Receptors in Asthmatic Airways1-3 R. G. GOLDIE
Introduction Asthma is an inflammatory lung disease involving reversible airway obstruction. The characteristic features of asthma include eosinophilia, nonspecific bronchial hyperreactivity to inhaled spasmogens, airway epithelial damage, mucosal edema, and mucosal gland hypersecretion. Examining the role of inflammatory mediators in these processes is critical to an understanding of the causes of asthma. A major element in such an assessment is the study of the distribution, density, and function of receptors for such chemicals in asthmatics compared with that in healthy human lung. Histamine Receptors Inhalation of allergens causes significant degranulation of mast cells in the bronchial wall in allergic asthmatics with release of histamine and several other putative mediators of asthma, including prostaglandin (PO) D, and sulfidoleukotrienes, These mediators may increase airway smooth muscle tone and promote mucosal edema and glandular secretion, resulting in narrowing of the airways and limiting airflow. This is seen in the immediate bronchial response in asthma (1).
Airway Smooth Muscle The causes of asthma-associated bronchial hyperreactivity to spasmogens such as histamine are unknown. Recent evaluations in vitro of the responsiveness of human bronchial preparations obtained postmortem from healthy and asthmatic lung to histamine and to carbachol indicate that no major intrinsic abnormality exists with respect to airway smooth muscle contractility. Indeed the potencies of these spasmogens were marginally lower in asthmatic bronchi than in tissue from healthy lung (2), while maximal responsiveness was similar in bronchi from both sources. These data also indicate that histamine H I receptor and muscarinic cholinoceptor function were not up-regulated in bronchial smooth muscle in asthma. Interestingly, histamine H I receptors have been detected using immunohistochemical techniques in greater numbers in airway epithelial cells than in either guinea pig airway or vascular smooth muscle (3). Histamine Hj-receptors might be expected to mediate relaxation of human airway smooth muscle since they are linked to the generation of cyclic adenosine monophosphate (cAMP). However, there is little evidence for their existence in functionally significant numbers in either normal or asthmatic subjects (4). No role for histamine Hs-receptors in airways has yet been established. Bronchial Epithelium Desquamation of the bronchial epithelium
SUMMARY Airway tissue obtained postmortem from nondiseased and severely asthmatic human lung was used in functional, radlollgand binding and autoradiographic studies to investigate aspects of various receptor systems for putative mediators of asthma. Results indicate that asthma does not involve an intrinsic abnormality of smooth muscle contractility to spasmogens. Nor was there any evidence for up-regulation of histamine H,-receptor, muscarinic cholinoceptor, or a,·adrenoceptor function. Conversely, severe asthma involving intense airway inflammation resulted in significant I}-adrenoceptor dysfunction, probably caused by receptor uncoupling from adenylate cyclase. Evidence was also obtained for histamine and methacholine-induced release of a nonprostanoid, airway epithelium-derived Inhibitory factor (EpDIF), which may have a significant dilator effect In the adjacent bronchial circulation. The activity of this potentially protective Inhibitory autacoid system would be expected to be reduced in asthma, which commonly involves epithelium damage. The autoradiographic distribution of specific binding sites for "'I-labeled substance P (I-SP) was also determined in bronchi from healthy humans and asthmatics. In sharp contrast to guinea pig airways where high levels of binding were detected over smooth muscle, specific I-SP binding was sparse over human airway smooth muscle from both sources, while dense binding was associated with structures immediately beneath the epithelium and with deep submucosal glands. These data suggest a more significant role for SP in secretory processes than In spasmogenic processes in human bronchi and highlight the potential for drawing Invalid conclusions concerning human airway function from studies using animal models. AM REV RE5PIR DIS 1990; 141:5151-5156
is a consistent and striking feature in asthma (5) and may be causally related to bronchial hyperreactivity. Recent studies have demonstrated that deliberate removal of the airway epithelium causes increased smooth muscle responsiveness to histamine, providing indirect evidence for the existence of an epithelium-derived inhibitory factor (EpDlF), generated in response to some spasmogens (6, 7). In nonasthmatics, bronchial reactivity to inhaled histamine is limited, while asthmatics respond as if lacking a braking mechanism opposing bronchoconstriction (8). EpDlF activity, which might be absent or reduced in asthma as a result of epithelial damage, may contribute to such a braking mechanism in healthy lung. Direct evidence for EpDIF release from guinea pig trachea and from human bronchus in response to histamine and methacholine (9) has now been demonstrated in a coaxial bioassay system (figure 1). These data also indicate the existence of epithelial H, histamine receptors and muscarinic cholinoceptors linked to the generation of a spasmolytic autacoid in both guinea pig trachea and human bronchus. The possibility that EpDIF acts as a bronchial circulation vasodilator and the significance of this to airway responsiveness to chemical stimuli in vivo requires investigation.
Receptors for Lipid-derived Mediators Prostanoids and Leukotrienes Phospholipase A 2 is a membrane-bound enzyme responsible for the mobilization of arachidonic acid and platelet-activating factor (PAF) from membrane phospholipids. The cyclooxygenase products of arachidonic acid metabolism POE, and POE2 are bronchodilators in normal and asthmatic subjects, al-
though constrictor responses have been observed (10). POF,a, POD 2, thromboxane A, and B2 contract human bronchial smooth muscle (11). In addition, bronchial obstruction results from the inhalation of POF2a in asthmatic and healthy volunteers. Leukotriene C. and D.. also contract human isolated bronchial preparations (12) and cause bronchoconstriction when inhaled by nonasthmatic and asthmatic subjects (13). However, there is no clear evidence for altered receptor distribution, number, or function in asthma.
Platelet-activating Factor PAF is an ether-linked glycerophospholipid generated in allergic reactions from lipid precursors found in abundance in the cell membrane. Platelets and various inflammatory cells, including eosinophils, basophils, neutrophils, and macrophages, can generate PAF in response to cell activation. PAF causes white cell and platelet migration, increases vascular permeability producing lung edema, and causes bronchoconstriction (14)and bronchial hyperreactivity to inhaled spasmogens in humans (15). These effects clearly suggest a significant role for PAF in the pathogenesis of asthma. Specific cell surface receptors for PAF have been detected in human lung membrane I From the Department of Pharmacology, University of Western Australia, Nedlands, Western Australia. 2 Supported by the National Health and Medical Research Council of Australia. J Correspondenceand requestsfor reprintsshould be addressed to Dr. R.G. Goldie, Department of Pharmacology, University of Western Australia, Perth, Nedlands 6009, Australia.
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Fig. 1. Responses to histamine (H = 100 11M) and methacholine (M = 25 11M) in phenylephrine (.A. = 0.05 11M) contracted endothelium-intact rat aorta (a), endothelium-denuded rat aorta (b), and endothelium-denuded rat aorta in coaxial assemblies with (c) epithelium-intact and (d) epithelium-denuded guinea pig tracheal tube segments. (Reproduced from Fernandes and colleagues [9] with permission.)
