The purposes of this project are to study the roles of peptides and peptidases, especially enkephalinase in regulating airways. I will determine which peptides stimulate secretion, smooth muscle contraction, and modulate neurotransmission by exposing tissues to suspected peptide agonists in vitro. I will measure the release of macromolecules labeled biochemically using radioactive precursors, immunological, and morphological methods. I will measure isometric contraction of airway smooth muscle using standard methods. I will determine the response of airways to peptides in the presence of selected peptidase inhibitors. I will determine the identities of endogenous peptidases within the airways, and determine what physiological and biochemical properties of these enzymes make them important in regulating airways. To accomplish these aims, I will perform a series of physiological and biochemical studies and will apply strategies that I used to determine the presence and function of enkephalinase in the airway. Preliminary experiments showed that selected peptides stimulated secretion, muscle contraction, and alter neurotransmission. These include substance P (SP), and neurokinins A and B (NK-A, NK-B). SP stimulated secretion and contraction in concentration-dependent fashions. A combination of proteinase inhibitors potentiated SP-induced effects as did inhibitors of enkephalinase alone. Enkephalinase inhibitors potentiated cholinergic neurotransmission implying that endogenous tachykinins modify cholinergic transmission. These inhibitors also potentiated effects of SP on neurotransmission. Inhibitors of enkephalinase also potentiated the responses to NK- A and NK-B, but did not potentiate secretion induced by VIP or bradykinin. Extracts of airways cleaved an artificial substrate for enkephalinase, effects inhibited by specific inhibitors and substrates for enkephalinase. These results suggest that endogenous peptidases are important in regulating peptide- induced airway responses. These results also suggest that abnormalities in the peptide or peptidase regulation of airways might result in diseases of the airways. Understanding the roles of airway peptides and peptidases will assist in the design and interpretation of other studies of airway function, and may provide knowledge necessary to design effective treatments for diseases characterized by abnormalities in mucociliary clearance, such as asthma, cystic fibrosis, and chronic bronchitis.
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