Airway epithelium-derived inhibitory factor.

Various bronchoactive agents can induce the release from the airway epithelium of an inhibitory substance that is able to relax certain tissues including rat aorta and possibly also airway smooth muscle. This substance, whose existence has recently been confirmed using a new bioassay system, is distinct from nitric oxide (EDRF) and is also known to be non-prostanoid in nature. Roy Goldie and colleagues describe the properties of this factor, and its potential clinical significance.

Ganglion cells of the cat accessory optic system: morphology and retinal topography.

The ganglion cells of the cat's retina form classes that are distinct in their cell morphology, retinal distribution, central projections, and functional properties. By means of the retrograde transport of horseradish peroxidase injected into the accessory optic nuclei of the cat midbrain, we have characterized the retinal ganglion cells projecting to these nuclei. The retinal projection is virtually completely crossed to the medial terminal nucleus and to the lateral terminal nucleus. This appears to be true for the dorsal terminal nucleus as well, although difficulties of the technique limit our findings for this region. No differences were found in either the spatial distribution, or the somal size distribution, or the morphological characteristics of the ganglion cells projecting to these three nuclei. In spatial distribution, these cells are concentrated in the area centralis and visual streak and show no evidence of a nasotemporal division. Morphologically, they have small to medium-sized somas and delicate, sparsely branching dendrites. They do not appear to belong to any of the morphological cell type thus far identified.

The retinal projection to the cat pretectum.

Retinal ganglion cells were labeled retrogradely by localized injections of HRP into different regions of the pretectum, tectum, and optic tract in 26 cats. Retinal projection zones in the pretectum were labeled anterogradely in the same cats by intravitreal injections of 3H-proline. This allowed the HRP injection sites to be located with respect to the retinal termination zones. The form of the projection zones from retina to pretectum was determined from serial reconstructions of either coronal or horizontal sections. The zones are best distinguished in horizontal sections, where they are seen as four roughly parallel strips on either side of the brain. They are more-or-less parallel to the anterior border of the tectum, and appear to traverse the entire width of the retinal projection to the tectum. Each zone is similar in form for the ipsilateral and contralateral projections, although the contralateral projection is thicker and denser. Binocular injections of 3H-proline showed that the projections from the two eyes were in register and did not interdigitate. Cells labeled by HRP injections in the anteromedial end of the pretectum were concentrated in the lower nasal quadrant of the contralateral retina, and the lower temporal quadrant of the ipsilateral retina. Posterolateral injections labeled cells in the upper quadrants. There is thus a rough retinotopic mapping along the elongated axis of the pretectum. When the distributions of ganglion cells labeled by HRP injections to different parts of the pretectum are combined, they show a concentration in both the visual streak and area centralis, and thereby reflect, at least qualitatively, the relative spatial distribution of the entire ganglion-cell population. About 85% of the retinal projection to the pretectum is contralateral. For all of the HRP injections, the spatial density of labeled cells was always low, accounting for no more than 3% of the total spatial density of ganglion cells in any retinal region. Several types of ganglion cells were labeled following injections to most regions of the pretectum; these included alpha, beta, and epsilon cells, as well as small-bodied cells showing a variety of morphologic forms. Alpha cells were labeled mainly from the anterolateral end of the pretectum, but other cell types were labeled from all injected regions. In the peripheral retina, 2% of the labeled cells were alpha cells, 32% were beta cells, 19% were epsilon cells, and the remaining 47% were small cells whose dendrites only occasionally filled to any significant extent.(ABSTRACT TRUNCATED AT 400 WORDS)

Aminoalkynyldithianes. A new class of calcium channel blockers.

