RNA metabolism in tracheal epithelium: alteration in hamsters deficient in vitamin A.

The electrophoretic pattern of RNA molecules that are synthesized in vitro in tracheal epithelium from hamsters deficient in vitamin A differs from that of RNA synthesized in normal, pair-fed control hamsters. There is less RNA of low electrophoretic mobility in the epithelial cells deficient in vitamin A. This alteration is reversed after the deficient animals have been treated with vitamin A.

Purification and reconstitution of chloride channels from kidney and trachea.

Chloride channels mediate absorption and secretion of fluid in epithelia, and the regulation of these channels is now known to be defective in cystic fibrosis. Indanyl-oxyacetic acid 94 (IAA-94) is a high-affinity ligand for the chloride channel, and an affinity resin based on that structure was developed. Solubilized proteins from kidney and trachea membranes were applied to the affinity matrix, and four proteins with apparent molecular masses of 97, 64, 40, and 27 kilodaltons were eluted from the column by excess IAA-94. A potential-dependent 36Cl- uptake was observed after reconstituting these proteins into liposomes. Three types of chloride channels with single-channel conductances of 26, 100, and 400 picosiemens were observed after fusion of these liposomes with planar lipid bilayers. Similar types of chloride channels have been observed in epithelia.

Factors influencing the viscous properties of chicken tracheal mucins.

1. Reduced viscosities, in water, of different types of mucin, such as fibrillar, gelatinous and soluble phase, separated from chicken tracheal secretions were measured. 2. H-bond breaking agents caused a significant decrease in the reduced viscosity of these mucins, but thiol-reagents alone did not have any effect. 3. Papain and Pronase did not cause any decrease in the reduced viscosity of these mucins. Neuraminidase decreased the reduced viscosity of soluble phase mucin by 50% by removing about 30% of its N-acetylneuraminic acid but had no effect on fibrillar and gelatinous mucins. Sulphatase neither removed any sulphate ester groups nor decreased the reduced viscosity. Due to some nonspecific intermolecular interaction, mixtures of mucins and enzymes or ovalbumin exhibited elevated reduced viscosities. 4. Ionic strength of the solutions appeared to decrease the reduced viscosity of these mucins. Increasing concentrations of Ca2+ in solutions of ionic strength of approx. 0.1 caused significant decrease in the reduced viscosity, but had no such effect in solutions of ionic strength of more than 0.1. 5. N-Acetylneuraminic acid and sulphate ester residues were 46.6 +/- 0.2, 43.4 +/- 0.6, 27.9 +/- 3.3 mg/g and 66.0 +/- 2.0, 34.2 +/- 3.3, 2.5 +/- 0.8 mg/g for fibrillar, gelatinous and soluble phase mucins, respectively. There appeared to be a good correlation between viscosity and N-acetylneuraminic acid contents among mucins of low reduced viscosities and between viscosity and sulphate ester residues among mucins of high reduced viscosities.

Isolation and characterization of glycoproteins from canine tracheal mucus.

Three homogeneous glycoproteins were isolated from reduced and S-carboxy-methylated canine tracheal pouch mucus by gel filtration and ion-exchange chromatography. Initial fractionation was carried out on Sephadex G-200; chromatography of the excluded Sephadex G-200 fraction on Bio-Gel A-15 m yielded two high molecular weight glycoprotein fractions. Following rechromatography on the same column, the main fraction behaved as an electrophoretically homogeneous high molecular weight (581 600) glycoprotein, with a high carbohydrate content (80%) and a single amino-terminal amino acid (arginine). Ion-exchange chromatography (DEAE-cellulose) of the included Sephadex G-200 fraction yielded two electrophoretically homogeneous glycoproteins of lower molecular weight (20 800 and 24 600, respectively). A single amino-terminal amino acid, glycine and alanine, respectively, was detected for each glycoprotein. Chemical analysis of these three glycoproteins revealed the presence of fucose, galactose, N-acetylgalactosamine, N-acetylglucosamine, N-acetylneuraminic acid and sulfate monoester. The high molecular weight glycoprotein had a higher hexose, sialic acid and sulfate content, per mg of protein, than the low molecular weight glycoproteins. The results of the alkaline borohydride treatment indicated that the majority of the carbohydrate chains of these glycoproteins are linked to the protein core through O-glycosidic bonds involving N-acetylgalactosamine and serine or threonine.