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preparations (16), although the distribution, localization, and relative densities in lung are not established. Wehave investigated the autoradiographic localization of specific binding sites for [3H]PAFin healthy and asthmatic human lung. However, the very high levels of nonspecific binding have thus far confounded these studies. The clinical efficacy of specific PAF antagonists in asthma is yet to be established, although the ginkolide mixture BN 52063 selectivelyinhibits the cutaneous late phase response to allergen in atopic subjects (17). p-Adrenoceptors p-Adrenoceptor agonist bronchodilators continue to be the mainstay of asthma therapy worldwide. Human bronchial smooth muscle contains a relatively pure population of p,-adrenoceptors (18, 19).Selectivep,-adrenoceptor agonists, such as salbutamol, terbutaline, and fenoterol, relax airway smooth muscle, reduce the release of chemical spasmogens from mast cells, improve mucociliary transport, decrease cholinergic neurotransmission, and reduce airway mucosal edema (20, 21). It has been suggested that the symptoms and pathophysiology of asthma could be explained in terms of reduced pulmonary p-adrenoceptor function (22). The status of p-adrenoceptor function in asthmatics has been extensively investigated since that time, but it remains controversial (20, 21). No significant difference in the potency of isoprenaline was observed between asthmatic and nonasthmatic bronchi obtained from lung tissue resected at surgery (23). Similarly, there
was no significant difference in p-adrenoceptor function in response to intravenous salbutamol between nonasthmatic volunteers and mild asthma sufferers (24). However, we have shown that both isoprenaline and fenoterol were approximately five times less potent in bronchi from subjects who had died during asthma attacks than in healthy human bronchi, whilethe potency of theophylline was similar in preparations from both groups (25). Hyporesponsiveness to ~agonists in two asthmatic subjects was apparently unrelated to previous exposure to p-agonists (25). These results suggest specific p-adrenoceptor hypofunction in bronchi from severely asthmatic lung and are consistent with similar studies using diseased human bronchi (26). Furthermore, a significant negative correlation was observed between the severity of asthma (as assessed by the level of baseline lung function) and the bronchodilator potency of inhaled salbutamol (27). This might have been due to reduced access of inhaled drug to p,-adrenoceptors or to functional antagonism between salbutamol and the progressively greater levels of airway tone present in more severely asthmatic individuals (20). However, the more severe asthmatics might also have been less sensitive to salbutamol as a result of bronchial 13-adrenoceptor dysfunction. In peripheral lymphocytes from allergic asthmatics,13-adrenoceptor/adenylate cyclase function was dramatically reduced 24 h after allergen challenge, while that in lymphocytes from healthy volunteers was unaffected (28). While radio ligand binding and autoradiographic studies have shown that the density
of 13-adrenoceptors associated with asthmatic bronchial smooth muscle and lung parenchyma was not reduced (29), decreased tissue sensitivity to 13-adrenoceptor agonists in asthma (25) may have resulted from impaired coupling between 13-adrenoceptors and adenylate cyclase associated with the inflammatory response in this disease (28). Activation of phospholipase A, is linked to both inflammatory changes in the airways and to diminished 13-adrenoceptor function (21, 30). Elevation of the activity of this enzyme may be an important cause of the reduction in pulmonary 13-adrenoceptor function observed in severe asthma. It has also been proposed that autoantibodies to 13,-adrenoceptors and the generation of oxygen free radicals during inflammation may cause 13-adrenoceptor hypofunction in asthmatics (20). The importance ofthese mechanisms still remains to be established. Studies both in vitro (23) and in vivo (24) have shown that asthmatics do not necessarily have diminished bronchial ~-adrenoceptor function. Furthermore, it is well established that blockade of 13-adrenoceptorsin nonasthmatics does not induce asthma or bronchial hyperresponsiveness (20, 21). Contrary to Szentivanyi's hypothesis, these findings indicate that 13-adrenoceptor dysfunction is not a fundamental cause of asthma. The marked worsening of asthma often produced by 13-blockers (21)illustrates the role of effective 13-adrenoceptor function in maintaining adequate airway caliber in asthmatics. Thus, significant 13-adrenoceptor hypofunction secondary to asthma may contribute to the further deterioration of this disease by rendering the subject more vulnerable to spasmogens and less responsive to 13-agonist therapy. a-Adrenoceptors Airway Smooth Muscle u-Adrenoceptor agonists have been shown to cause bronchial obstruction in asthmatics (31). This may be caused by stimulation of airway smooth muscle u-adrenoceptors. Early binding studies suggested increased a-adrenoceptor number in asthmatic lung (32). A twofold increase in pulmonary c-adrenoceptor density wasalso detected in a guinea pig model of asthma (33). However, phenylephrine-induced airway obstruction could only be demonstrated in asthmatics after l3-adrenoceptor blockade with propranolol (31). Prior to this treatment, phenylephrine caused marked bronchodilatation in these subjects but had no such effect in nonasthmatics. Thus, in the absence of l3-adrenoceptor blockade, u-adrenoceptor function was clearly subordinate to l3-adrenoceptor function even in asthmatics. In the presence of propranolol, adrenaline and noradrenaline (34) caused phentolaminesensitivecontractions of human isolated bronchial preparations. These responses were weak, being never greater than 10 to 20070 of maximal responses induced by carbachol or histamine. In addition, bronchi from subjects
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with lung disease (including pneumonia, chronic obstructive lung disease, and pulmonary fibrosis) contracted strongly in response to noradrenaline. These data suggest that airway u-adrenoceptor function was markedly enhanced in diseased lung. Conversely, our data indicates that asthma does not involve any significant increase in airway smooth muscle a-adrenoceptor activity. Indeed, only very weak contractions of bronchial preparations from both healthy and asthmatic human lung obtained postmortem were induced by noradrenaline or phenylephrine. Furthermore, these weak effects were almost never detected unless a 13-adrenoceptor antagonist was present (18, 35). Even then, the effect of the a-agonist was highly variable with no response occurring in most preparations (18). Certainly no evidence of asthma-induced enhancement of bronchial smooth muscle c-adrenoceptor function was found. However, responses of several other pulmonary tissues to c-agonists may alter airway caliber and airflow resistance in vivo. These include modification of neurotransmitter release, stimulation of airway mucus secretion, and enhancement of chemical mediator release from mast cells.
Autonomic Nerves In human airways, sympathetic postganglionic nerve endings terminate on ganglion cells of both the excitatory cholinergic system (36) and NANC inhibitory system (36, 37). In human isolated bronchial tissue, noradrenaline can inhibit parasympathetic neurotransmission via prejunctional a2-adrenoceptors (38). Thus, it is possible that presynaptic a 2-adrenoceptors can attenuate acetylcholine release from cholinergic nerves, substance P release from NANC excitatory nerves, noradrenaline release from sympathetic nerves, and neurotransmitter release from the NANC inhibitory system. Given the dominant role of the parasympathetic nervous system in the regulation of human airway caliber, the net effect of prejunctional aradrenoceptor stimulation should favor bronchodilatation rather than bronchoconstriction.
Secretory Glands In sharp contrast to airway smooth muscle, where u-adrenoceptors are sparse, submucosal glands are apparently well endowed with u-adrenoceptors (39). It is also established that stimulation of these receptors induces an increase in the secretion of glandular mucus into the airway lumen (40). In human bronchi, both a- and 13-adrenoceptor agonists have been shown to enhance total mucus output, although a-adrenoceptor function predominates (37, 40). This strongly suggests that glandular c-adrenoceptors are important in the physiologic control of secretory function in human lung. However, there is not evidence that a-adrenoceptor-mediated glandular function is greater in asthmatic than in healthy lung.
Mast Cells
Substance P
c-Adrenoceptors can function to facilitate the release of chemical mediators from human lung mast cells in vitro (41). However, in the absence of 13-adrenoceptor blockade, noradrenaline inhibited rather than enhanced mediator release, presumably as a result of predominant 13-adrenoceptor function (41). Thus, mast cell u-adrenoceptors are unlikely to be important in the modulation of mediator release when exposed to physiologic concentrations of adrenaline or noradrenaline in vivo. Furthermore, there is no evidence for an increase in u-adrenoceptor numbers or function in mast cells from asthmatic lung.