Several dithiane derivatives, prepared as intermediates for compounds structurally related to the therapeutically useful antimuscarinic agent oxybutynin, were effective inhibitors of calcium ion induced contraction of guinea pig ileal strips and of KCl-induced calcium entry into neuronal cells. Although the first member of this series, 2-[5-(diethylamino)-3-pentynyl]-1,3-dithiane (2a), was only marginally effective, its condensation product with diphenyl ketone, i.e. 2-[5-(diethylamino)-3-pentynyl]-2-(a,a-diphenyl-a- hydroxymethyl)-1,3-dithiane (3a), demonstrated weak, but significant, calcium channel antagonist activity. As part of a structure-activity relationship (SAR) study, various structural analogues of 2a and 3a were prepared and examined for calcium antagonist properties. In addition to these structural types, ring bridged (tricyclic) congeners of 3, i.e. 4, related bicyclic compounds 5, dehydroxylated derivatives 6, some homologous 2-[[[(N,N-disubstituted-amino)methyl]2- phenyl-1,3-dithianes (7), and a series of 2-[6-[N,N-disubstituted-amino)methyl]-1-hydroxy-1-phenyl- 4-hexynyl]-1,3-dithianes (8) were prepared and studied for calcium channel blocking activity. In general, greatest potency was noted in the tricyclic series 4; however, a definitive SAR could not be established. A structural similarity between several potent calcium antagonists having the structures 7c, 8b, and 8d and the well-known calcium channel blockers verapamil and tiapamil suggests these compounds may act at the same site. Compounds in the other classes (2-6) failed to show clearly defined SAR and their potency differed markedly in two tests for calcium channel antagonist activity. These results may indicate that the dithiane derivatives 2-6 produce their effects in a manner differing from that of the calcium channel antagonists diltiazem, verapamil, and nitrendepine.

Severe bleeding diathesis associated with invasive aspergillosis in transplant patients.

A severe bleeding disorder developed in eight renal transplant patients with invasive aspergillosis. The hemorrhagic diathesis was characterized by wound oozing, severe upper and lower gastrointestinal tract hemorrhage, and mucosal bleeding at other sites. This unusual coagulopathy was characterized by a prolonged thrombin time, which was corrected with protamine sulfate, and an abnormal Reptilase time. The bleeding disorder antedated the diagnosis of invasive aspergillosis in all cases. The probability that the coagulopathy was due to proteolytic enzymes elaborated by Aspergillus sp. is discussed.

Effects of a novel nonpeptide bradykinin B2 receptor antagonist on intestinal and airway smooth muscle: further evidence for the tracheal B3 receptor.

1. We examined the effects of phosphonium, [[4-[[2- [[bis(cyclohexylamino)methylene]amino]-3-(2-naphthalenyl) 1-oxopropyl]amino]-phenyl]-tributyl, chloride, monohydrochloride (WIN 64338), a novel, nonpeptide bradykinin B2 receptor antagonist, on bradykinin-induced contractions of guinea-pig isolated ileum, and guinea-pig and ferret trachea. 2. WIN 64338 potently and competitively antagonized ileal contractions, in response to bradykinin, exhibiting a pA2 value of 7.97 +/- 0.10. The compound was without effect on contractions elicited by methacholine, a muscarinic receptor antagonist. Thus, WIN 64338 is a competitive and selective antagonist of ileal B2 receptors. 3. In contrast, WIN 64338 was completely without effect on bradykinin-induced contractions of guinea-pig or ferret trachea. Thus, even at a concentration of 1 microM, which was sufficient to cause a 100 fold decrease in ileal sensitivity to bradykinin, WIN 64338 failed to shift the bradykinin log concentration-response curves in trachea isolated from either species. 4. These data confirm that WIN 64338 represents the first reported nonpeptide antagonist of guinea-pig ileal B2 receptors. They also provide additional evidence for heterogeneity of bradykinin receptors within the same species (guinea-pig) and, furthermore, indicate that the tracheal bradykinin receptor (B3?) is different from that in ileal tissue (B2).

The effects of vasoactive intestinal peptide (VIP) antagonists, and VIP and peptide histidine isoleucine antisera on non-adrenergic, non-cholinergic relaxations of tracheal smooth muscle.

1. The effects of several drugs, including antagonists of vasoactive intestinal peptide (VIP), and antisera to VIP or peptide histidine isoleucine (PHI), on relaxation responses of guinea-pig isolated trachea to electrical field stimulation (EFS) have been examined. 2. beta-Adrenoceptor blockade with propranolol only partially blocked the inhibitory response to EFS, but had no effect in tissues from animals pretreated with 6-hydroxydopamine or reserpine. 3. Neither adenosine deaminase, in the presence of dipyridamole, nor the potent adenosine antagonist NPC205 (1,3-n-dipropyl-8-(4-hydroxyphenyl)-xanthine) had any effect on the inhibitory response to EFS. 4. The VIP antagonists, [Ac-Tyr1, D-Phe2]-GRF(1-29)-NH2 and [4-Cl-D-Phe6, Leu17]-VIP had no effect on the inhibitory response to EFS. Moreover, they were without effect on responses to exogenous VIP or PHI. 5. Overnight incubation with VIP antisera markedly reduced the inhibitory response to EFS. PHI antisera had a similar, but smaller effect. 6. In the presence of a concentration of VIP that is maximal for its relaxant effect, inhibitory responses to electrical stimulation were greatly inhibited. 7. Naloxone and reactive blue 2 each had no effect on inhibitory responses indicating that endogenous opioids and adenosine 5'-triphosphate (ATP) respectively are not involved. 8. The results suggest that VIP and PHI, but not adenosine, contribute to non-adrenergic, noncholinergic inhibitory nerve responses of guinea-pig trachea. Moreover, the surprising lack of effect of both VIP antagonists on these responses, and in particular, on responses to exogenous VIP, suggests that the receptors mediating VIP-induced tracheal relaxation are different from those that mediate pancreatic secretion.