Mucus-glycoproteins (mucins) of the cat trachea: characterisation and control of secretion.

Glycoproteins produced by the tracheae of anaesthetized cats were radiolabelled biosynthetically by a pulse administration of Na2 35SO4 and [3H]glucose into the tracheal lumen. Subsequently, radiolabelled secretions were washed from the tracheal lumen. Repeated doses of pilocarpine and then ammonia vapour were given to stimulate secretion. Pilocarpine-stimulated glycoproteins, which came mainly from the submucosal glands, were particularly enriched with 35S. Ammonia-stimulated secretions, which probably came mostly from the microvillous border of the surface epithelium, contained mainly 3H radioactivity but little 35S. Two negatively-charged glycoproteins of different molecular size were identified in the secretions: the larger component was excluded on Sepharose CL-4B and it had a higher 3H 35S ratio than the smaller component which was retarded on Sepharose CL-4B. The relative amount of the smaller component decreased progressively with repeated pilocarpine stimulation and it was not detected in secretions induced by ammonia. Pilocarpine stimulation caused little alteration in carbohydrate composition of the secreted glycoproteins. In response to ammonia, glycoproteins were secreted with a high sialic acid content but quantitatively they represented a small amount of material compared with that induced by pilocarpine. These findings suggest that tracheal glycoproteins from different epithelial-cell sources have distinctive chemical compositions and that their secretions may be independently regulated. The 35S-rich high-molecular-weight glycoproteins from the submucosal glands were of the mucin-type but those derived from the microvillus border may represent a different class of airway glycoproteins from typical epithelial mucins.

Somatomedin activity in tracheal fluid from the newborn infant.

Somatomedin activity was determined by porcine cartilage bioassay and somatomedin-C RIA in matched samples of tracheal fluid (TF), amniotic fluid (AF), and cord (CP) and maternal plasma collected during the delivery of normal infants of 36-40 weeks gestation. Somatomedin levels determined by both assay systems were significantly greater in TF than in AF [TF, 0.40 +/- 0.04 (+/- SE) U/ml; AF, 0.20 +/- 0.02 U/ml by bioassay in 11 infants; TF, 0.25 +/- 0.04 U/ml; AF, 0.18 +/- 0.03 U/ml, by RIA in 6 infants; P less than 0.005]. When chromatographed on Bio-Gel P10 at pH 3.5, the major part of the somatomedin immunoreactivity from both TF and CP eluted in the same position as iodinated somatomedin-C (mol wt, 7600). Somatomedin activity recovered after chromatography of TF and CP increased the incorporation of radiothymidine by growth-restricted human fetal fibroblasts in vitro. Maternal plasma contained 2.49 U/ml radioimmunoassayable somatomedin-C, a value higher than that in the nonpregnant adult. The results suggest that the human fetal lung contributes somatomedin to TF and raises the possibility that these peptides are involved in lung growth or maturation.

Changes in myosin heavy chain stoichiometry in pig tracheal smooth muscle during development.

The stoichiometry of the myosin heavy chains (MHCs) has been measured in the tracheal smooth muscle of the pig after electrophoresis on SDS 4% polyacrylamide gel. The ratio of slower migrating MHC to the faster migrating MHC was 2.1 neonates, 1.5 in young and 0.95 in old pigs (P less than 0.01) showing that MHC composition changes with development. The unequal proportion of MHCs was not compatible with a heterodimeric arrangement of the MHCs in the native molecule as proposed earlier by Rovner et al. [(1986) Am. J. Physiol. 250, C861-870] and it is suggested that native molecules may be composed of homodimer heavy chains.

Quantitative histochemistry of mucosubstance in tracheal epithelium of the macaque monkey.

