Molecular basis of modulation of the vascular TMEM16A channel by GPCR pathways: Therapeutic potential for control of artery tone

Title : Molecular basis of modulation of the vascular TMEM16A channel by GPCR pathways: Therapeutic potential for control of artery tone

Abstract:

Introduction: The TMEM16A channel is a Ca²⁺-activated Cl^- channel (CaCC) highly expressed in mammalian arterial smooth muscle. The channel connects Ca²⁺ release triggered by G_q protein-coupled receptor (G_qPCR) activation with membrane depolarisation and smooth muscle contraction. The channel is a proposed drug target for vascular diseases such as hypertension and stroke. While the activation of the channel by inositol 3-phosphate (IP₃)-mediated Ca²⁺ release is well established, whether other GPCR signalling pathways modulate the channel is poorly defined. The aim of this study is to define the GPCR signalling pathways that control TMEM16A channel activity and elucidate the physiological significance of these mechanisms with emphasis on the role of G-protein βγ subunits.

Materials and methods: Whole-cell patch-clamp electrophysiology and assessment of the isometric force of isolated artery rings (wire myography) were used to gain insights into the mechanisms of TMEM16A channel regulation following GPCR activation.

Results: The TMEM16A channel heterologously expressed in human embryonic kidney 293 T (HEK-293T) cells gave rise to large outwardly rectifying currents similar to CaCC currents observed in arterial smooth muscle cells. Co-expression of TMEM16A channel with the α₁ adrenoceptor (a G_qPCR receptor) in HEK-293T cells gave rise to Cl^- currents in response to phenylephrine, an α₁ adrenoceptor agonist. Inhibition of βγ subunits with gallein did not affect the properties of the TMEM16A current in both the absence or presence of unstimulated α₁ adrenoceptor. When the α₁ adrenoceptor was stimulated with phenylephrine, however, gallein dampened the activation of the TMEM16A current by the agonist. Furthermore, the inclusion of βγ subunits in the pipette solution potentiated the TMEM16A current. Unexpectedly, treatment of isolated rat aortic rings with gallein did not affect the response of the vessel to phenylephrine.

Conclusions: The study reveals that the TMEM16A channel is controlled by the βγ subunit during the activation of the α₁ adrenoceptor. In isolated aortic rings, inhibition of βγ subunits did not result in an altered response to phenylephrine, possibly because multiple membrane currents are modulated by βγ subunits counteracting their contribution to vessel tone.

Audience Take Away: 

• This is a basic research project focused on the mechanisms of regulation of the vascular TMEM16A chloride channel by GPCR pathways. The audience will learn about the potential for therapeutic exploitation of the TMEM16A channel for the control of artery tone
• The advanced approaches used in this project can be applied to the study of ion channels in a variety of cardiovascular cell types, helping colleagues to implement new approaches in their own cellular pharmacology research
• Understanding the regulatory mechanisms of ion channels like TMEM16A can help develop more targeted, accurate therapeutic strategies for managing cardiovascular diseases including hypertension and stroke
• This poster is likely to attract the interest of conference participants broadly involved in research of pathophysiology and pharmacology of cardiovascular ion channels

Biography:

Catherine Choi studied Integrated Pharmacology and Physiology at King’s College London, United Kingdom and graduated as MSci in 2017. She then joined the research group of Professor Paolo Tammaro at the Department of Pharmacology, University of Oxford.