Sensory neuropeptides such as substance P (SP), neurokinin (NK) A, and calcitonin gene-related peptide (CGRP) have a variety of effects that suggest a role in asthma, including contraction of airway smooth muscle, stimulation of mucosal gland secretion, and increasing mucosal vascular permeability. However, no direct evidence for such a role has been obtained (10). SP-containing afferent sensory nerves have been found in both animal and human airways (47). Since this peptide can contract airway smooth muscle, enhance bronchial mucosal edema, and stimulate mucous secretion, it may be the neurotransmitter for a NANC excitatory nervous system in human lung (37). However, while SP-containing nerves appear to be functionally important in the guinea pig trachea (48), the majority of human isolated bronchi tested did not contain such nerves (49). Thus, SP-containing nerves do not appear to playa major role in the regulation of airway tone in man. Furthermore, SP was almost 1,000 times less potent in human isolated airway preparations than in guinea pig trachea (49). The autoradiographic distribution of receptors for 125I-labeled Bolton-Hunter SP (I-SP) has recently been described in guinea pig and human lung (50). This study indicated high densities of specific binding sites for I-SP in airway smooth muscle from the trachea to small bronchioles, with lesser binding associated with vascular smooth muscle and epithelium. We have also demonstrated high levels of specific I-SP binding associated with guinea pig bronchial smooth muscle and bronchial epithelium (figure 2A). Relatively high levels of binding were also detected over vascular muscle, with virtually no binding detected over alveolar septa. In sharp contrast, in both healthy and asthmatic human bronchi, I-SP binding was very sparse over airway smooth muscle and epithelium, with no evidence for differential binding capacity between healthy and asthmatic tissue (figure 2, D and G). I-SP binding was most dense over tissue structures immediately beneath the airway epithelium and over tissue beneath the airway smooth muscle. Some binding was also associated with deep submucosal glands. I-SP binding was virtually absent in human alveolar wall tissue and peripheral blood vessels. Less specific I-SP binding in the asthmatic bronchus than in the healthy bronchus (figure 2D) may indicate down-regulation of SP receptors in the diseased airway. While speculative, such down-regulation is consistent with the concept of excessive stimulation ot SP receptors following epithelial damage and the exposure of sensory nerve endings forming part of airway axon reflex loops (10, 37). However, these data do not support the view that SP might have a significant spasmogenic role in asthmatic or healthy bronchi, although an influence on airway secretions is suggested. Once again, the dangers of extending fmdings from animal data to humans are highlighted.
Binding and Autoradiographic Studies We have studied the distribution of c-adrenoceptors in nondiseased human lung using the a,-selective radioligands l2SI-labeled BE2254 and [3H]prazosin. Only very low levels of specific binding were detected throughout the lung. Binding in bronchial and bronchiolar tissue was no more intense than at other sites. The low density of a-ligand binding contrasted sharply with the high densities of 13-adrenoceptors detected using l2SI_ labeled iodocyanopindolol in bronchiolar and alveolar tissue in adjacent tissue sections from the same lung. Our functional and autoradiographic data are consistent with the dominance of 13- over e-adrenoceptor function in human lung. They also emphasize that extrapolations of data from animal lung studies to man must be made with great care. At present the evidence does not support the concept of a major role for c-adrenoceptors in the pathophysiology of asthma.
Neuropeptides
Vasoactive Intestinal Peptide Experiments in vitro have demonstrated the existence of a NANC inhibitory system in human airway smooth muscle (42). Vasoactive intestinal peptide (VIP) is favored as the likely inhibitory neurotransmitter, although this view has been questioned. It is possible that defective NANC inhibitory nerve function in asthma could lead to elevated bronchial tone (37), although in the absence of supportive experimental data, this suggestion is purely speculative. Autoradiography has revealed VIP receptors in proximal airway smooth muscle, epithelium, and submucosal glands in both guinea pig and human lung, although no receptors were detected in bronchioles (43). The functional significance of the NANC inhibitory system in human airways is also unclear in view of evidence demonstrating much smaller effects in human bronchi than in guinea pig trachea (44). Furthermore, inhaled (45) and infused VIP produced only weak bronchodilatation in both asthmatic and nonasthmatic subjects (46), although some protection against histamine-induced bronchoconstriction was observed (45).
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RECEPTORS IN ASTHMATIC AIRWAYS
Conclusions The nonspecific bronchial hyperreactivity of asthma does not seem to involve increased airway smooth muscle contractility. Rather, alterations in systemscontrolling airway tone, mucosal vascular and epithelial function, and airway secretions seem to play a vital role. Airway u-adrenoceptor function is of little significance either in healthy or asthmatic bronchi. However, reduced ~-adrenoceptor function can be detected in severe asthma, probably resulting from widespread and intense inflammatory responses in the lung. Clearly, this is not causally linked to asthma but may be critical to the maintenance of airway caliber in severe disease. A growing number of chemical substances have been suggested as putative mediators of asthma symptoms. However, the pharmacologic profile of no single substance can completely account for the altered airway function seen in this disease. Thus, it seems that several mediators and their interactions will prove to be important to the pathophysiology of asthma. Epithelial desquamation resulting from airway inflammation is also clearly a major factor influencing airwayfunction in asthma since this exposes sensory afferent nerves perhaps resulting in neurogenic inflammation and bronchial obstruction, as well as the loss of protective EpDIF. The striking differences between human airway and guinea pig airway tissue, with respect to the distribution of specific binding sites for I-SP, highlight the potential dangers of extrapolating conclusions derived from animal modelsto human airway systems. While animal studies continue to provide important insights into respiratory function in health and disease, more data must be derived from human lung, and from asthmatic airways in particular, before these issues can be finally resolved. References 1. Robinson C, Holgate ST. Mast cell dependent inflammatory mediators and their putative role in bronchial asthma. Clin Sci 1985; 68:103-12. 2. Goldie RG, Spina D, Henry PJ, Lulich KM, Paterson JW. In vitro responsivenessof human asthmatic bronchus to carbachol, histamine, l3-adrenoceptor agonists and theophylline. Br J Clin Pharmacol 1986; 22:669-76. 3. Sert! K, Casale TB, Wescott SL, Kaliner MA. Immunohistochemical localization of histaminestimulated increases in cyclic GMP in guinea pig lung. Am Rev Respir Dis 1987; 135:456-62. 4. White JP, Mills J, Eiser NM. Comparison of the effects of histamine HI and Hj-receptor agonists on large and small airwaysin normal and asthmatic subjects. Br J Dis Chest 1987; 81:155-69. 5. Laitinen LA, Heino M, Laitinen A, Kava T, Haahtels T. Damage of airwayepithelium and bron-
