pneumoniae to host cells and the colonization of the respiratory tract of guinea pigs.

Improvement of therapeutic index of phosphodiesterase type IV inhibitors as anti-Asthmatics.

A new series of catechol hydrazines was synthesized and their structure-activity relationship (SAR) was analyzed for developing an effective phosphodiesterase 4 (PDE4) inhibitor as an anti-asthmatic drug candidate. Among the (E)-Analogues tested using in vitro assays, 5CC showed a strong PDE4 inhibitory activity and a significantly improved rolipram binding profile compared with rolipram, a prototype PDE4 inhibitor. Moreover, from in-vivo asthma model, we observed that (E)-Analogue 5CC had a good efficacy against guinea-pig respiratory tract inflammation and bronchoconstriction, along with a remarkably reduced emetic side effect, compared with rolipram. Conclusively, (E)-Analogue 5CC seems to be a promising candidate for the development of anti-asthmatic PDE4 inhibitors.

COMBINED RESPIRATORY EFFECTS OF COLD AIR WITH SO2 OR NO2 IN SINGLE 1-HOUR EXPOSURES OF HYPERVENTILATING GUINEA PIGS

The present study is a continuation of our previous experiments on repeated 10-min exposures of anesthetized, mechanically ventilated guinea pigs to clean cold dry air (Hälinen et al., 2000a), and to cold air plus gaseous air pollutants (Hälinen et al., 2000b). This time we made continuous 60-min exposures to clean cold dry air, cold air + SO2 at 1 ppm, cold air + NO2 at 1 ppm, and warm humid air + NO2 at 1 ppm, and focused on responses at 10-60 min. Clean cold dry air and cold air + pollutants (n = 8-9 in each group) produced similar cooling in the guinea pig lower respiratory tract. The decreases in intratracheal temperature (Ttr) reached a plateau at 20 min with mean maximal decreases of 9.7-11.3°C from the pre-exposure control values of 36.0-37.3°C. In contrast, there were progressive decreases in esophageal temperature (Toe) during the exposures, indicating constant conductive and evaporative heat losses from the tracheobronchial tissues. The mean maximal decreases in Toe were 1.2-1.4°C from the preexposure control values of 37.8-38.0°C. Clean cold dry air induced 4.5-10.8% mean decreases in peak expiratory flow (PEF) at 10-60 min of exposure, which were statistically nonsignificant due to a relatively large variation between animals. Cold air + SO2 at 1 ppm induced a mean decrease of 11.4% in PEF at 10 min (p < .05), which was spontaneously abolished during the next 10 min of exposure. Cold air + NO2 at 1 ppm caused no decrease, but in fact small, nonsignificant increases in PEF at 30-60 min of exposure. Cold air + NO2 at 1 ppm, and to some extent also cold air + SO2 at 1 ppm, attenuated significantly the mechanical ventilation induced gradual decrease in tidal volume (VT), when compared to clean cold dry air exposure. Cold air + NO2 at 1 ppm, but not warm humid air + NO2 at 1 ppm, increased significantly the proportion of macrophages in the differential count of bronchoalveolar lavage fluid (BALF) white cells when compared to both clean warm humid air and cold dry air. None of the exposure conditions caused morphological or inflammatory changes in the respiratory tissues. In conclusion, continuous 60-min exposures to clean cold dry air, cold air + SO2, and cold air + NO2 produced weaker functional effects on the lower respiratory tract of guinea pigs than our previous consecutive 10-min exposures to these air conditions. After the first 10 min, there was a strong attenuation of the bronchoconstrictor responses, especially to cold air + NO2 at 1 ppm. The small airway effects of prolonged mechanical ventilation were significantly modified by NO2 at 1 ppm in both cold dry and warm humid breathing air. Finally, cold air + NO2 at 1 ppm increased the proportion of macrophages in BALF white cells.

IMMUNOHISTOCHEMICAL DETECTION OF TOLUENE DIISOCYANATE (TDI) ADDUCTS IN PULMONARY TISSUE OF GUINEA PIGS FOLLOWING INHALATION EXPOSURE