Experimentally applied irritants and chronic respiratory diseases appear to alter the amount and composition of secretory cell product in surface epithelium and submucosal glands of pulmonary airways. Previous methods used to quantify these changes have been very time-consuming or have not measured the same components of the airway wall. The present study describes a rapid, reproducible, and standardized automated method for quantifying secretory products. The tracheas from eight macaque monkeys were fixed with glutaraldehyde-paraformaldehyde, embedded in glycol methacrylate, serially sectioned at 2 microns, and histochemically stained to demonstrate neutral, sialylated, and sulfated mucosubstances in the cartilaginous, intercartilaginous, and membranous regions of both proximal and distal trachea. Volume densities were determined using an image analyzer and are expressed as volume of stained mucosubstance per unit surface area of epithelial basal lamina. Comparison of the automated method to manual point counting and evaluation of internal variance showed that the automated method had a twelve-fold increase in efficiency with no significant differences in measurements. After weighting the values of each region according to their anatomical contribution, the total secretory product (TSP) for the entire trachea was determined. Periodate-reactive acid material predominated (73%) in luminal surface epithelium, and neutral material predominated (78%) in submucosal glands. Surface epithelium contained 66% of the TSP. The greater contribution by surface epithelium and predominance of acid mucins there resulted in a TSP from the trachea that consisted of 59% acid material (most of which was sulfated) and 41% neutral material. The method proved to be a valid, reproducible, and rapid technique for evaluating variability in abundance of mucosubstances within airway epithelium.

Histochemical and ultrastructural characterization of serotonin-containing cells in rabbit tracheal epithelium.

Tracheal endocrine cells (TECs) that contain serotonin have been characterized previously by staining with ferric ferricyanide. In the present article, the ferric ferricyanide staining reaction has been used to locate the TECs in deplasticized thick sections of Epon-embedded rabbit tracheas. Adjacent thin sections of the same cell were subsequently observed by electron microscopy. The TECs were filled with dense-core vesicles (DCVs) located in the cytoplasm between the nucleus and the lumen and also lateral to the nucleus. In a separate experiment, pieces of rabbit trachea were treated with a solution of glutaraldehyde-dichromate to demonstrate the presence of amines. High levels of chromium were detected in the DCVs by energy-dispersive X-ray analysis. The results from these studies have correlated the ultrastructure of a serotonin-containing endocrine cell present in rabbit tracheal epithelium with a cell type previously characterized only by light and fluorescence histochemical methods. The results also indicate that serotonin in these cells is stored in the DCVs.

Light microscopic histochemical detection of sugar residues in secretory glycoproteins of rodent and human tracheal glands with lectin-horseradish peroxidase conjugates and the galactose oxidase-Schiff sequence.

Lectins conjugated to horseradish peroxidase were used to stain paraffin sections of mouse, rat, and human respiratory tract tissues. An additional method was applied utilizing galactose oxidase to oxidize the C-6 hydroxyl of galactose and N-acetylgalactosamine residues and the resulting aldehyde was visualized with 2% Schiff's reagent. Sections were stained prior to and after removal of sialic acid residues. Oligosaccharides with terminal beta-galactose and alpha-N-acetylgalactosamine residues were found in all serous cells in the mouse trachea but were never seen in human tracheal serous cells. About 5-10% of serous cells in the rat trachea contained terminal beta-galactose, whereas all human tracheal serous cells and the remainder of those in the rat contained oligosaccharides with terminal sialic acid and penultimate beta-galactose residues. Fucose was not detected in tracheal serous cells of any species. Mucous cells in the mouse and rat trachea produced heterogeneous oligosaccharides containing terminal alpha-N-acetylgalactosamine and/or terminal sialic acid residues in various proportions. The structure of oligosaccharides in human tracheal mucous cells varied between individuals and was related to ABO blood group reactivity. The majority of oligosaccharides in type A individuals contained terminal alpha-N-acetylgalactosamine, whereas type AB individuals had approximately equal amounts of terminal alpha-galactose and alpha-N-acetylgalactosamine residues. Mucous cells in the two type O specimens examined contained a large amount of terminal beta-galactose and surprisingly, terminal fucose was not detected in these individuals. These results support biochemical studies showing structural diversity in oligosaccharide chains of respiratory tract secretions and reveal differences in glycoprotein secretion of different cell types.

Identification of contractile proteins in basal bodies of ciliated tracheal epithelial cells.

To determine the molecular composition of the components of basal bodies and the interbasal body apparatus of ciliated cells in rat tracheal epithelium, we used rabbit anti-actin, anti-alpha-actinin, anti-tropomyosin, and anti-myosin as primary antisera applied to the tissue in an indirect immunoperoxidase technique. The antisera was proven to be monospecific by elution of antibody after affinity chromatography. Sheep anti-rabbit immunoglobulin Fab fragments coupled to peroxidase were used for ultrastructural localization of the bound rabbit antibody. Antibodies against alpha-actinin were demonstrated around peripheral microtubules of cilia and linking these microtubules to central doublet and plasma membrane. Alpha-actinin was also shown in the basal foot processes. Anti-actin antibodies were associated with microtubules of the cilium and basal bodies, except in the region of the ciliary necklace. The antibodies directed against actin also had affinity for rootlets, basal foot processes, and communications between basal bodies and foot processes. Both anti-myosin and anti-tropomyosin antibodies were localized to part of the region of the constriction of the cilium, to the central basal density and the outer surfaces of basal body microtubules, and to the basal foot processes together with their communications to the basal body. The data suggest active contractile function of basal bodies.

