Neurogenic inflammation, through the release and action of tachykinins, is an important mechanism contributing to lung pathophysiology. The neurotransmitter peptides, substance P, neurokinin A and neurokinin B, have long been appreciated as small peptides capable of potent actions including bronchoconstriction and edema formation. Data generated in the previous funding period demonstrate that the tachykinin receptors are encoded by single copy multi-exon genes located on chromosomes 2 (NK-1 receptor) and 10 (NK-2 receptor). cDNAs encoding receptors selective for each of the tachykinin ligands have been cloned from human, mouse and several other species, and much has been learned about the ligand- receptor interactions. Additionally, several non-peptide tachykinin receptor antagonists have recently been developed, providing new tools to further investigate the system. This continuation application outlines a molecular biological approach, using both cell culture and in vivo experiments to continue to probe neurogenic inflammation mediated via tachykinin receptors.
For Specific Aim 1, we will examine structure- function relationships of the receptor molecules, identifying interaction sites that confer ligand selectivity.
In Specific Aim 2, we will examine receptor-effector relationships to determine G-protein selectivity and its effect on ligand binding and signal transduction.
For Specific Aim 3, we have created transgenic mice in which exon 1 of the NK-1 receptor is deleted and replaced by the gene for beta-galactosidase, which will be valuable both for reporter functions to determine the spatiotemporal expression of the NK-1 receptor, as well as for characterizing the phenotype of NK-1 receptor deficient animals. Results of these studies will lead to greater understanding of the mechanistics of tachykinin ligand-receptor interactions and their role in lung pathophysiology as well as providing valuable reagents to the field.
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