5155 chial reactivity in patients with asthma. Am Rev 24. Tattersfield AE, Holgate ST, Harvey JE, Gribbin HR. Is asthma due to partial l3-blockade of Respir Dis 1985; 131:599-606. 6. Flavahan NA, Aarhus LL, Rimele TJ, Van- airways? Agents Actions [Suppl] 1983; 13:265-71. houtte PM. Respiratory epithelium inhibits bron- 25. Goldie RG, Spina D, Henry PJ, Lulich KM, chial smooth muscle tone. J Appl Physiol 1985; Paterson JW. In vitro responsivenessof human asth8:834-8. matic bronchus to carbachol, histamine, l3-adre7. Goldie RG, Papadimitriou JM, Paterson JW, noceptor agonists and theophylline. Br J Clin PharRigby PJ, Self HM, Spina D. Influence of the epi- macol 1986; 22:669-76. thelium on responsiveness of guinea-pig trachea 26. Cerrina J, Ladurie ML, Labat C, Raffestin to contractile and relaxant agonists. Br J Pharmacol B, Bayol A, Brink C. Comparison of human bronchial muscle responses to histamine in vivo with 1986; 87:5-14. 8. Woolcock AJ, Salome C, Yan K. The shape of histamine and isoproterenol agonists in vitro. Am the dose-response curve to histamine in asthmatic Rev Respir Dis 1986; 134:57-61. and normal subjects. Am Rev Respir Dis 1984; 27. Barnes PJ, Pride NB. Dose-response curves to inhaled l3-adrenoceptor agonists in normal and 13:71-5. 9. Fernandes LB, Paterson JW, Goldie RG. Coasthmatic subjects. Br J Clin Pharmacol 1983; axial bioassay of an epithelium-derived inhibitory 15:677-82. factor (EpDIF) from guinea trachea. Br J Phar- 28. Meurs H, Koeter GH, De Vries K, Kauffman HE The l3-adrenergicsystem and allergic bronchial macol 1989; 96:117-24 10. BarnesPJ,ChungKF,PageCPo Inflammatory me- asthma: changes in lymphocyte l3-adrenergic recepdiators and asthma. Pharmacol Rev 1988; 40:49-84. tor number and adenylate cyclase activity after an allergen-induced asthmatic attack. J Allergy Clin 11. Gardiner PJ, Collier HOJ. Specific receptors for prostaglandins in airways. Prostaglandins 1980; Immunol 1982; 70:272-80. 19:819-41. 29. Goldie RG, Spina D, Lulich KM, Paterson Jw. 12. Hanna CJ, Bach MK, Pare PD, Schellenberg The status of l3-adrenoceptor function in asthma. RR. Slowreacting substance (Ieukotrienes) contract Pharmacology: Excerpta Medica Congress Series human airway and pulmonary vascular smooth 1987; 750:465-8. muscle in vitro. Nature 1981; 290:343-4. 30. TakiF, TakagiK, Satake T, SugiyamaS, Ozawa 13. Barnes NC, Piper PJ, Costello JE Compara- T. The role of phospholipasein reduced l3-adrenergic tive effects of inhaled leukotriene C., leukotriene responsiveness in experimental asthma. Am Rev Respir Dis 1986; 133:362-6. D. and histamine in normal human subjects. Tho31. Patel KR, Kerr JW. The airways response to rax 1984; 39:500-4. 14. PageCP, ArcherCB, Paul W, Morley J. Paf- phenylephrine after blockade of a and l3-receptors acether: a mediator of inflammation in asthma. in extrinsic bronchial asthma. Clin Allergy 1973; 3:439-48. Trends Pharmacol Sci 1984; 5:239-41. 15. Cuss FM, Dixon CMS, Barnes PJ. Effects of 32. Szentivanyi A. The radio ligand binding apinhaled platelet activating factor on pulmonary proach in the study of lymphocytic adrenoceptors function and bronchial responsivenessin man. Lan- and the constitutional basis of atopy. J Allergy Clin cet 1986; 2:189-92. Immunol 1980; 65:5-11. 16. Hwang SB, Lam MH, Shen TY. Specific bind- 33. Barnes PJ, Dollery CT, MacDermot J. Ining sites for platelet activating factor in human lung creased pulmonary a-adrenergic and reduced l3-adtissues. Biochem Biophys Res Commun 1985; 128: renergic receptors in experimental asthma. Nature 1980; 285:569-71. 972-9. 17. Chung KF, Dent G, McCusker M, Guinot P, 34. Kneussl MP, Richardson JB. Alpha-adrenergic Page CP, Barnes PJ. Effect of a ginkolide mixture receptors in human and canine tracheal and bron(BN 52063) in antagonizing skin and platelet re- chial smooth muscle. J Appl Physiol 1978; 45: sponses to platelet activating factor in man. Lan- 307-11. 35. Goldie RG, Lulich KM, Paterson JW. Broncet 1987; 1:248-51. 18. Goldie RG, Paterson JW, Spina D, Wale J. chial c-adrenoceptor function in asthma. Trends Classification of l3-adrenoceptors in human isolated Pharmacol Sci 1985; 6:469-72. bronchus. Br J Pharmacol 1984; 81:611-5. 36. Richardson JB. Nerve supply to the lungs. Am 19. Zaagsma J, Van der Heijden PJ, Van der Rev Respir Dis 1979; 119:785-802. Schaar MW, Bank CM. Differentiation of func37. Barnes PJ. The third nervous system in the tional adrenoceptors in human and guinea-pig air- lung: physiology and clinical perspectives. Thorax ways. Eur J Respir Dis 1984;65(Suppl 135:16-33). 1984; 39:561-7. 20. Barnes PJ. Neural control of human airways 38. Grundstrom N, Andersson RGG. Inhibition in health and disease. Am Rev Respir Dis 1986; of the cholinergic neurotransmission in human airwaysvia prejunctional a,-adrenoceptors. Acta Phys134:1289-314. 21. Paterson JW, Lulich KM, Goldie RG. Drug iol Scand 1985; 125:513-7. effects on l3-adrenoceptor function in asthma. In: 39. Barnes PJ. Localization and functionof airMorley J, ed. Perspectives in asthma. 2.I3-adreno- way autonomic receptors. Eur J Respir Dis 1984; ceptors in asthma. London: Academic Press, 1984; 65(Suppl 135:187-97). 40. Phipps RJ, Williams IP, Richardson PS, Pell 245-68. 22. Szentivanyi A. The l3-adrenergictheory of the J, Pack RJ, Wright N. Sympathomimetic drugs atopic abnormality in bronchial asthma. J Allergy stimulate the output of secretory glycoproteinsfrom human bronchi in vitro. Clin Sci 1982; 63:23-8. 1968; 42:203-32. 23. SvedrnyrNLV, Larsson SA, Thiringer GK. De- 41. Kaliner M, Orange RP, Austen KE Immunovelopment of "resistance" in l3-adrenergicreceptors logical release of histamine and slow reacting substance of anaphylaxis from human lung. J Exp Med of asthmatic patients. Chest 1976; 69:479-83.
Fig. 2. Autoradiographic distribution of binding sites for 1lSl-labeled Bolton Hunter Substance P (I-SP; 0.2 nM) in guinea pig lung (A-C), nondiseased human bronchus (D-F), and asthmatic human bronchus (G-I). Plates A, D, and G are dark-field photomicrographs showing totall-SP binding in the frozen tissue sections shown in light-field photomicrographs B, E, and H, respectively. Note that the smooth muscle (SM) and epithelium (Epi) of the guinea pig bronchus (Br) and bronchiole (BI), as well as vascular (V) tissue, were heavily labeled. In sharp contrast, SM and Epi were very sparsely labeled as were submucosal glands (GI) and cartilage (C) in both nondiseased and asthmatic bronchi. However, there is intense labeling of submucosal tissue surrounding these structures in human bronchi from both sources. Nonspecific I-SP binding in respective adjacent sections was determined in the presence of 1 11M unlabeled SP (plates C, F, and I). Bar = 100 11m.
5156 1972; 136:556-67. 42. Davis C, Karman MS, Jones TR, Daniel EE.
Control of human airway smooth muscle: in vitro studies. J Appl Physiol 1982; 53:1080-7. 43. Carstairs JR, Barnes PJ. Visualization of vasoactive intestinal peptide receptors in human and guinea-pig lung. J Pharmacol Exp Ther 1986; 239:249-55.
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45. Barnes PJ, Dixon CMS. The effects of inhaled
vasoactive intestinal peptide on bronchial hyperreactivity in man. Am Rev Respir Dis 1984; 130: 162-6. 46. Morice A, Unwin RJ, Sever PS. Vasoactive
intestinalpeptide causesbronchodilatation and protects against histamine-induced bronchoconstriction in asthmatic subjects. Lancet 1983; 1225-6. 47. Polack JM, Bloom SR. Regulatory peptides in the respiratory tract of man and other animals. Exp Lung Res 1982; 3:313-28. 48. Lundberg JM, Saria A, Brodin E, Rosell S,
Folkers K. A substance P antagonist inhibits vagally induced increase in vascular permeability and bronchial smooth muscle contraction in the guinea pig. Proc Nat! Acad Sci USA 1983; 80:1120-4. 49. Lundberg JM, Mart!ing C-R, Saria A. Substance P and capsaicin-induced contraction of human bronchi. ActaPhysiol Scand 1983; 119:49-53. 50. Carstairs JR, Barnes PJ. Autoradiographic mapping of substance P receptors in lung. Eur J Pharmacol 1986; 127:295-6.