Effects of p-chlorophenylalanine on the sensitivity of rat intestine to agonists and on intestinal 5-hydroxytryptamine levels during Nippostrongylus brasiliensis infection.

Infection of rats with the nematode N. brasiliensis caused non-specific increases in maximum response of isolated intestine to acetylcholine and 5-hydroxytryptamine (5-HT), and a specific subsensitivity to 5-HT. Intestinal levels of 5-HT, measured fluorimetrically, increased approximately 2 fold during infection. Treatment of infected rats with parachlorophenylalanine (PCPA) depleted the gut of 5-HT, and prevented the specific subsensitivity to the amine but not the increases in maximum response. Depletion of intestinal 5-HT did not prevent the immune expulsion of the parasites. It is concluded that the specific subsensitivity of the gut is due to the elevated levels of 5-HT during infection, but that the increased maximum responses are due to some other factor. Further, the lack of effect of PCPA on parasite rejection casts doubt on the proposed role of 5-HT in this process.

Effects of epithelium removal on relaxation of airway smooth muscle induced by vasoactive intestinal peptide and electrical field stimulation.

1. We have studied the effect of epithelium removal on relaxation of guinea-pig isolated tracheal smooth muscle induced by vasoactive intestinal peptide (VIP) or stimulation of non-adrenergic, non-cholinergic (NANC) inhibitory nerves. Also examined were the effects of inhibitors of neutral endopeptidase (NEP) and angiotensin-converting enzyme (ACE). 2. Epithelium removal produced a 3.6 +/- 0.4 fold leftward shift in the VIP concentration-response curve. The supersensitivity to VIP, following epithelium removal was abolished by phosphoramidon or thiorphan (NEP inhibitors), but unaffected by captopril (an ACE inhibitor). In intact trachea, the NEP inhibitors produced leftward shifts in the VIP curves similar to those produced by epithelium removal. 3. In contrast to responses to exogenous VIP, neurogenic NANC inhibitory responses to electrical field stimulation were affected neither by epithelial denudation nor by the peptidase inhibitors. 4. As in previous studies, epithelium removal increased tracheal sensitivity to isoprenaline. This was not altered by pretreatment with a cocktail of peptidase inhibitors. Thus, the effect of the NEP inhibitors on responses to VIP appears to be relatively specific. 5. These data indicate that exogenous VIP is a substrate for airway NEP, since inhibition of the enzyme potentiates the peptide. This is further evidence that the airway epithelium provides a source for the metabolism of mediators. 6. In guinea-pig trachea the NEP responsible for cleaving VIP may be located largely in the epithelial layer, since NEP inhibition was without effect on sensitivity to VIP in epithelium-denuded preparations. If VIP is a NANC inhibitory neurotransmitter in this tissue its degradation endogenously does not appear to involve epithelial NEP.

Effects of peptidases on non-adrenergic, non-cholinergic inhibitory responses of tracheal smooth muscle: a comparison with effects on VIP- and PHI-induced relaxation.

1. The effects of peptidase enzymes on non-adrenergic, non-cholinergic (NANC) inhibitory responses of guinea-pig trachea to electrical field stimulation (EFS), and on relaxations induced by vasoactive intestinal peptide (VIP) and peptide histidine isoleucine (PHI) have been examined. 2. alpha-Chymotrypsin reduced both the magnitude and, particularly, the duration of the inhibitory response to EFS, whereas papain reduced only the magnitude. Aprotinin, a peptidase inhibitor prevented the effects of alpha-chymotrypsin but was without effect on papain. 3. alpha-Chymotrypsin and papain both abolished relaxant responses to exogenous VIP and PHI. The action of alpha-chymotrypsin was prevented by aprotinin, whereas that of papain was not affected. 4. The peptidases were without effect on concentration-response curves to methacholine or to isoprenaline. It was also observed that, in the absence of the peptidases, aprotinin had no effect on inhibitory responses either to EFS or to exogenous VIP and PHI. 5. It is suggested that neuropeptides, possibly VIP and PHI, released during EFS of guinea-pig trachea, partly mediate NANC relaxations, and that their action may be inhibited by peptidases. However, the lack of effect of aprotinin alone, on responses to EFS, suggests that, if endogenous peptidases are important in terminating the action of neuropeptides, they are resistant to the effect of this particular peptidase inhibitor. It is further suggested that neurogenic relaxation of guinea-pig trachea is also partly mediated by a substance, possibly non-peptide, other than VIP or PHI.

