Odorant and taste receptor systems in the heart: investigation of novel cardiac biology

Foster, Simon R. (2013). Odorant and taste receptor systems in the heart: investigation of novel cardiac biology PhD Thesis, School of Biomedical Sciences, The University of Queensland.

       
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Author Foster, Simon R.
Thesis Title Odorant and taste receptor systems in the heart: investigation of novel cardiac biology
School, Centre or Institute School of Biomedical Sciences
Institution The University of Queensland
Publication date 2013
Thesis type PhD Thesis
Total pages 362
Language eng
Subjects 1115 Pharmacology and Pharmaceutical Sciences
1102 Cardiovascular Medicine and Haematology
0601 Biochemistry and Cell Biology
Formatted abstract
G protein-coupled receptors (GPCRs) are seven transmembrane-spanning proteins that mediate cellular and physiological responses – they are critical for cardiovascular function and are targeted by frontline therapies for the treatment of hypertension and heart failure. Nevertheless, these therapies only target a small fraction of the cardiac GPCR repertoire, indicating that there are opportunities to investigate unappreciated aspects of heart biology.

The odorant and taste receptors account for more than half of the human GPCR superfamily, and have generally been considered as exclusive mediators of olfaction and taste. However, recent evidence suggests that members of the large, distinct taste and odorant GPCR families have specific functions in tissues beyond the oronasal cavity, including in the skeletal muscle, brain, gastrointestinal tract and respiratory system. In spite of their poorly characterised pharmacology, these GPCRs are emerging as novel therapeutic targets, although they have yet to be systematically studied in the heart. Thus, the primary aim of this thesis was to test the hypothesis that odorant and taste receptors are expressed in multiple cardiac cell types and that these receptors play an important role in heart function.

Previous microarray data generated in our laboratory have indicated that several odorant GPCRs are regulated in ventricular cardiomyocytes following hypertrophic stimuli, providing the impetus for their further characterisation in this project. In promising early experiments, numerous ORs were identified in rodent heart and cardiac cells and were subsequently cloned. However, it proved very challenging to progress the work, given the large number of potential odorant GPCR targets and lack of pharmacological tools and ligands. Accordingly, the focus of this project was shifted towards the more tractable family of taste receptors.

RT-qPCR screens in rodent hearts and in cultured cardiac cells revealed discrete subsets of type 2 taste receptors (Tas2) as well as Tas1r1 and Tas1r3 (comprising the umami receptor) were expressed. These taste GPCRs were present in cardiac myocytes and fibroblasts, and were visualised across the myocardium in isolated cardiac cells by in situ hybridisation. Tas1r1 gene-targeted mice (Tas1r1Cre/Rosa26tdRFP) strikingly recapitulated these data. Moreover, the cardiac-expressed Tas2rs are located in genomic clusters and shared developmental and physiological/pathological expression patterns, suggesting common regulatory control. Intriguingly, several Tas2rs were upregulated in cultured rat myocytes and in mouse heart in vivo following starvation, and in a model of pathological hypertrophy and heart failure, suggesting that the expression of these taste receptors in the heart may play a role in cardiac nutrient sensing. Accordingly, I investigated a potential role for these taste GPCRs in autophagy but siRNA knockdown of Tas1r3 and Tas2r126 had no discernible effect on the autophagic process.

In order to interrogate the physiology of the cardiac-expressed taste GPCRs, I sought to identify agonist ligands. Five Tas2 GPCRs were cloned from rat heart and were screened against a panel of 102 synthetic and natural bitter compounds in a heterologous expression system. Using intracellular Ca2+ mobilisation as a readout of receptor activation, new ligands were identified for three receptors (Tas2r108, Tas2r137 and Tas2r143), providing the necessary tools to probe T2R function in the heart.

Subsequently, these novel taste GPCR ligands were applied in an ex vivo perfused mouse heart model to study their effects on cardiac function. All putative T2R ligands tested exhibited concentration-dependent effects, notably a biphasic change of aortic pressure with sodium benzoate and a sodium thiocyanate-mediated decrease in systolic pressure. Strikingly, both of these responses were abrogated in the presence of Gαi (pertussis toxin) and Gβγ (gallein) inhibitors, suggesting these bitter ligands elicit G protein-dependent effects on rodent cardiac function.

In addition, taste receptors are expressed in the healthy and failing human heart. Remarkably, 15 of the 25 human TAS2 GPCRs were detected, ranging in abundance between that of the important cardiovascular GPCRs, angiotensin II type 1 receptor and the expressed β1-adrenoceptor. To begin to investigate the cardiac function of these GPCRs, explanted human right atrial tissue was stimulated with T2R ligands, corresponding to the most highly expressed receptors. Albeit at very high ligand concentrations, consistent cardiodepressant (negative inotropic) effects were observed, opening the possibility of T2R-dependent effects in human heart.

In summary, I provide the first evidence that taste GPCRs are expressed in human and rodent heart, foreshadowing an exciting new field of cardiac research. Using a combination of molecular, physiological and pharmacological approaches, several members of a GPCR family previously unrecognised in the heart have been identified, cloned and deorphanised. Moreover, ex vivo experiments in human and rodents have demonstrated profound bitter ligand-induced and G protein–dependent effects on cardiac contractility. The novel cardiovascular depressant effects of these ligands are of significant interest, and the natural extension of this work will be to establish the specific G protein and taste GPCR-dependent responses in the heart.
Keyword G protein-coupled receptor
Taste receptor
Odorant receptor
Heart
Cardiac physiology
Cell signalling
Human myocardium

 
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Created: Tue, 11 Mar 2014, 12:03:04 EST by Simon Foster on behalf of Scholarly Communication and Digitisation Service