Introduction Asthma is an inflammatory lung disease involving reversible airway obstruction. The characteristic features of asthma include eosinophilia, nonspecific bronchial hyperreactivity to inhaled spasmogens, airway epithelial damage, mucosal edema, and mucosal gland hypersecretion. Examining the role of inflammatory mediators in these processes is critical to an understanding of the causes of asthma. A major element in such an assessment is the study of the distribution, density, and function of receptors for such chemicals in asthmatics compared with that in healthy human lung. Histamine Receptors Inhalation of allergens causes significant degranulation of mast cells in the bronchial wall in allergic asthmatics with release of histamine and several other putative mediators of asthma, including prostaglandin (PO) D, and sulfidoleukotrienes, These mediators may increase airway smooth muscle tone and promote mucosal edema and glandular secretion, resulting in narrowing of the airways and limiting airflow. This is seen in the immediate bronchial response in asthma (1).
Airway Smooth Muscle The causes of asthma-associated bronchial hyperreactivity to spasmogens such as histamine are unknown. Recent evaluations in vitro of the responsiveness of human bronchial preparations obtained postmortem from healthy and asthmatic lung to histamine and to carbachol indicate that no major intrinsic abnormality exists with respect to airway smooth muscle contractility. Indeed the potencies of these spasmogens were marginally lower in asthmatic bronchi than in tissue from healthy lung (2), while maximal responsiveness was similar in bronchi from both sources. These data also indicate that histamine H I receptor and muscarinic cholinoceptor function were not up-regulated in bronchial smooth muscle in asthma. Interestingly, histamine H I receptors have been detected using immunohistochemical techniques in greater numbers in airway epithelial cells than in either guinea pig airway or vascular smooth muscle (3). Histamine Hj-receptors might be expected to mediate relaxation of human airway smooth muscle since they are linked to the generation of cyclic adenosine monophosphate (cAMP). However, there is little evidence for their existence in functionally significant numbers in either normal or asthmatic subjects (4). No role for histamine Hs-receptors in airways has yet been established. Bronchial Epithelium Desquamation of the bronchial epithelium
SUMMARY Airway tissue obtained postmortem from nondiseased and severely asthmatic human lung was used in functional, radlollgand binding and autoradiographic studies to investigate aspects of various receptor systems for putative mediators of asthma. Results indicate that asthma does not involve an intrinsic abnormality of smooth muscle contractility to spasmogens. Nor was there any evidence for up-regulation of histamine H,-receptor, muscarinic cholinoceptor, or a,·adrenoceptor function. Conversely, severe asthma involving intense airway inflammation resulted in significant I}-adrenoceptor dysfunction, probably caused by receptor uncoupling from adenylate cyclase. Evidence was also obtained for histamine and methacholine-induced release of a nonprostanoid, airway epithelium-derived Inhibitory factor (EpDIF), which may have a significant dilator effect In the adjacent bronchial circulation. The activity of this potentially protective Inhibitory autacoid system would be expected to be reduced in asthma, which commonly involves epithelium damage. The autoradiographic distribution of specific binding sites for "'I-labeled substance P (I-SP) was also determined in bronchi from healthy humans and asthmatics. In sharp contrast to guinea pig airways where high levels of binding were detected over smooth muscle, specific I-SP binding was sparse over human airway smooth muscle from both sources, while dense binding was associated with structures immediately beneath the epithelium and with deep submucosal glands. These data suggest a more significant role for SP in secretory processes than In spasmogenic processes in human bronchi and highlight the potential for drawing Invalid conclusions concerning human airway function from studies using animal models. AM REV RE5PIR DIS 1990; 141:5151-5156
is a consistent and striking feature in asthma (5) and may be causally related to bronchial hyperreactivity. Recent studies have demonstrated that deliberate removal of the airway epithelium causes increased smooth muscle responsiveness to histamine, providing indirect evidence for the existence of an epithelium-derived inhibitory factor (EpDlF), generated in response to some spasmogens (6, 7). In nonasthmatics, bronchial reactivity to inhaled histamine is limited, while asthmatics respond as if lacking a braking mechanism opposing bronchoconstriction (8). EpDlF activity, which might be absent or reduced in asthma as a result of epithelial damage, may contribute to such a braking mechanism in healthy lung. Direct evidence for EpDIF release from guinea pig trachea and from human bronchus in response to histamine and methacholine (9) has now been demonstrated in a coaxial bioassay system (figure 1). These data also indicate the existence of epithelial H, histamine receptors and muscarinic cholinoceptors linked to the generation of a spasmolytic autacoid in both guinea pig trachea and human bronchus. The possibility that EpDIF acts as a bronchial circulation vasodilator and the significance of this to airway responsiveness to chemical stimuli in vivo requires investigation.
Receptors for Lipid-derived Mediators Prostanoids and Leukotrienes Phospholipase A 2 is a membrane-bound enzyme responsible for the mobilization of arachidonic acid and platelet-activating factor (PAF) from membrane phospholipids. The cyclooxygenase products of arachidonic acid metabolism POE, and POE2 are bronchodilators in normal and asthmatic subjects, al-
though constrictor responses have been observed (10). POF,a, POD 2, thromboxane A, and B2 contract human bronchial smooth muscle (11). In addition, bronchial obstruction results from the inhalation of POF2a in asthmatic and healthy volunteers. Leukotriene C. and D.. also contract human isolated bronchial preparations (12) and cause bronchoconstriction when inhaled by nonasthmatic and asthmatic subjects (13). However, there is no clear evidence for altered receptor distribution, number, or function in asthma.
Platelet-activating Factor PAF is an ether-linked glycerophospholipid generated in allergic reactions from lipid precursors found in abundance in the cell membrane. Platelets and various inflammatory cells, including eosinophils, basophils, neutrophils, and macrophages, can generate PAF in response to cell activation. PAF causes white cell and platelet migration, increases vascular permeability producing lung edema, and causes bronchoconstriction (14)and bronchial hyperreactivity to inhaled spasmogens in humans (15). These effects clearly suggest a significant role for PAF in the pathogenesis of asthma. Specific cell surface receptors for PAF have been detected in human lung membrane I From the Department of Pharmacology, University of Western Australia, Nedlands, Western Australia. 2 Supported by the National Health and Medical Research Council of Australia. J Correspondenceand requestsfor reprintsshould be addressed to Dr. R.G. Goldie, Department of Pharmacology, University of Western Australia, Perth, Nedlands 6009, Australia.
S151
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Fig. 1. Responses to histamine (H = 100 11M) and methacholine (M = 25 11M) in phenylephrine (.A. = 0.05 11M) contracted endothelium-intact rat aorta (a), endothelium-denuded rat aorta (b), and endothelium-denuded rat aorta in coaxial assemblies with (c) epithelium-intact and (d) epithelium-denuded guinea pig tracheal tube segments. (Reproduced from Fernandes and colleagues [9] with permission.)