Toluene diisocyanate (TDI) is currently the most prevalent cause of occupational asthma. Inhalation of vapors of the reactive chemical may result in irritation and inflammation of the respiratory tract as well as in pulmonary sensitization. Although numerous studies have characterized the pulmonary inflammation, the mechanism underlying sensitization remains controversial. This investigation sought to address the problem by following the localization of TDI in the respiratory tract of guinea pigs using a well-characterized animal model that displays both immunologic and respiratory hypersensitivity to TDI. Nonanesthetized and minimally restrained guinea pigs were exposed to a sensitizing regimen of TDI that consisted of inhalation of 1.0 ppm TDI vapor for 3 h/day on 5 consecutive days. Pulmonary inflammation was present in animals sacrificed immediately following the exposure as evidenced histologically, and by increased protein and inflammatory cells in the bronchoalveolar lavage. Using a specific and sensitive rabbit antiserum prepared to TDI-keyhole limpet hemocyanin, the presence of TDI adducts in respiratory tissues was investigated. TDI adducts were detected primarily in the apical region of the epithelium of the nares, trachea, bronchi, and bronchioles. The amount of adduct was greatly reduced in animals sacrificed 18 days later. Sensitized guinea pigs that were challenged on day 23 with 80-250 ppb TDI demonstrated the same distribution of TDI adducts as did shamexposed animals similarly challenged. These results are the first to employ immunohistochemistry to detect TDI adducts in respiratory tract tissue. Findings indicate that one exposure to the chemical results in detectable adducts, and that the location of adducts is dependent upon both the concentration of TDI inhaled and the number of exposures. Isolation and characterization of the cellular adducts should assist in elucidation of the role of adducts in the sensitization process.

CILIARY ULTRASTRUCTURE AND BEATING ACTIVITY IN RAT AND GUINEA‐PIG RESPIRATORY MUCOSA

1. The rat and the guinea-pig are commonly used animals when the effects of drugs on ciliary activity in respiratory airways are studied. There are few data concerning the possible differences in ciliary function between these two animals. 2. Using a photodetector method we measured the ciliary beating frequency (CBF) from the upper part of the trachea, the lower part of the trachea and the distal part of the main bronchi (subsegmental bronchi) of rat and guinea-pig respiratory tract. In addition, the structure of the cilia was studied using scanning electron microscopy (SEM). 3. CBF in the rat respiratory tract was significantly lower than in the guinea-pigs. In the upper trachea, the CBF for rat was 12.7 beats/s and 15.3 beats/s for guinea-pig. The respective values were 9.2/16.0 beats/s in the lower part of the trachea and 6.9/13.8 beats/s in subsegmental bronchi. In both rats and guinea-pigs CBF was lower in the subsegmental bronchi than in the trachea (rat: 25.0-45.7%, guinea-pig: 9.8-13.8%). 4. In addition to higher CBF, the quality of the photo-electrical signal was better from guinea-pig tissues, probably as a result of the larger amounts of ciliated cells and longer cilia of guinea-pig respiratory epithelial compared to those in rat mucosa. 5. SEM showed that the rat cilia were on average shorter (3.6 vs 4.3 microns) and thinner (0.19 vs 0.22 microns) than those of the guinea-pig. Rat mucosa was markedly less ciliated than the respiratory mucosa of the guinea-pig.(ABSTRACT TRUNCATED AT 250 WORDS)

Autoradiographic localization of calcitonin gene-related peptide receptors in guinea pig respiratory tract: effect of capsaicin pretreatment.

Calcitonin gene-related peptide (CGRP) is a potent vasodilator peptide present in capsaicin-sensitive neurons innervating the respiratory tract. In this study, the autoradiographic distribution of [125I]CGRP binding sites was investigated in guinea pig airways. Extremely dense specific binding occurred over parenchymal tissue, with moderate specific binding over tracheal glands, the endothelium of pulmonary veins and arteries, and small blood vessels in the bronchial wall. The localization of binding sites for [125I]CGRP over blood vessels but not bronchial smooth muscle correlates well with the physiological actions of this peptide, although the function of the parenchymal sites is unknown. No significant difference in binding was seen in vehicle- or capsaicin-pretreated animals, suggesting that sites are not reliant on factors from capsaicin-sensitive neurons.

NADPH-diaphorase activity as a marker for nitric oxide synthase in neurons of the guinea pig respiratory tract.

Recent studies in physiology have suggested that part of the inhibitory nonadrenergic noncholinergic (iN-ANC) response of airway smooth muscle is mediated by nitric oxide (NO). To examine this point morphologically, the guinea pig respiratory tract was investigated histochemically for nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase (NADPH-d), a marker for NO synthase (NOS). In addition, coexpression of NOS and vasoactive intestinal peptide (VIP) or calcitonin gene-related peptide (CGRP) was studied using a combination of histochemistry for NADPH-d and immunohistochemistry for VIP or CGRP. Nerve fibers showing NADPH-d activity were abundantly observed in the respiratory tract. They were distributed throughout smooth-muscle bundles, lamina propria, submucosal glands, and around bronchial and pulmonary arteries. NADPH-d-containing nerve-cell bodies were occasionally found within airway ganglia. The colocalization study demonstrated that NADPH-d-containing nerve fibers frequently coincided with VIP-like immunoreactive nerve fibers but not with CGRP-like immunoreactive nerve fibers. Among nonneural tissues, NADPH-d activity was noticed in the endothelium of both bronchial and pulmonary vessels, and in the pleura. These observations indicated that NO may be produced by neurons and vascular endothelium of the guinea pig respiratory tract, and may function as a neuronal mediator as well as endothelium-derived relaxing factor (EDRF). Colocalization of NADPH-d and VIP-like immunoreactivity in nerve fibers suggested that NO and VIP may function as cotransmitters.