Localization of low molecular weight protease inhibitor in serous secretory cells of the respiratory tract.

We prepared in rabbits an antiserum against low molecular weight protease inhibitor (LMI) purified from the sputum of patients with purulent bronchitis. Using this antiserum in an immunoperoxidase staining method we found that this inhibitor was located exclusively in the serous cells of the submucosal glands of human upper and lower airways. The inhibitor was localized also in serous cells of the sublingual and submandibular glands. In contrast, LMI could not be demonstrated in the serous cells of the parotid gland. In the tissues investigated a strong association between the localization of the protease inhibitor and lysozyme was observed. Our observations indicate that the inhibitor may be present together with lysozyme as a secretory product in the serous cell granules. The possible consequences of the coexistence of these two proteins in the defense mechanism of the respiratory tract is discussed.

Immunohistological localization of alpha 1-microglobulin in normal rat tissues.

The tissue distribution of rat alpha 1-microglobulin (alpha 1-m) was studied by indirect immunofluorescence in various rat tissues using a polyvalent rabbit antiserum to the purified antigen and a monoclonal antibody (H23) to the human homologue, in parallel with a polyclonal anti-rat IgA antiserum. It was found that all tissues stained by anti-IgA were also alpha 1-m positive; these tissues included tissues of the stomach, duodenum, ileum, colon, pancreas, trachea, esophagus and jejunum. However, the observation that IgA plasma cells as well as secretory cells, while positively stained by anti-IgA, are alpha 1-m negative suggests that the association between IgA and alpha 1-m occurs at a postsecretory stage, after the IgA molecules have been transported across the epithelial cells. Additionally, hepatocytes were intensely stained by anti-alpha 1-m antibodies, indicating that the liver, as already suggested by metabolic studies on isolated guinea-pig liver explants, may be responsible for the synthesis of this protein. Among lymphoid tissues, an intense and homogeneous staining was observed in the thymus and the white pulp of the spleen. Sections of lymph nodes, however, showed differential staining; apart from a few isolated dendritic cells in the mantle region of the lymphoid follicles, the germinal centers and medullary cords showed no staining with anti-alpha 1-m antibodies. The paracortical cells, macrophages in the subcapsular sinus, and interfollicular lymphocytes showed intense cytoplasmic staining with anti-alpha 1-m antibodies. In other tissues, macrophages, monocytes, tissue histiocytes, and dendritic cells were alpha 1-m positive. Although they confirm the presence of alpha 1-m in the lymphoid tissues, as already reported in man, these results show that the protein is also present in hepatocytes and in exocrine fluids containing IgA. Since alpha 1-m, like secretory component, can bind to IgA to form stable complexes, these two heavily glycosylated proteins may have similar biologic properties.

Surfactant in the lung and tracheal fluid of the fetal lamb and acceleration of its appearance by dexamethasone.

We measured tracheal fluid production rate and pulmonary surfactant flux in 12 sets of twin fetal lambs of 102 to 135 days' gestation. In nine of these sets we also measured surfactant flux and the concentration of saturated lecithin in the lungs before and during dexamethasone infusion into one of the twins. The average tracheal fluid flow rate was 3.25 ml/kg/hr (SD 1.6) and, relative to body weight, did not change between 103 and 135 days' gestation or during the infusion of dexamethasone. In untreated fetuses, surfactant was detected between 108 and 130 days and its flux gradually increased but remained less than 150mug/kg/hr at 135 days' gestation; the amount of disaturated lecithin in the lung relative to body weight increased 13-fold from 108 to 134 days. From these data we calculated that the minimal rate of synthesis of disaturated lecithin was increased about 4-fold by dexamethasone from 108 to 120 days; this enhancement fell to about 1.8-fold by 135 days. The rate of secretion of disaturated lecithin, as estimated by surfactant flux in tracheal fluid, was increased by dexamethasone throughout the interval of the study.

Systemic vasculitis associated with eosinophilia and marked degranulation of tissue eosinophils.