•
preparations (16), although the distribution, localization, and relative densities in lung are not established. Wehave investigated the autoradiographic localization of specific binding sites for [3H]PAFin healthy and asthmatic human lung. However, the very high levels of nonspecific binding have thus far confounded these studies. The clinical efficacy of specific PAF antagonists in asthma is yet to be established, although the ginkolide mixture BN 52063 selectivelyinhibits the cutaneous late phase response to allergen in atopic subjects (17). p-Adrenoceptors p-Adrenoceptor agonist bronchodilators continue to be the mainstay of asthma therapy worldwide. Human bronchial smooth muscle contains a relatively pure population of p,-adrenoceptors (18, 19).Selectivep,-adrenoceptor agonists, such as salbutamol, terbutaline, and fenoterol, relax airway smooth muscle, reduce the release of chemical spasmogens from mast cells, improve mucociliary transport, decrease cholinergic neurotransmission, and reduce airway mucosal edema (20, 21). It has been suggested that the symptoms and pathophysiology of asthma could be explained in terms of reduced pulmonary p-adrenoceptor function (22). The status of p-adrenoceptor function in asthmatics has been extensively investigated since that time, but it remains controversial (20, 21). No significant difference in the potency of isoprenaline was observed between asthmatic and nonasthmatic bronchi obtained from lung tissue resected at surgery (23). Similarly, there
was no significant difference in p-adrenoceptor function in response to intravenous salbutamol between nonasthmatic volunteers and mild asthma sufferers (24). However, we have shown that both isoprenaline and fenoterol were approximately five times less potent in bronchi from subjects who had died during asthma attacks than in healthy human bronchi, whilethe potency of theophylline was similar in preparations from both groups (25). Hyporesponsiveness to ~agonists in two asthmatic subjects was apparently unrelated to previous exposure to p-agonists (25). These results suggest specific p-adrenoceptor hypofunction in bronchi from severely asthmatic lung and are consistent with similar studies using diseased human bronchi (26). Furthermore, a significant negative correlation was observed between the severity of asthma (as assessed by the level of baseline lung function) and the bronchodilator potency of inhaled salbutamol (27). This might have been due to reduced access of inhaled drug to p,-adrenoceptors or to functional antagonism between salbutamol and the progressively greater levels of airway tone present in more severely asthmatic individuals (20). However, the more severe asthmatics might also have been less sensitive to salbutamol as a result of bronchial 13-adrenoceptor dysfunction. In peripheral lymphocytes from allergic asthmatics,13-adrenoceptor/adenylate cyclase function was dramatically reduced 24 h after allergen challenge, while that in lymphocytes from healthy volunteers was unaffected (28). While radio ligand binding and autoradiographic studies have shown that the density
of 13-adrenoceptors associated with asthmatic bronchial smooth muscle and lung parenchyma was not reduced (29), decreased tissue sensitivity to 13-adrenoceptor agonists in asthma (25) may have resulted from impaired coupling between 13-adrenoceptors and adenylate cyclase associated with the inflammatory response in this disease (28). Activation of phospholipase A, is linked to both inflammatory changes in the airways and to diminished 13-adrenoceptor function (21, 30). Elevation of the activity of this enzyme may be an important cause of the reduction in pulmonary 13-adrenoceptor function observed in severe asthma. It has also been proposed that autoantibodies to 13,-adrenoceptors and the generation of oxygen free radicals during inflammation may cause 13-adrenoceptor hypofunction in asthmatics (20). The importance ofthese mechanisms still remains to be established. Studies both in vitro (23) and in vivo (24) have shown that asthmatics do not necessarily have diminished bronchial ~-adrenoceptor function. Furthermore, it is well established that blockade of 13-adrenoceptorsin nonasthmatics does not induce asthma or bronchial hyperresponsiveness (20, 21). Contrary to Szentivanyi's hypothesis, these findings indicate that 13-adrenoceptor dysfunction is not a fundamental cause of asthma. The marked worsening of asthma often produced by 13-blockers (21)illustrates the role of effective 13-adrenoceptor function in maintaining adequate airway caliber in asthmatics. Thus, significant 13-adrenoceptor hypofunction secondary to asthma may contribute to the further deterioration of this disease by rendering the subject more vulnerable to spasmogens and less responsive to 13-agonist therapy. a-Adrenoceptors Airway Smooth Muscle u-Adrenoceptor agonists have been shown to cause bronchial obstruction in asthmatics (31). This may be caused by stimulation of airway smooth muscle u-adrenoceptors. Early binding studies suggested increased a-adrenoceptor number in asthmatic lung (32). A twofold increase in pulmonary c-adrenoceptor density wasalso detected in a guinea pig model of asthma (33). However, phenylephrine-induced airway obstruction could only be demonstrated in asthmatics after l3-adrenoceptor blockade with propranolol (31). Prior to this treatment, phenylephrine caused marked bronchodilatation in these subjects but had no such effect in nonasthmatics. Thus, in the absence of l3-adrenoceptor blockade, u-adrenoceptor function was clearly subordinate to l3-adrenoceptor function even in asthmatics. In the presence of propranolol, adrenaline and noradrenaline (34) caused phentolaminesensitivecontractions of human isolated bronchial preparations. These responses were weak, being never greater than 10 to 20070 of maximal responses induced by carbachol or histamine. In addition, bronchi from subjects
5153
RECEPTORS IN ASTHMATIC AIRWAYS
with lung disease (including pneumonia, chronic obstructive lung disease, and pulmonary fibrosis) contracted strongly in response to noradrenaline. These data suggest that airway u-adrenoceptor function was markedly enhanced in diseased lung. Conversely, our data indicates that asthma does not involve any significant increase in airway smooth muscle a-adrenoceptor activity. Indeed, only very weak contractions of bronchial preparations from both healthy and asthmatic human lung obtained postmortem were induced by noradrenaline or phenylephrine. Furthermore, these weak effects were almost never detected unless a 13-adrenoceptor antagonist was present (18, 35). Even then, the effect of the a-agonist was highly variable with no response occurring in most preparations (18). Certainly no evidence of asthma-induced enhancement of bronchial smooth muscle c-adrenoceptor function was found. However, responses of several other pulmonary tissues to c-agonists may alter airway caliber and airflow resistance in vivo. These include modification of neurotransmitter release, stimulation of airway mucus secretion, and enhancement of chemical mediator release from mast cells.
Autonomic Nerves In human airways, sympathetic postganglionic nerve endings terminate on ganglion cells of both the excitatory cholinergic system (36) and NANC inhibitory system (36, 37). In human isolated bronchial tissue, noradrenaline can inhibit parasympathetic neurotransmission via prejunctional a2-adrenoceptors (38). Thus, it is possible that presynaptic a 2-adrenoceptors can attenuate acetylcholine release from cholinergic nerves, substance P release from NANC excitatory nerves, noradrenaline release from sympathetic nerves, and neurotransmitter release from the NANC inhibitory system. Given the dominant role of the parasympathetic nervous system in the regulation of human airway caliber, the net effect of prejunctional aradrenoceptor stimulation should favor bronchodilatation rather than bronchoconstriction.
Secretory Glands In sharp contrast to airway smooth muscle, where u-adrenoceptors are sparse, submucosal glands are apparently well endowed with u-adrenoceptors (39). It is also established that stimulation of these receptors induces an increase in the secretion of glandular mucus into the airway lumen (40). In human bronchi, both a- and 13-adrenoceptor agonists have been shown to enhance total mucus output, although a-adrenoceptor function predominates (37, 40). This strongly suggests that glandular c-adrenoceptors are important in the physiologic control of secretory function in human lung. However, there is not evidence that a-adrenoceptor-mediated glandular function is greater in asthmatic than in healthy lung.
Mast Cells
Substance P
c-Adrenoceptors can function to facilitate the release of chemical mediators from human lung mast cells in vitro (41). However, in the absence of 13-adrenoceptor blockade, noradrenaline inhibited rather than enhanced mediator release, presumably as a result of predominant 13-adrenoceptor function (41). Thus, mast cell u-adrenoceptors are unlikely to be important in the modulation of mediator release when exposed to physiologic concentrations of adrenaline or noradrenaline in vivo. Furthermore, there is no evidence for an increase in u-adrenoceptor numbers or function in mast cells from asthmatic lung.
