Purpose of the Conference

Chairman's Summary

The overall aim of the Transatlantic Airway Conference (TAC)is to reviewrecent advances in research directed at various aspects of airway physiology and pathophysiology. The fourth conference was held in Key Biscayne, Florida in January 1989. This year's theme was receptors and second messengers. In the last fewyears, there have been remarkable advances in the understanding of receptor mechanism due to recent progress in pharmacology, biochemistry, and molecular biology. A summary of new information was therefore timely. Under the experiencedand skillful guidance of Peter J. Barnes, chairman of the meeting, the participants, who are acknowledged leaders in their respective scientific areas of research, identified basic processes involved in the activation of receptors and second messengers, and established a link to airway disease. The free exchange of ideas also raised relevant new questions to be answered by future investigations. The scientific papers included in this supplement formed the basis of these productive discussions. We would like to thank the following participants: P. Barnes, C. Basbaum, N. Birdsall, L. Birnbaumer, L. Block, J. K. Brown, R. F. Coburn, J. Douglas, B. Fredholm, R. Goldie, B. Grandordy, M. Kaliner, M. Kotlikoff, R. D. Krell, D. Lew, J. MacDermott, P. Minette, S. Nahorski, F. Nijkamp, J. Putney, J. A. M. Raaijmakers, C. Roberts, U. Ryan, D. Ukena, C. Venter, J. Westwick, and H. Zaagsma. In addition, we are indebted to G. Krick, M. T. Lopez-Vidriero, and the Boehringer Ingelheim Company for their professional support of the conference.

Scientific Committee: (Chairman) University oj Miami Miami, FL, U.S.A.

ADAM WANNER

HOMER BOUSHEY

University oj California San Francisco, CA, U.S.A. ALAIN JUNOD

University oj Geneva Geneva, Switzerland ANDRE ~ PERRUCHOUD

University oj Basel Basel, Switzerland

There have recently been quite remarkable advances in our understanding of receptor mechanisms, and recent advances in pharmacology, biochemistry, and molecular biology have now made it possible to understand the mechanisms of receptor action in considerable detail. Many of these advances are unknown to clinical scientists working on diseases in which abnormal function of receptors may be critical. The Fourth Transatlantic Airway Conference was designed to bring together recent research into receptor mechanisms and second messengers with the possible application of this research to airway disease. The interaction between the basic scientist and the clinical scientist is, of course, a two-way process; the clinician can learn about the outstanding advances in receptor science and might use such information to try to further understand disease processes or to develop new approaches to therapy. Conversely, the basic scientist can learn from clinicians what questions may be relevant to pursue and how disease can provide information about receptor regulation. Knowledge of receptors and second messengers is fundamental to understanding the normal and abnormal control of airway function. Many types of receptors are involved in regulation of target cells in the airways, and the inflammatory cells that may infiltrate the airwaysin disease. More than 80 distinct receptors have now been identified. Surface membrane receptors relevant to airway disease can be broadly divided into neurotransmitter receptors that are operated by transmitters (including neuropeptides) released from airway nerves, hormonal receptors that are responsive to circulating agonists, receptors for the multiplicity of cell mediators released in inflammatory conditions, and receptors that respond to various growth factors. Receptors determine cell responsivenessand can be selectively activated by low concentrations of endogenous agonists. Receptors have the quality of specificity and allow a cell to respond selectively to the myriad of chemical signals to which it is exposed. Activation of receptors then leads to the typical cellular response. The black box between stimulus and response is now becoming smaller and great strides have now been made in understanding the second messengers involved. There are relatively few second messenger mechanisms since specificity is provided by the surface receptor population and the number of ways in which a cell can respond is limited. The meeting was structured into three sections. Initially, basic mechanisms of receptor function were considered, followed by discussion on particular cell types relevant to airway disease, and finally the role of receptors in airway disease was discussed. Advances in molecular biology have had a major impact on receptor research. It has now become possible to clone and express

