Bitter tasting compounds dilate airways by inhibiting airway smooth muscle calcium oscillations and calcium sensitivity

Abstract

While selective, bitter tasting, TAS2R agonists can relax agonist-contracted airway smooth muscle (ASM), their mechanism of action is unclear. However, ASM contraction is regulated by Ca²⁺ signalling and Ca²⁺ sensitivity. We have therefore investigated how the TAS2R10 agonists chloroquine, quinine and denotonium regulate contractile agonist-induced Ca²⁺ signalling and sensitivity. Airways in mouse lung slices were contracted with either methacholine (MCh) or 5HT and bronchodilation assessed using phase-contrast microscopy. Ca²⁺ signalling was measured with 2-photon fluorescence microscopy of ASM cells loaded with Oregon Green, a Ca²⁺-sensitive indicator (with or without caged-IP₃). Effects on Ca²⁺ sensitivity were assessed on lung slices treated with caffeine and ryanodine to permeabilize ASM cells to Ca²⁺ . The TAS2R10 agonists dilated airways constricted by either MCh or 5HT, accompanied by inhibition of agonist-induced Ca²⁺ oscillations. However, in non-contracted airways, TAS2R10 agonists, at concentrations that maximally dilated constricted airways, did not evoke Ca²⁺ signals in ASM cells. Ca²⁺ increases mediated by the photolysis of caged-IP₃ were also attenuated by chloroquine, quinine and denotonium. In Ca²⁺-permeabilized ASM cells, the TAS2R10 agonists dilated MCh- and 5HT-constricted airways. TAS2R10 agonists reversed bronchoconstriction by inhibiting agonist-induced Ca²⁺ oscillations while simultaneously reducing the Ca²⁺ sensitivity of ASM cells. Reduction of Ca²⁺ oscillations may be due to inhibition of Ca²⁺ release through IP₃ receptors. Further characterization of bronchodilatory TAS2R agonists may lead to the development of novel therapies for the treatment of bronchoconstrictive conditions.