Sensory neuropeptides such as substance P (SP), neurokinin (NK) A, and calcitonin gene-related peptide (CGRP) have a variety of effects that suggest a role in asthma, including contraction of airway smooth muscle, stimulation of mucosal gland secretion, and increasing mucosal vascular permeability. However, no direct evidence for such a role has been obtained (10). SP-containing afferent sensory nerves have been found in both animal and human airways (47). Since this peptide can contract airway smooth muscle, enhance bronchial mucosal edema, and stimulate mucous secretion, it may be the neurotransmitter for a NANC excitatory nervous system in human lung (37). However, while SP-containing nerves appear to be functionally important in the guinea pig trachea (48), the majority of human isolated bronchi tested did not contain such nerves (49). Thus, SP-containing nerves do not appear to playa major role in the regulation of airway tone in man. Furthermore, SP was almost 1,000 times less potent in human isolated airway preparations than in guinea pig trachea (49). The autoradiographic distribution of receptors for 125I-labeled Bolton-Hunter SP (I-SP) has recently been described in guinea pig and human lung (50). This study indicated high densities of specific binding sites for I-SP in airway smooth muscle from the trachea to small bronchioles, with lesser binding associated with vascular smooth muscle and epithelium. We have also demonstrated high levels of specific I-SP binding associated with guinea pig bronchial smooth muscle and bronchial epithelium (figure 2A). Relatively high levels of binding were also detected over vascular muscle, with virtually no binding detected over alveolar septa. In sharp contrast, in both healthy and asthmatic human bronchi, I-SP binding was very sparse over airway smooth muscle and epithelium, with no evidence for differential binding capacity between healthy and asthmatic tissue (figure 2, D and G). I-SP binding was most dense over tissue structures immediately beneath the airway epithelium and over tissue beneath the airway smooth muscle. Some binding was also associated with deep submucosal glands. I-SP binding was virtually absent in human alveolar wall tissue and peripheral blood vessels. Less specific I-SP binding in the asthmatic bronchus than in the healthy bronchus (figure 2D) may indicate down-regulation of SP receptors in the diseased airway. While speculative, such down-regulation is consistent with the concept of excessive stimulation ot SP receptors following epithelial damage and the exposure of sensory nerve endings forming part of airway axon reflex loops (10, 37). However, these data do not support the view that SP might have a significant spasmogenic role in asthmatic or healthy bronchi, although an influence on airway secretions is suggested. Once again, the dangers of extending fmdings from animal data to humans are highlighted.
Binding and Autoradiographic Studies We have studied the distribution of c-adrenoceptors in nondiseased human lung using the a,-selective radioligands l2SI-labeled BE2254 and [3H]prazosin. Only very low levels of specific binding were detected throughout the lung. Binding in bronchial and bronchiolar tissue was no more intense than at other sites. The low density of a-ligand binding contrasted sharply with the high densities of 13-adrenoceptors detected using l2SI_ labeled iodocyanopindolol in bronchiolar and alveolar tissue in adjacent tissue sections from the same lung. Our functional and autoradiographic data are consistent with the dominance of 13- over e-adrenoceptor function in human lung. They also emphasize that extrapolations of data from animal lung studies to man must be made with great care. At present the evidence does not support the concept of a major role for c-adrenoceptors in the pathophysiology of asthma.
Neuropeptides
Vasoactive Intestinal Peptide Experiments in vitro have demonstrated the existence of a NANC inhibitory system in human airway smooth muscle (42). Vasoactive intestinal peptide (VIP) is favored as the likely inhibitory neurotransmitter, although this view has been questioned. It is possible that defective NANC inhibitory nerve function in asthma could lead to elevated bronchial tone (37), although in the absence of supportive experimental data, this suggestion is purely speculative. Autoradiography has revealed VIP receptors in proximal airway smooth muscle, epithelium, and submucosal glands in both guinea pig and human lung, although no receptors were detected in bronchioles (43). The functional significance of the NANC inhibitory system in human airways is also unclear in view of evidence demonstrating much smaller effects in human bronchi than in guinea pig trachea (44). Furthermore, inhaled (45) and infused VIP produced only weak bronchodilatation in both asthmatic and nonasthmatic subjects (46), although some protection against histamine-induced bronchoconstriction was observed (45).
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5154
RECEPTORS IN ASTHMATIC AIRWAYS
Conclusions The nonspecific bronchial hyperreactivity of asthma does not seem to involve increased airway smooth muscle contractility. Rather, alterations in systemscontrolling airway tone, mucosal vascular and epithelial function, and airway secretions seem to play a vital role. Airway u-adrenoceptor function is of little significance either in healthy or asthmatic bronchi. However, reduced ~-adrenoceptor function can be detected in severe asthma, probably resulting from widespread and intense inflammatory responses in the lung. Clearly, this is not causally linked to asthma but may be critical to the maintenance of airway caliber in severe disease. A growing number of chemical substances have been suggested as putative mediators of asthma symptoms. However, the pharmacologic profile of no single substance can completely account for the altered airway function seen in this disease. Thus, it seems that several mediators and their interactions will prove to be important to the pathophysiology of asthma. Epithelial desquamation resulting from airway inflammation is also clearly a major factor influencing airwayfunction in asthma since this exposes sensory afferent nerves perhaps resulting in neurogenic inflammation and bronchial obstruction, as well as the loss of protective EpDIF. The striking differences between human airway and guinea pig airway tissue, with respect to the distribution of specific binding sites for I-SP, highlight the potential dangers of extrapolating conclusions derived from animal modelsto human airway systems. While animal studies continue to provide important insights into respiratory function in health and disease, more data must be derived from human lung, and from asthmatic airways in particular, before these issues can be finally resolved. References 1. Robinson C, Holgate ST. Mast cell dependent inflammatory mediators and their putative role in bronchial asthma. Clin Sci 1985; 68:103-12. 2. Goldie RG, Spina D, Henry PJ, Lulich KM, Paterson JW. In vitro responsivenessof human asthmatic bronchus to carbachol, histamine, l3-adrenoceptor agonists and theophylline. Br J Clin Pharmacol 1986; 22:669-76. 3. Sert! K, Casale TB, Wescott SL, Kaliner MA. Immunohistochemical localization of histaminestimulated increases in cyclic GMP in guinea pig lung. Am Rev Respir Dis 1987; 135:456-62. 4. White JP, Mills J, Eiser NM. Comparison of the effects of histamine HI and Hj-receptor agonists on large and small airwaysin normal and asthmatic subjects. Br J Dis Chest 1987; 81:155-69. 5. Laitinen LA, Heino M, Laitinen A, Kava T, Haahtels T. Damage of airwayepithelium and bron-