many types of surface receptor, and this has allowed questions to be answered that would have previously been inconceivable. One surprising discovery with sequence analysis of receptor proteins has been the homology between completely different receptors. Receptors that interact with guanine nucleotide regulatory proteins (G proteins) have certain common sequences, such that they can be classified as members of a large supergene family, that differ in their extracellular domains, which are involved in regulation of different ligands. Advances in technology are now making it possible to study the physical shape or conformation of these proteins and this will presumably aid the development of novel drugs that interact with these receptors. The use of site-directed mutagenesis to substitute at critical sites in the molecule has also provided important insights into the working of surface receptors. The development of cDNA probes for receptor messenger RNA provides a powerful tool to study genetic expression of receptors and, when combined with in situ hybridization, allows precise localization of receptor synthesis. The potential of this approach in airways disease is enormous, yet there have been rather few studies using this powerful technology in lung disease. The coupling of receptors to intracellular biochemical events is now much better understood, and the structure of G proteins has been elucidated. G proteins that stimulate and inhibit adenylate cyclase, that stimulate phospholipases (including phosphoinositidase), and that activate certain membrane channels are now recognized, and the genetic expression of the components of G proteins is an area of intense research activity. Because G proteins may determine the responsiveness of cells, they may be an important site of abnormality in certain diseases and may be a suitable target for future drug therapy. The second messengers involved in signal transduction have been extensively investigated. The importance of the adenylate cyclase regulation of intracellular cyclic AMP has been obvious for some time, although the relevance of inhibition of adenylate cyclase by some receptors (such as muscarinic receptors in airway smooth muscle) is still not entirely clear. Intracellular calcium is an important signaling mechanism, and the mechanisms by which surface receptors may lead to release of calcium ions from intracellular stores, via phosphoinositide (PI) hydrolysis, are now much better understood. However, many mysteries remain and the role of inositol tetrakisphosphate (IP4), IP5, and IP6 on in cell signaling is not yet clear. The regulation of PI hydrolysis is complex and incompletely understood. The interaction between PI hydrolysis and protein kinase C activation are potentially very important for normal and abnormal cell function, and may have particular relevance for airways disease. The interaction between receptors and ion channels is also an important area of study, particularly

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for epithelial cellsand inflammatory cells.The mechanism by which growth factor receptors and receptors for cytokines influence cell function is also of great relevance in disease. The role of tyrosine kinase activation in these events is an important area of research that has barely been applied to airway disease. Subtypes of receptor have been recognized for many years and were initially defined in terms of agonist potency and later by the development of selective antagonist drugs. Their existenceis now confirmed by molecular cloning techniques. Indeed, receptor subtypes have been cloned for which no drugs are yet able to discriminate. Thus, for muscarinic receptors five distinct receptor proteins have now been cloned and expressed,yet only three subtypes of receptor can be differentiated with currently available drugs. Why should so many subtypes of receptor exist? Sometimes this may be because the natural agonist differs - thus for beta-adrenoceptors, beta-l receptors are preferentially activated by norepinephrine released from adrenergic nerves, whereas beta-2 receptors are regulated by circulating epinephrine. Similarly, for tachykinin receptors, substance P preferentially activates a receptor (called NK-I receptor) that is distinct from the receptor activated by neurokinin A (NK-2 receptor) and neurokinin B (NK-3 receptor). But sometimes the same agonist activates different receptor subtypes; thus, acetylcholine is the only natural agonist for the multiple muscarinic and nicotinic receptors now recognized. In such cases, it is likely that each receptor subtype may be linked to a different intracellular mechanism and can thus givea different signal to the cell. Regulation of receptor function and second messengers is obviously of very great relevance to understanding normal physiology and disease states. Many factors that can influence receptor function, both homologous and heterologous, are now recognized, and increasingly,there are demonstrations that one type of receptor can influence the expression or activity of a quite different receptor (receptor transmodulation). This may have particular relevance in disease when cells are bathed in abnormal stimuli, such as inflammatory mediators, such that the cell changes its responsiveness to other stimuli. More needs to be discoveredabout how receptors talk to each other to lead to the final cellular response, and how intracellular events can influence the function of surface receptors. Of course, molecular biology should greatly advance our

