Cellular and Molecular Mechanisms of bitter tastant-induced bronchodilation

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): Bitter tastant-induced bronchodilation is a newly discovered form of airway smooth muscle (ASM) relaxation, and bitter tastants hold great promise as bronchodilators, which are indispensable for 300 million patients worldwide with asthma and chronic obstructive pulmonary disease. We seek to address two fundamental issues related to this relaxation: the cellular and molecular mechanisms of action of bitter tastants in ASM, and the effectiveness of bitter tastants in chronic asthma. Bitter tastants activate the type 2 taste receptor (TAS2R)-gustducin-PLCbeta2 pathway in taste cells and some extra-oral cells. Our preliminary data, however, suggest that TAS2Rs and gustducin, but not PLCbeta2, are essential for bitter tastant-induced bronchodilation. To firmly establish this view, we will study the effects of deleting TAS2R105, alpha-gustducin or PLCbeta2 on bitter tastant-induced changes in [Ca2+]i, cell length and tension in ASM (Aim 1). The uniqueness of Tas2r105-/- mice is that the resultant function changes can be assessed reliably with cycloheximide, a ligand specific to TAS2R105. On the other hand, the advantage of alpha-gustducin-/- mice is that although the mouse genome contains 35 TAS2Rs, all of them couple with gustducin. As a result, gustducin deletion could block bronchodilation in response to a broad spectrum of bitter tastants. Our preliminary data further revealed that inhibition of L-type CaV1.2 channels is the key molecular event responsible for bitter tastant-induced bronchodilation, and this inhibition depends on pertussis toxin sensitive gustducin but not PLCbeta. Using patch clamp, pharmacology, genetic knockout mice and heterologous expression systems, we will uncover the molecular mechanism by which bitter tastants inhibit this channel (Aim 2). Finally, bitter tastants are effective bronchodilators in a mouse model of acute asthma, implying their tremendous therapeutic potential in this disorder. Yet, asthma is a chronic disease; it is thus imperative to establish the effectiveness of bitter tastants in chronic asthma. Our preliminary study revealed that bitter tastants reverse the contraction of airways from two mouse models of chronic asthma including one induced by Asperigillus fumigates crude protein extract, a common allergen of human asthmatics. In Aim 3, we will systematically characterize this effect in vitro and in vivo in these mice, and determine its molecular basis using allergen sensitized Tas2r105-/- or alpha-gustducin-/- mice. To translate our findings in mice to human, we will uncover the mechanisms of bitter tastant-induced bronchodilation using human lung specimens. This work should establish not only how bitter tastants cause bronchodilation in mouse and human, but also their usefulness in treating airway diseases in mouse models of chronic asthma. Such advances will deepen our understanding of ASM biology and facilitate the development of bitter tastants as new bronchodilators.