5155 chial reactivity in patients with asthma. Am Rev 24. Tattersfield AE, Holgate ST, Harvey JE, Gribbin HR. Is asthma due to partial l3-blockade of Respir Dis 1985; 131:599-606. 6. Flavahan NA, Aarhus LL, Rimele TJ, Van- airways? Agents Actions [Suppl] 1983; 13:265-71. houtte PM. Respiratory epithelium inhibits bron- 25. Goldie RG, Spina D, Henry PJ, Lulich KM, chial smooth muscle tone. J Appl Physiol 1985; Paterson JW. In vitro responsivenessof human asth8:834-8. matic bronchus to carbachol, histamine, l3-adre7. Goldie RG, Papadimitriou JM, Paterson JW, noceptor agonists and theophylline. Br J Clin PharRigby PJ, Self HM, Spina D. Influence of the epi- macol 1986; 22:669-76. thelium on responsiveness of guinea-pig trachea 26. Cerrina J, Ladurie ML, Labat C, Raffestin to contractile and relaxant agonists. Br J Pharmacol B, Bayol A, Brink C. Comparison of human bronchial muscle responses to histamine in vivo with 1986; 87:5-14. 8. Woolcock AJ, Salome C, Yan K. The shape of histamine and isoproterenol agonists in vitro. Am the dose-response curve to histamine in asthmatic Rev Respir Dis 1986; 134:57-61. and normal subjects. Am Rev Respir Dis 1984; 27. Barnes PJ, Pride NB. Dose-response curves to inhaled l3-adrenoceptor agonists in normal and 13:71-5. 9. Fernandes LB, Paterson JW, Goldie RG. Coasthmatic subjects. Br J Clin Pharmacol 1983; axial bioassay of an epithelium-derived inhibitory 15:677-82. factor (EpDIF) from guinea trachea. Br J Phar- 28. Meurs H, Koeter GH, De Vries K, Kauffman HE The l3-adrenergicsystem and allergic bronchial macol 1989; 96:117-24 10. BarnesPJ,ChungKF,PageCPo Inflammatory me- asthma: changes in lymphocyte l3-adrenergic recepdiators and asthma. Pharmacol Rev 1988; 40:49-84. tor number and adenylate cyclase activity after an allergen-induced asthmatic attack. J Allergy Clin 11. Gardiner PJ, Collier HOJ. Specific receptors for prostaglandins in airways. Prostaglandins 1980; Immunol 1982; 70:272-80. 19:819-41. 29. Goldie RG, Spina D, Lulich KM, Paterson Jw. 12. Hanna CJ, Bach MK, Pare PD, Schellenberg The status of l3-adrenoceptor function in asthma. RR. Slowreacting substance (Ieukotrienes) contract Pharmacology: Excerpta Medica Congress Series human airway and pulmonary vascular smooth 1987; 750:465-8. muscle in vitro. Nature 1981; 290:343-4. 30. TakiF, TakagiK, Satake T, SugiyamaS, Ozawa 13. Barnes NC, Piper PJ, Costello JE Compara- T. The role of phospholipasein reduced l3-adrenergic tive effects of inhaled leukotriene C., leukotriene responsiveness in experimental asthma. Am Rev Respir Dis 1986; 133:362-6. D. and histamine in normal human subjects. Tho31. Patel KR, Kerr JW. The airways response to rax 1984; 39:500-4. 14. PageCP, ArcherCB, Paul W, Morley J. Paf- phenylephrine after blockade of a and l3-receptors acether: a mediator of inflammation in asthma. in extrinsic bronchial asthma. Clin Allergy 1973; 3:439-48. Trends Pharmacol Sci 1984; 5:239-41. 15. Cuss FM, Dixon CMS, Barnes PJ. Effects of 32. Szentivanyi A. The radio ligand binding apinhaled platelet activating factor on pulmonary proach in the study of lymphocytic adrenoceptors function and bronchial responsivenessin man. Lan- and the constitutional basis of atopy. J Allergy Clin cet 1986; 2:189-92. Immunol 1980; 65:5-11. 16. Hwang SB, Lam MH, Shen TY. Specific bind- 33. Barnes PJ, Dollery CT, MacDermot J. Ining sites for platelet activating factor in human lung creased pulmonary a-adrenergic and reduced l3-adtissues. Biochem Biophys Res Commun 1985; 128: renergic receptors in experimental asthma. Nature 1980; 285:569-71. 972-9. 17. Chung KF, Dent G, McCusker M, Guinot P, 34. Kneussl MP, Richardson JB. Alpha-adrenergic Page CP, Barnes PJ. Effect of a ginkolide mixture receptors in human and canine tracheal and bron(BN 52063) in antagonizing skin and platelet re- chial smooth muscle. J Appl Physiol 1978; 45: sponses to platelet activating factor in man. Lan- 307-11. 35. Goldie RG, Lulich KM, Paterson JW. Broncet 1987; 1:248-51. 18. Goldie RG, Paterson JW, Spina D, Wale J. chial c-adrenoceptor function in asthma. Trends Classification of l3-adrenoceptors in human isolated Pharmacol Sci 1985; 6:469-72. bronchus. Br J Pharmacol 1984; 81:611-5. 36. Richardson JB. Nerve supply to the lungs. Am 19. Zaagsma J, Van der Heijden PJ, Van der Rev Respir Dis 1979; 119:785-802. Schaar MW, Bank CM. Differentiation of func37. Barnes PJ. The third nervous system in the tional adrenoceptors in human and guinea-pig air- lung: physiology and clinical perspectives. Thorax ways. Eur J Respir Dis 1984;65(Suppl 135:16-33). 1984; 39:561-7. 20. Barnes PJ. Neural control of human airways 38. Grundstrom N, Andersson RGG. Inhibition in health and disease. Am Rev Respir Dis 1986; of the cholinergic neurotransmission in human airwaysvia prejunctional a,-adrenoceptors. Acta Phys134:1289-314. 21. Paterson JW, Lulich KM, Goldie RG. Drug iol Scand 1985; 125:513-7. effects on l3-adrenoceptor function in asthma. In: 39. Barnes PJ. Localization and functionof airMorley J, ed. Perspectives in asthma. 2.I3-adreno- way autonomic receptors. Eur J Respir Dis 1984; ceptors in asthma. London: Academic Press, 1984; 65(Suppl 135:187-97). 40. Phipps RJ, Williams IP, Richardson PS, Pell 245-68. 22. Szentivanyi A. The l3-adrenergictheory of the J, Pack RJ, Wright N. Sympathomimetic drugs atopic abnormality in bronchial asthma. J Allergy stimulate the output of secretory glycoproteinsfrom human bronchi in vitro. Clin Sci 1982; 63:23-8. 1968; 42:203-32. 23. SvedrnyrNLV, Larsson SA, Thiringer GK. De- 41. Kaliner M, Orange RP, Austen KE Immunovelopment of "resistance" in l3-adrenergicreceptors logical release of histamine and slow reacting substance of anaphylaxis from human lung. J Exp Med of asthmatic patients. Chest 1976; 69:479-83.
Fig. 2. Autoradiographic distribution of binding sites for 1lSl-labeled Bolton Hunter Substance P (I-SP; 0.2 nM) in guinea pig lung (A-C), nondiseased human bronchus (D-F), and asthmatic human bronchus (G-I). Plates A, D, and G are dark-field photomicrographs showing totall-SP binding in the frozen tissue sections shown in light-field photomicrographs B, E, and H, respectively. Note that the smooth muscle (SM) and epithelium (Epi) of the guinea pig bronchus (Br) and bronchiole (BI), as well as vascular (V) tissue, were heavily labeled. In sharp contrast, SM and Epi were very sparsely labeled as were submucosal glands (GI) and cartilage (C) in both nondiseased and asthmatic bronchi. However, there is intense labeling of submucosal tissue surrounding these structures in human bronchi from both sources. Nonspecific I-SP binding in respective adjacent sections was determined in the presence of 1 11M unlabeled SP (plates C, F, and I). Bar = 100 11m.
5156 1972; 136:556-67. 42. Davis C, Karman MS, Jones TR, Daniel EE.
Control of human airway smooth muscle: in vitro studies. J Appl Physiol 1982; 53:1080-7. 43. Carstairs JR, Barnes PJ. Visualization of vasoactive intestinal peptide receptors in human and guinea-pig lung. J Pharmacol Exp Ther 1986; 239:249-55.
44. Taylor SM, Pare PD, Schellenberg RR. Cholinergic and nonadrenergic mechanisms in human and guinea pig airways. J Appl Physiol 1984; 56:958-65.
R. G. GOLDIE
45. Barnes PJ, Dixon CMS. The effects of inhaled
vasoactive intestinal peptide on bronchial hyperreactivity in man. Am Rev Respir Dis 1984; 130: 162-6. 46. Morice A, Unwin RJ, Sever PS. Vasoactive
intestinalpeptide causesbronchodilatation and protects against histamine-induced bronchoconstriction in asthmatic subjects. Lancet 1983; 1225-6. 47. Polack JM, Bloom SR. Regulatory peptides in the respiratory tract of man and other animals. Exp Lung Res 1982; 3:313-28. 48. Lundberg JM, Saria A, Brodin E, Rosell S,
Folkers K. A substance P antagonist inhibits vagally induced increase in vascular permeability and bronchial smooth muscle contraction in the guinea pig. Proc Nat! Acad Sci USA 1983; 80:1120-4. 49. Lundberg JM, Mart!ing C-R, Saria A. Substance P and capsaicin-induced contraction of human bronchi. ActaPhysiol Scand 1983; 119:49-53. 50. Carstairs JR, Barnes PJ. Autoradiographic mapping of substance P receptors in lung. Eur J Pharmacol 1986; 127:295-6.
