Sebastian, S;
(2015)
The role of G-proteins in the sino-atrial node in controlling heart rate.
Doctoral thesis , UCL (University College London).
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Abstract
Reciprocal physiological modulation of heart rate is controlled by the sympathetic and parasympathetic systems acting on the sino-atrial (SA) node. However there is little direct in vivo work examining the role of stimulatory and inhibitory G-protein signalling in the SA node. We have previously shown that the inhibitory G-proteins Gαi2, is important in mediating vagal tone to the murine heart with global Gαi2 knock-out mice developing tachycardia with loss of high frequency power and being resistant to the effects of the muscarinic agonist, carbachol (Zuberi et al. 2008). However, it is unclear whether the effect is mediated entirely via Gαi2 signalling at the end-organ level or whether there is a central component. I sought to investigate this by studying mice with conditional knock outs (KO) of Gαi2 and Gαo in cardiac tissue by using αMHC cre mice. Studies with αMHC cre mice using the cre lox-P approach were negative as there was no difference in heart rate variability (HRV) and electrocardiograph (ECG) parameters between controls and conditional KO of Gαi2 and Gαo, though by real-time polymerase chain reaction (RT-PCR) we could see reduction in the level of their expression. I assumed deletion of Gαi2 and Gαo may not be complete in the SA nodal region where G-protein inwardly rectifying K+ channels (GIRK) channels are active. I sought to investigate this further by studying mice with conditional deletion of Gαi2 and Gαs in the conduction system of the murine heart using cre lox-P technology. I crossed mice in which cre recombinase expression was driven by a tamoxifen-inducible conduction system-specific construct with Gαs floxed and Gαi2 floxed mice. I studied the heart rate responses of adult mice compared with littermate controls by using ECG radio-telemetry before and after administration of tamoxifen. The mice with conditional deletion of Gαs and Gαi2 had a loss of diurnal variation and were bradycardic or tachycardic in the daytime respectively. In mice with conditional deletion of Gαs there was a selective loss of low frequency power whilst with deletion of Gαi2 there was a loss of high frequency power in power spectral analysis of heart rate variability. There was no evidence of pathological arrhythmia. Pharmacological modulation of heart rate by isoprenaline was impaired in the Gαs conditional knock-out mice but a muscarinic agonist was still able to slow the heart rate in Gαi2 conditional knock-out mice. We conclude that Gαs and Gαi2 mediated signalling in the sino-atrial node is important in the reciprocal regulation of heart rate through the autonomic nervous system. Another way to look at the role of specific gene function is via gain of function mutation. Regulators of G-protein signalling (RGS) proteins are a recently discovered family of proteins that negatively regulate G-protein-coupled signalling pathways. Thus, I used RGS-insensitive (RGSi) mutant Gα subunits, which contain a G184S point mutation in their switch I region that prevents binding of RGS proteins and the subsequent Gα deactivation. RGS insensitive Gαi2 and Gαo mice (RGSiGαi2 and RGSiGαo) were found to be tachycardic and lost diurnal variation in heart rate. The A1 receptor agonist, 2-chloro-N6-cyclo-pentlyadenosine (CCPA) and M2 receptor agonist, carbachol showed an enhanced inhibitory response (bradycardic) to RGSiGαo but this enhanced bradycardic response was observed only with carbachol for RGSiGαi2. HRV signature was found to be lost in RGSiGαo but not with RGSiGαi2. Interestingly, studies with RGSiGαi2 and RGSiGαo prove a role of the central nervous system in controlling the heart rate in these knock-ins. Gαi2 levels are up-regulated in heart failure. However, it was not clear whether its role is protective or maladaptive in the context of heart failure. Hence, I subjected the mice in which cre recombinase expression was driven by a tamoxifen-inducible conduction system-specific construct with Gαs floxed and Gαi2 floxed mice to myocardial infarction by left anterior descending (LAD) artery ligation. I found that cardiac-specific knock-out of Gαs attenuates left ventricular (LV) dilation and improves LV contractile function after myocardial infarction (MI). Conditional deletion of Gαs was found to be protective as this was reflected in increased fractional shortening (FS%), ejection fraction (EF%) and fractional area change (FAC%) on echocardiographic analysis after MI. LV remodelling was observed in cardiac-specific knock-out of Gαi2 to a certain extent with chamber dilatation and decreased EF% and FS% and FAC%. In summary absence of Gαs is found to preserve LV function may be by its bradycardic nature may protect the heart against the ischemic insult, although the precise mechanism that helps against the adverse LV remodelling need to be explored. On the other hand absence of Gαi2 and moderate tachycardia in these mice didn’t have any detrimental effects on LV remodelling.
Type: | Thesis (Doctoral) |
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Title: | The role of G-proteins in the sino-atrial node in controlling heart rate |
Open access status: | An open access version is available from UCL Discovery |
Language: | English |
UCL classification: | UCL UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Medicine |
URI: | https://discovery.ucl.ac.uk/id/eprint/1469611 |
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