Sunday, May 18, 2025
5/18/2025
Connexin hemichannels: an unexplored critical component in endothelium of resistance arteries - Project Summary/Abstract Blood pressure is controlled mainly at the arterioles in the microcirculation. The endothelium plays a critical role in controlling function of the vessel by the production of vasodilators as vasoconstrictors agents. In arterioles the main vasodilator agents are nitric oxide (NO) and endothelium-derived hyperpolarization (EDH) by direct activation of Ca2+-activated K+ channels (KCa) of small (SKCa) and intermediate (IKCa) conductance which cause vasodilation in resistance arteries. NO and EDH require endothelium Ca2+ increases. NO modulates endothelial cell function via either guanylyl cyclase 1 (GC1) or protein S- nitrosylation. While much attention has focused on GC1-associated vasodilation, the role of endothelium evoking S- nitrosylated on calcium permeable channels which will be critical to regulate the vasomotor tone is an interestingly mechanism that has not been fully explored yet. TRPV4 has been described as an essential component in the endothelial Ca2+ increases, promoting an influx of Ca2+ from the extracellular space under shear stress or upon endothelium dependent vasodilators. Thus, TRPV4 activation is critical to promote NO and EDH-driven relaxation. However, even after TRPV4 activation by vasodilators or specific TRPV4 agonists, the endothelial Ca2+ influx and EDH-driven relaxation are dramatically diminished by blocking Cx43 hemichannels. This implies that: 1) TRPV4 is necessary but not sufficient for proper endothelial Ca2+ entry, EDH and dilation; 2) TRPV4-stimulated dilation crucially involves endothelial Ca2+ entry via Cx43 hemichannels. In this context, recent results indicate TRPV4-induced Cx43 hemichannels opening in lens epithelium. Our preliminary results in arteriolar endothelial cells (primary culture as intact endothelium) revealed TRPV4-induced endothelial Cx43 hemichannels activity, endothelial Ca2+ influx, and endothelial hyperpolarization by Cx43 hemichannels activation. Connexin hemichannels form two types of channels. Gap junction channels and hemichannels. Connexin proteins in non-junctional plasma membrane allow tightly regulated extracellular Ca2+ influx in astrocytes, muscle cells, microglia, cardiac cells, etc. Endothelial peripheral arterioles cells express connexin (Cx) 37,40 and 43. However according to our previous data and preliminary results using primary culture endothelial cells, isolated mesenteric arteries the S-nitrosylated Cx43 hemichannels is a critical component in the TRPV4/Cx43 hemichannels signaling. The objective of this proposal is to identify the mechanisms through which TRPV4 activates endothelial Cx43 hemichannels which will be a critical step to trigger Ca2+ increases promoting endothelial hyperpolarization. In addition, we want to test whether i n h y p e r t e n s i o n m o d e l s t h e r e i s a dysregulation of T R P V 4 / Cx43 hemichannels s i g n a l i n g . Besides, we postulate that interaction TRPV4-induced Cx43 hemichannels currents which evoke Ca2+-activated K+ channels (KCa) of small (SKCa) and intermediate (IKCa) conductance promoting endothelial hyperpolarization. Our central hypothesis is that S-nitrosylation of endothelial Cx43 hemichannels by TRPV4 activation is required for the Ca2+ entry, endothelial hyperpolarization and vasodilation upon endothelium-dependent agonist. We will test this hypothesis through two mechanistic Specific Aims (SA): • SA1 Is a TRPV4/Cx43 hemichannel signaling node a key component in arteriolar dilation? • SA2 Do hypertensive mice display a dysfunction in endothelial TRPV4/Cx43 hemichannel activity? These novel studies will yield mechanistic insights into the role of endothelial Cx43 hemichannels. They will also advance knowledge of vascular disease and assist physicians to manage hypertension.
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