Adenosine receptor-mediated contraction and relaxation of guinea-pig isolated tracheal smooth muscle: effects of adenosine antagonists.

1. The effects of several adenosine analogues and antagonists on guinea-pig isolated trachea have been examined. 2. 5'-N-ethylcarboxamidoadenosine (NECA), 5'-N-methylcarboxamidoadenosine (MECA) and adenosine (in the presence and absence of dipyridamole) elicited concentration-dependent tracheal relaxation. 3. The R(-)- and S(+)-enantiomers of N6-(2-phenylisopropyl)adenosine (R-PIA and S-PIA respectively), N6-cyclohexyladenosine (CHA) and 2-chloroadenosine (CADO) caused contractions at low concentrations (0.05-2.0 microM), whereas at higher concentrations, relaxation resulted. 4. For tracheal relaxation, the adenosine analogues exhibited the following rank order of potency: NECA greater than CADO greater than R-PIA = MECA greater than S-PIA greater than adenosine. The rank order of potency for inducing contractions was R-PIA greater than CHA greater than CADO greater than S-PIA. These data suggest that relaxation is mediated by adenosine A2-receptors, whereas contraction is the result of activation of A1-receptors. 5. 8-Phenyltheophylline (8-PT), aminophylline, the triazoloquinazoline CGS 15943A and NPC205 (1,3-di-n-propyl-8-(4-hydroxyphenyl)xanthine) each inhibited the R-PIA-induced contractile response, whereas enprofylline was without effect. NPC205, aminophylline and 8-PT were competitive antagonists, but CGS15943A was non-competitive. 6. That the most potent antagonist was the A1-selective agent, NPC205 (pA2 = 7.80), further suggests that the contraction is mediated by A1-receptors. Moreover, NPC205 was 13 times more potent as an antagonist of R-PIA-induced contractions (A1) than of NECA-induced relaxations (A2). 7. The antagonists were also found to relax the trachea by an unknown mechanism. That enprofylline did not antagonize the R-PIA-induced contractions, but was 3-4 times more potent a tracheal relaxant than aminophylline, further suggests that a direct effect on airway smooth muscle, rather than antagonism of endogenous adenosine, is more relevant to the bronchodilator effect of alkylxanthines in the treatment of asthma.

The effects of epithelium removal on the sensitivity of guinea-pig isolated trachealis to bronchodilator drugs.

Mechanical removal of the epithelium increased the sensitivity of tracheal strips to isoprenaline, sodium nitroprusside, and to adenosine (only in the presence of inhibitors of its uptake and metabolism). Epithelium removal was without effect on sensitivity to salbutamol or papaverine. Preincubation of tracheal strips with an inhibitor of extraneuronal uptake, corticosterone (50 microM), had no effect on tissue sensitivity to either salbutamol or papaverine. However, the steroid both increased sensitivity to isoprenaline, and abolished the effect of epithelium removal on sensitivity to this catecholamine. These results suggest that in the guinea-pig, the tracheal epithelium is a major source of extraneuronal uptake for catecholamines. Furthermore, the increase in trachealis sensitivity to isoprenaline following epithelium removal is probably due to loss of these sites of extraneuronal uptake. The fact that sensitivity to salbutamol, papaverine and adenosine (in the absence of metabolic inhibitors) was not increased by denuding the epithelium indicates that loss of a diffusion barrier to drugs is not the mechanism of increased sensitivity. Adenosine (and possibly nitroprusside) may cause the epithelium to release a smooth muscle excitatory factor. Thus, removal of the epithelium attenuates this excitatory influence and enhances smooth muscle responsiveness to adenosine. These results provide further evidence that the epithelium has an important role in modulating the sensitivity of guinea-pig trachealis to drugs.

Relaxation of guinea-pig tracheal smooth muscle to arachidonate is converted to contraction following epithelium removal.

