Bitter taste receptors (TAS2Rs) have been implicated in roles other than taste perception. TAS2R agonists exert vasorelaxant effects in vascular smooth muscle from guinea pig, mouse and humans. We hypothesis that bitter taste agonists could modulate the pulmonary artery vascular tone. The mechanisms of action of bitter taste agonists, which are used to investigate the functions of TAS2Rs, are unclear. The purpose of this study was to pharmacologically characterise the effects of three common bitter taste agonists in the rat pulmonary artery in vitro to elucidate their mechanisms of action. In addition, this study sought to investigate which TAS2R subunits are expressed in the rat pulmonary artery. Myographic studies were conducted in rat pulmonary artery rings pre-contracted with phenylephrine or U46619 to study the pharmacological effect of bitter taste agonists as well as to investigate the possible mechanisms involved in their effects. The calcium fluorescence dye Cal-520 AM was used to measure the intracellular calcium levels in freshly isolated pulmonary artery SMCs. RNA sequencing analysis was conducted to determine the mRNA expression of the Tas2r subunits in the rat pulmonary artery. In addition, RT-qPCR was conducted using Tas2r-specific primers to validate the results from the RNA sequencing analysis.PCR-based genotyping was also carried out to examine the presence of any potential variations on rat TAS2R genes. Quinine, denatonium, and dextromethorphan caused a concentration-dependent relaxation of the pulmonary artery pre-contracted with phenylephrine. Removal of endothelium had no effect in the concentration-response curves for quinine, denatonium, and dextromethorphan, indicating that the effect of these agonists appears to be endothelium-independent in the rat pulmonary artery. Moreover, BKCa channels and the βγ subunit did not appear to be involved in the mechanism of action of bitter taste agonists on the rat pulmonary artery, as blocking these did not result in an overall significant change in the vasorelaxation. In summary, the results from this in vitro study shows that quinine, denatonium, and dextromethorphan have vasorelaxant activities in the rat pulmonary artery. The vasorelaxant action of quinine and denatonium appears to be through inhibition of extracellular calcium influx, most likely through inhibition of ROCC in pulmonary artery SMCs. Dextromethorphan-induced increases in [Ca2+]i in the rat pulmonary artery isolated SMCs. Dextromethorphan seems to has dual actions; involving the extracellular in nature in addition to inhibition of ROCC, partially via VDCC, and mostly from intracellular Ca2+ stores via the PLC-IP3 pathway in pulmonary artery SMCs. The molecular biology studies have showed that Tas2r108 and Tas2r139 were expressed in the rat pulmonary artery. These findings suggest that these bitter taste agonists are effective vasorelaxant that might be used in pulmonary hypertension.
|Date of Award||20 Mar 2020|
- University Of Strathclyde
|Supervisor||Robert Drummond (Supervisor) & Edward Rowan (Supervisor)|