A 9-year-old girl had fever, life-threatening interstitial pneumonia, a vesicular skin rash, parotid and submandibular swelling, and marked blood eosinophilia (WBC count 47,000/microL, 39% eosinophils). Results of lung and skin biopsies showed vasculitis with intense eosinophil infiltration. Immunofluorescence analyses of these biopsies, as well as analysis of tissue from the parotid, lip, and trachea, showed striking deposition of eosinophil granule major basic protein associated with areas of tissue damage. Treatment with glucocorticoids and hydroxyurea produced clinical improvement. The association between tissue damage and deposition of the cytotoxic major basic protein suggests that the eosinophil contributed to the pathophysiology of this disease. Recognition of the capacity of the eosinophil to produce tissue damage can be helpful in the selection of therapy.

Evidence for an A2/Ra adenosine receptor in the guinea-pig trachea.

1 An attempt was made to determine whether the extracellular adenosine receptor that mediates relaxation in the guinea-pig trachea is of the A(1)/R(i) or A(2)/R(a) subtype.2 Dose-response curves to adenosine and a number of 5'- and N(6)-substituted analogues were constructed for the isolated guinea-pig trachea, contracted with carbachol.3 The 5'-substituted analogues of adenosine were the most potent compounds tested, the order of potency being 5'-N-cyclopropylcarboxamide adenosine (NCPCA) > 5'-N-ethylcarboxamide adenosine (NECA) > 2-chloroadenosine > L-N(6)-phenylisopropyladenosine (L-PIA) > adenosine > D-N(6)-phenylisopropyladenosine (D-PIA).4 The difference in potency between the stereoisomers D- and L-PIA on the isolated trachea was at the most five fold.5 Responses to low doses of adenosine and its analogues were attenuated after treatment with either theophylline or 8-phenyltheophylline. The responses to 2-chloroadenosine were affected to a lesser extent than were those to the other purines.6 Adenosine transport inhibitors, dipyridamole and dilazep, potentiated responses to adenosine, did not affect those to NCPCA, NECA, L-PIA and D-PIA but significantly reduced the responses to high doses of 2-chloroadenosine.7 Relaxations evoked by 9-beta-D-xylofuranosyladenosine which can activate intracellular but not extracellular adenosine receptors, were attenuated by dipyridamole but unaffected by 8-phenyltheophylline.8 The results support the existence of an extracellular A(2)/R(a) subtype of adenosine receptor and an intracellular purine-sensitive site, both of which mediate relaxation.

Ontogeny of epidermal growth factor receptor and lipocortin-1 in fetal and neonatal human lungs.

The ontogeny and distribution of the epidermal growth factor (EGF) receptor and lipocortin-1, a major cellular substrate of the EGF receptor, were evaluated in a developmental series of fetal and neonatal human lungs (8 to 41 weeks' gestation and stillborn to 16 days' postnatal age). The peroxidase anti-peroxidase technique with two polyclonal antibodies recognizing the EGF receptor and one polyclonal antibody recognizing lipocortin-1 were used for immunohistochemical localization. Extensive or scattered bronchiolar EGF receptor immunoreactivity appeared in the entire series of frozen lung specimens from 15 to 32 weeks' gestation. Bronchial glands exhibited EGF receptor immunostaining from 19 weeks onward, and immunoreactivity in bronchial epithelium was detected from 23 weeks onward. Most tracheas showed extensive lipocortin-1 immunoreactivity in the epithelium beginning at 10 weeks' gestation. Immunostaining was also seen in cells lining the ducts of submucosal glands after 15 weeks' gestation and in nonmucous acinar cells of tracheal glands after their appearance at 18 weeks' gestation. Bronchial epithelium exhibited lipocortin-1 immunoreactivity from 12 weeks' gestation onward. Bronchial gland necks became immunostained from 16 weeks' gestation onward, followed by acinar immunostaining as they subsequently developed. Bronchiolar epithelium was immunostained as early as 12 weeks, beginning with the largest airways, and by 24 weeks extending distally to the bronchioloalveolar portals. Lipocortin-1 immunostaining of larger conducting airway epithelium was primarily confined to ciliated cells. Neither EGF receptor nor lipocortin-1 immunoreactivity was detected in alveolar type I or type II cells, fibrocytes, chondrocytes, or smooth muscle cells at any gestational age. These developmental patterns suggest that the EGF receptor and lipocortin-1 may participate in normal growth factor-induced proliferation of the conducting airways and their glands in the human fetal lung and trachea.