1 The effect of epithelium removal on responses of guinea-pig isolated trachealis to sodium arachidonate has been examined. 2 Arachidonate (100 microM) caused relaxation of epithelium-intact preparations, but following epithelium removal, the response to arachidonate was converted to contraction. In the presence of indomethacin (1 microM), arachidonate caused contraction in intact and denuded trachea. 3 Arachidonate also produced concentration-dependent effects, the qualitative nature of which varied with the presence or absence of the epithelium. In the presence of indomethacin, tracheal strips contracted in a concentration-dependent manner whether or not the epithelium had been removed. 4 Nordihydroguaiaretic acid (NDGA; 10 microM) markedly inhibited the contractile response of denuded strips to arachidonate. In intact tissues this lipoxygenase inhibitor converted the arachidonate-induced relaxation to a concentration-dependent contraction. The contraction to arachidonate, in the presence of NDGA, was epithelium-dependent. In the presence of both indomethacin and NDGA, responses to arachidonate were abolished. 5 It is concluded that the relaxation of guinea-pig trachea to arachidonic acid is epithelium-dependent and is mediated by an inhibitory product of the cyclo-oxygenase metabolic pathway. The contraction in denuded trachea, and trachea in the presence of indomethacin, may be mediated by lipoxygenase products of arachidonic acid metabolism, i.e. peptidoleukotrienes. The mediator of the epithelium-dependent contraction in NDGA-treated tissues is unknown.

Effects of bradykinin receptor antagonists on antigen-induced respiratory distress, airway hyperresponsiveness and eosinophilia in guinea-pigs.

1. We examined effects of bradykinin (BK) receptor antagonists on airway hyperresponsiveness and eosinophilia in sensitized guinea-pigs that had been administered single, as well as repeated (chronic) challenges with inhaled ovalbumin. In addition, the effects of BK antagonists on antigen-induced respiratory distress during the chronic study were noted. 2. At 24 h following single antigen challenge, guinea-pigs exhibited airway hyperresponsiveness to the bronchoconstrictor effect of i.v. histamine, characterized by a left shift in the dose-response curve. In addition, responses to the maximum dose of histamine that could be used were significantly increased in hyperresponsive guinea-pigs. The percentages of bronchoalveolar fluid, eosinophil and neutrophils also increased. 3. A BK B1 receptor antagonist, desArg9-[Leu8]-BK, significantly inhibited airway hyperresponsiveness induced by single antigen challenge. A B2 receptor antagonist, D-Arg-[Hyp3, Thi5,8,D-Phe7]-BK (NPC 349) had a small, but statistically significant inhibitory effect on responsiveness to the highest histamine dose in challenged animals. DesArg9-[Leu8]-BK significantly inhibited the neutrophilia, whereas NPC 349 inhibited infiltration by both cell types. 4. Chronic antigen challenge also caused airway hyperresponsiveness to i.v. acetylcholine (ACh), distinguished by an increase in the slope of the dose-response curve. Thus, the magnitude of the bronchoconstrictor responses to the maximum dose of ACh that could be used was significantly increased. No change in sensitivity to ACh was evident. Marked eosinophilia was also noted in the trachea, bronchi and lung parenchyma. 5. Airway hyperresponsiveness and eosinophilia, induced by chronic antigen challenge, were markedly inhibited by the B2 antagonists, D-Arg-[Hyp3,D-Phe7]-BK (NPC 567) or D-Arg-[Hyp3,Thi5d-Tic7,Tic8]-BK (NPC 16731).NPC 16731 also abolished antigen-induced cyanosis, and delayed the onset of dyspnoea,doubling the time taken for animals to exhibit respiratory distress.6. The ability of BK receptor antagonists to inhibit antigen-induced airway hyperresponsiveness, in addition to eosinophilia, indicates an important role for endogenous kinins. Moreover, the abrogation of eosinophil infiltration suggests that BK has a significant function in maintaining allergic inflammation of the airways.

Differential effects of epithelium removal on the responsiveness of guinea-pig tracheal smooth muscle to bronchoconstrictors.

