Bronchial asthma is associated with inflammatory changes, swelling, stiffness of the airways, spontaneous, mechanically or chemically evoked bronchoconstriction. These changes can be initiated and maintained by autonomic imbalance in bronchodilatory and bronchoconstrictive influences. In this grant proposal we will stress how the nervous system via a number of differentiated patterns during prolonged and often repeated stimulations, affect airway structure and functions down to cellular and molecular level of organization. In particular, the role of sensory innervation will be studied. Subsets of sensory fibers, both in animals and humans, contain the tachykinin peptides, substance P (SP) and neurokinin A (NKA), which are colocalized with calcitonin gene related peptide (CGRP). In response to noxious agents and bradykinin (BK), SP, NKA and CGRP are released from sensory nerves and when released they exert a variety of local biological effects, including increase in cholinergic transmission, airway smooth muscle (ASM) contraction, mucus secretion and plasma protein extravasation. These changes are mediated through activation of typical features of bronchial asthma. We hypothesize that in bronchial asthma the regulation of the NK-1, NK-2, and NK-3 receptors in achieved via a feedforward mechanism in which peptide synthesis or secretion (or both) is enhanced. Increased levels of released SP and NKA would then cause increased in the expression and the sensitivity of NK-1, NK-2, and NK-3 receptors, and potentiate cholinergic influences. We will investigate changes in SPK, BK and NKA levels, NK-1, NK-2, CGRP, B-1 and B-2 receptor mRNAs and receptor peptide expression in airways of asthmatics and in the airways of sensitized guinea pigs following repeated exposure to antigen. Preprotachykinin (PTT) mRNA expression in the airways will be quantitated using solution hybridization-nuclease protection assays. Peptide and receptor mRNA expression levels will be normalized to glyceraldehyde-3- phosphatedydrogenase mRNA levels. Similarly, expression of the BK receptor message and receptor protein will be determined in guinea pigs. In the airway smooth muscle preparation, the link between changes in signal transduction pathways and airway hyper- responsiveness will be studied. Using biophysical and biochemical approaches, we will assess the changes in contractility of the airway smooth muscle which could account for the refractoriness to therapy by employing techniques well established in our laboratories. The results of proposed studies will contribute to better understanding of fundamental mechanisms of a feedforward system as a determinant of progression of the pathologic process, and will lead to improved methods in treatment of obstructive airway disorders.
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