Transepithelial prostacyclin gradient in isolated canine trachea.

Cyclooxygenase products of arachidonic acid, potential modulators of airway smooth muscle, have recently been described in bronchoalveolar lavage from canine lungs. To evaluate the possibility that airway epithelium represents a barrier to movement of prostacyclin (PGI2), an important bronchodilator synthesized by isolated airway, we measured the concentrations of 6-oxoprostaglandin F1 alpha (6-oxo-PGF1 alpha), the stable degradation product of PGI2, on the mucosal and serosal sides of isolated canine tracheal segments (CTS) mounted in Ussing chambers. 6-oxo-PGF1 alpha was measured by radioimmunoassay after purification by high-performance liquid chromatography. The concentration of 6-oxo-PGF1 alpha was significantly higher on the serosal than the mucosal side of CTS (1,262 +/- 252 vs. 390 +/- 168 pg.min-1.g-1, n = 8, P less than 0.05). A significant correlation was present between 6-oxo-PGF1 alpha measured on both sides of each CTS (r = 0.778, n = 26, P less than 0.01). 6-oxo-PGF1 alpha production from CTS stripped of mucosa was significantly greater than from isolated mucosa. Radiochromatograms obtained after incubation with [3H]arachidonic acid and calcium ionophore A23187 confirmed PGI2 as the predominant cyclooxygenase product of the submucosa, whereas the mucosa produced only small amounts of PGI2 in proportion to other cyclooxygenase products. PGI2 (10(-8) to 10(-6) M) applied to the mucosal surface of closed tracheal segments precontracted with histamine resulted in no significant relaxation, whereas serosal application showed a concentration-dependent effect. Radiolabeled 6-oxo-PGF1 alpha did not cross the isolated epithelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Electrolyte and other chemical concentrations in tracheal airway surface liquid and mucus.

With the ferret in vitro tracheal preparation, we measured the electrolyte and chemical composition of airway surface liquid (ASL) under control conditions and when drugs were added to promote submucosal gland secretion and to change epithelial ion transport. Control ASL was hyperosmolar (342 +/- 2.8 mosmol/kg) compared with ferret plasma and surrounding buffer. Higher values were also found for sodium (167 +/- 1.7 mmol/l), potassium (9.0 +/- 0.05 mmol/l), total calcium (3.46 +/- 0.11 mmol/l), and ionized calcium (2.55 +/- 0.18 mmol/l). pH was lower (7.12 +/- 0.03) than in plasma or buffer. Addition of methacholine to the surrounding buffer increased flow of ASL and potential difference across the mucosa and lowered pH, calcium, sodium, and chloride concentrations. Potassium concentration was increased. Phenylephrine increased flow and decreased calcium concentrations. Salbutamol (albuterol) had no effect on flow but decreased pH and increased calcium and potassium concentrations. Histamine increased flow and calcium concentrations and decreased pH. These changes are presumably due to changes in gland secretion and epithelial transport. Methacholine and phenylephrine increased the sugar content of the secretions, the changes with phenylephrine being larger. Thus resting ASL is hyperosmolar and relatively acid, with high cation contents, and administration of drugs changes its composition by actions on submucosal glands and epithelium.

Luminal receptors for bradykinin on the canine tracheal epithelium: functional subtyping.

Luminal addition of bradykinin (BK) to the open-circuited canine tracheal epithelium produces a biphasic response in transmucosal potential difference (P.D.): a rapid, transient decrease (dip) followed by a subsequent, more sustained increase (rise), both phases being associated with an increase in conductance. We have attempted to characterise the receptor subtype mediating the bradykinin response. Lys-bradykinin (Lys-BK) elicited a similar response, and its EC50 as judged from concentration-response relations was similar to that of BK. Cross-tachyphylaxis between the two peptides confirmed a common receptor. Des-Arg9-BK (a B1-agonist) neither elicited a response nor inhibited responses to BK. The novel B2-antagonist [Thi6,9-D-Phe8]kallidin reversibly inhibited responses to both BK and Lys-BK. The rapid changes in P.D. (dips) were unaffected by Na+ removal, but were eliminated by replacing luminal Cl- with isethionate. Thus, BK, acting on B2-receptors, transiently increases anion permeability of the luminal membrane of the canine tracheal epithelium.