1 The influence of the epithelium on contractions produced by the peptidoleukotrienes, 5-hydroxytryptamine (5-HT) and the thromboxane mimetic, U-44069, was examined in trachea from control and ovalbumin-sensitized guinea-pigs. 2 In control tissues removal of the epithelium produced an approximately 2 to 4 fold leftward shift in leukotriene C4 (LTC4) and LTD4 concentration-response curves, but no effect on LTE4-induced contractions. Similar results were obtained in preparations from ovalbumin-sensitized animals. 3 Responses produced by 5-HT or U-44069 were similar in the presence and absence of the epithelium in control guinea-pigs. 4 Indomethacin produced contrasting effects on leukotriene-induced contractions in control guinea-pigs: an increase in sensitivity to LTC4 in the presence but not absence of the epithelium, no effect on LTD4-induced contractions and a decrease in sensitivity to LTE4 in both epithelium-containing and epithelium-free preparations. 5 These results indicate that there is selectivity in the effects of epithelium removal on agonist-induced contractions of the guinea-pig trachea. This provides further evidence for the modulatory influence of the epithelium on the reactivity of mammalian airway smooth muscle and supports the postulated existence of an epithelium-derived inhibitory factor. The observation that in intact trachea indomethacin mimics the effects of epithelium removal on LTC4-induced responses, suggests the involvement of a prostanoid(s) in this phenomenon.

D-Arg[Hyp3-Thi5-D-Tic7-Tic8]-bradykinin, a potent antagonist of smooth muscle BK2 receptors and BK3 receptors.

D-Arg[Hyp3-Thi5-D-Tic7-Tic8]-bradykinin (NPC 16731) inhibited bradykinin (BK) binding and BK-induced contraction in guinea-pig ileum, being markedly more potent than D-Phe7-BK analogues as a BK2 receptor antagonist. In isolated trachea NPC 16731, unlike other BK2 antagonists, inhibited BK binding and BK-induced contraction, and 45Ca2+ efflux in tracheal smooth muscle cells. That NPC 16731 potently inhibits BK effects in trachea provides further evidence for the existence of the airway BK3 receptor.

Cutaneous microabscess formation from Alernaria alternata.

A 69-year-old farmer with acute lymphocytic lymphoma developed multiple ulcerated hemorrhagic cutaneous lesions. Alternaria alternata was cultured from the lesions and hyphae were seen in biopsies of the lesions. The potential role of Alternaria as a human pathogen is discussed.

Effects of bradykinin receptor agonists, and captopril and thiorphan in ferret isolated trachea: evidence for bradykinin generation in vitro.

Bradykinin B2 receptor agonists, but not a B1 receptor agonist, were potent spasmogens of ferret isolated trachea. Bradykinin-induced contractions were unaffected by several pharmacological agents, indicating a direct effect on airway smooth muscle B2 receptors. Captopril plus thiorphan caused contractions in approximately 70% of preparations that were abolished by B2 receptor antagonists, but not by B1 receptor antagonist. Thus, ferret tracheal tissues appear capable of releasing and degrading kinins in vitro. Inhibition of peptide degradation with captopril and thiorphan may allow the endogenous kinins to accumulate in concentrations sufficient to elicit tracheal contraction via activation of B2 receptors.

Epithelium removal increases the reactivity of human isolated tracheal muscle to methacholine and reduces the effect of verapamil.

Removal of the epithelium increased the sensitivity of human isolated tracheal smooth muscle to methacholine, producing a greater than 2-fold leftward shift in the concentration-response curve. Concomitantly, the ability of verpamil to lower the maximum contractile response was reduced in tissues without an intact epithelium. These findings suggest a role for the epithelium in modulating the reactivity of human and extend the findings of similar studies in experimental animals.

Cloning, sequencing and functional expression of a guinea pig lung bradykinin B2 receptor.

Kinin receptors are classified as B1 and B2 based upon agonist and antagonist potencies and cloning and expression studies. Using sequences from human and rat bradykinin B2 receptors, polymerase chain reaction (PCR) was utilized to isolate cDNA from guinea pig lung. The receptor obtained is predicted to have 372 amino acids and shares > 80% sequence homology with human, rat, rabbit and mouse B2 receptors. In competition binding experiments in Chinese hamster ovary (CHO-K1) cells in which the guinea pig cDNA was expressed, [3H]bradykinin was displaced by kinin receptor ligands with an order of potency consistent with a B2 subtype. In CHO cells expressing the guinea pig receptor, bradykinin caused a concentration 45Ca2+ efflux. A B1 receptor agonist, desArg9-bradykinin, also caused 45Ca2+ efflux but with a potency several orders of magnitude lower than bradykinin. Curiously, several B1 and B2 receptor antagonists induced 45Ca2+ efflux, indicating that this receptor may be coupled differently in CHO cells than in native tissues.