understanding of the regulation of receptor and G-protein synthesis. While there have been major advances in our understanding of basic receptor mechanisms, it has been rather disappointing that so little of the research has been applied to cells relevant to airway disease. It is rather obvious that different cells will show profoundly different responses to the same signals and what is true in a turkey erythrocyte cannot necessarily be applied to an airway smooth muscle cell or to a mast celL Yetthe study of airway cellsis not as easy as the study of cultured cell lines or blood cells since separation of pure cell populations is often difficult. Some of the target cells that may be important in airway disease were discussed in depth. Most information is known about airway smooth muscle since its function may be studied by classic pharmacologic techniques and it has been possible to obtain relatively pure piecesof airway smooth muscle for functional and for biochemical studies. The development of cultured airway smooth muscle cells is also a useful new technology since this allows the study of single cells both electrophysiologically and functionally. Yetthere are disadvantages of studying single cells and these were discussed throughout the meeting. Dissociation of cells from tissues may change the surface receptors quite markedly - thus trypsin digests surface beta-receptors and removal of cellsfrom their neighbors may also change their function. It may therefore be necessaryto integrate classicfunctional studies with singlecell studies to check any alterations in function. Airway smooth muscle is only one target cell in the airway, yet far less is known about receptors on other target cells, such as epithelial and gland cells, neurons and fibroblasts, endothelial cells, or on migratory cells such as mast cells, macrophages, lymphocytes, neutrophils, eosinophils, and platelets, which may participate in disease processes. The distribution of various receptors within the airway has been studied by autoradiographic mapping techniques, but it is sometimes difficult to perform the essential functional studies to link in with this approach. The development of micromethods should make this possible in the future. Although the receptor regulation of normal constituents of the airway is poorly understood, even less is known of changes in receptor function that might pertain in disease. Altered autonomic receptor function has

long been postulated in asthma and most studies of receptor function in airway disease have concentrated on the possible imbalance between bronchoconstrictor and bronchodilator receptors. While the initial belief that there was a fundamental abnormality in asthma, this is unlikely to be correct and there is compelling evidence that changes in receptor function may occur secondary to airway disease, perhaps as a result of a chronic inflammatory process. The molecular basis for such changes requires more detailed study, preferably on diseased tissue rather than on animal models, which have often proved to be misleading in this area. In the future, a deeper understanding of receptor function in airway disease may lead to novel approaches to asthma and other airway diseases. Drugs that interact with receptors in a new way or that interact with G proteins or with second messenger systems may prove to be usefuL Drugs that interfere with fundamental processes, such as breakdown of phosphoinositides, release of intracellular calcium via IP3 or with G-protein signaling, might be considered to be too nonspecific to be useful but, in airway disease, it is possible to obtain a certain degree of selectivity by delivering drugs via the aerosol route. Thus, airway diseases might provide a useful testing ground for some of the future therapeutic developments. The Fourth Transatlantic Airway Conference certainly achieved its objective ofbridging the gap between basic science and airway disease, and I believeeveryone benefited from such interaction. This workshop came up with a number of new ideas and many important questions that will need to be addressed over the next few years. It is clear that molecular biology will have a profound impact on airway research but should not be considered in isolation. It is the close link between molecular biology, pharmacology, biochemistry, and physiology that will answer the important questions. We have the tools; now we must apply them sensibly. The future is exciting, and it is obvious that this is an area of research in respiratory medicine that is ripe for important advances. PETER J. BARNES, M.D. Department of Thoracic Medicine National Heart and Lung Institute London, England