Modulation of neural bronchoconstrictor responses in the guinea pig respiratory tract by vasoactive intestinal peptide

Vasoactive intestinal peptide (VIP) is localised to cholinergic nerves in airways. We have investigated the effects of VIP on both cholinergic and non-adrenergic, non-cholinergic (NANC) neuronal bronchoconstrictor responses to electrical field stimulation (EFS) in guinea pig airways and on cholinergic neurotransmission following sensory nerve depletion. VIP significantly attenuated the cholinergic bronchoconstrictor responses to EFS in trachea (EC 50 values in upper and lower trachea of 3.7 ± 0.2 × 10 −9 M and 8.6 ± 0.3 × 10 −9 M, respectively) and bronchi (31.2 ± 1.6% inhibition in main and 15.1 ± 3.3% in hilar bronchi at 10 −7 M VIP) and the NANC bronchoconstrictor responses to EFS in bronchi (with maximum inhibitions of 93.1 ± 1.8% at 3 × 10 −8 M VIP in main and 40.2 ± 5.3% at 10 −8 M in hilar bronchi). VIP at 10 −7 M, but not at 10 −10 M, significantly attenuated the contractile responses to exogenously applied ACh in trachea (EC 50 values of 4.9 ± 0.2 × 10 −6 M in the absence and 8.4 ± 0.4 × 10 −5 M in the presence of VIP 10 −7 M VIP) to SP in main bronchi (EC 50 values of 5.7 ± 0.2 × 10 −8 M in the absence vs. 7.3 ± 0.3 × 10 −7 M in the presence of 10 −7 M VIP). Since the inhibition of these neural responses is greater than the inhibition of the equivalent responses elicited by the exogenous transmitters, this indicates that VIP may modulate release of acetylcholine from cholinergic nerves and of neuropeptides from sensory nerves, in addition to a post-junctional functional antagonist action.

Parainfluenza-3 infection of the guinea pig airways induces respiratory airway hyperreactivityin vitro

Guinea pigs were inoculated intra-tracheally with bovine parainfluenza type 3 virus (PI-3) to investigate airway responsiveness to histamine and arecoline in vitro. Two days after saline or PI-3 inoculation no difference in the reactivity of the tracheal spirals was observed after cumulative concentration/response curves with both drugs. Similarly, contractions of parenchymal strips, induced by histamine, did not differ between both groups. However, 4 days after PI-3 inoculation the histamine induced contraction of the tracheal spirals was increased by 47% and the arecoline induced contractions by 32% as compared to the control group. Further, the contractions induced by histamine in parenchymal strips were significantly enhanced (p less than 0.01) in the PI-3 treated group. In conclusion, PI-3 infection of guinea pig respiratory tract induces hyperreactivity of the central and peripheral airways in vitro.

Regional and time-dependent effects of inflammatory mediators on airway microvascular permeability in the guinea pig.

1. Airway oedema resulting from increased microvascular permeability is a characteristic pathological finding of asthma. The regional effects of putative mediators involved in asthma on airway microvascular permeability have been studied. 2. The effects of histamine, leukotriene (LT) D4 and platelet-activating factor (PAF) on microvascular permeability in the nasal mucosa, larynx, trachea, main bronchi and intrapulmonary airways of the guinea pig were assessed by measuring the extravasation of intravenously administered Evans Blue dye. 3. PAF and LTD4 caused increased microvascular leakage throughout the respiratory tract, although their effects were maximal in different regions. Histamine had no significant effect on intrapulmonary airways. PAF was more potent than LTD4 and histamine at all airway levels. For example, in the trachea the doses required to cause leakage of 50% of maximal (ED50) were 10.4 nmol/kg, 138 nmol/kg and 11.2 mumol/kg, respectively, for PAF, LTD4 and histamine. 4. The effect of the three mediators was maximal 5 min after intravenous administration. Histamine, but neither LTD4 nor PAF, still caused significant leakage 30 min after administration. 5. The increased microvascular leakage induced by the mediators was inhibited by their respective specific receptor antagonists, suggesting that the effect was mediated via specific receptors. 6. Histamine, LTD4 and PAF have varying potencies in increasing microvascular permeability in the guinea-pig respiratory tract, exert their maximal effect in different regions and have varying durations of action.