Endothelial-dependent dilation following chronic hypoxia involves TRPV4-mediated activation of endothelial BK...

Following chronic hypoxia (CH), the systemic vasculature exhibits blunted vasoconstriction due to endothelial-dependent hyperpolarization (EDH). Previous data demonstrate that subsequent to CH, EDH-mediated vasodilation switches from a reliance on SK_ca and IK_ca channels to activation of the endothelial BK_ca channels (eBK). The mechanism by which endothelial cell stimulation activates eBK channels following CH is not known. We hypothesized that following CH, EDH-dependent vasodilation involves a TRPV4-dependent activation of eBK channels. ACh induced concentration-dependent dilation in pressurized gracilis arteries from both normoxic and CH rats. Inhibition of TRPV4 (RN-1734) attenuated the ACh response in arteries from CH rats but had no effect in normoxic animals. In the presence of L-NNA and indomethacin, TRPV4 blockade attenuated ACh-induced vasodilation in arteries from CH rats. ACh elicited endothelial TRPV4-mediated Ca²⁺ events in arteries from both groups. GSK1016790A (GSK101, TRPV4 agonist) elicited vasodilation in arteries from normoxic and CH rats. In arteries from normoxic animals, TRAM-34/apamin abolished the dilation to TRPV4 activation, whereas luminal iberiotoxin had no effect. In CH rats, only administration of all three K_ca channel inhibitors abolished the dilation to TRPV4 activation. Using Duolink®, we observed co-localization between Cav-1, TRPV4, and BK channels in gracilis arteries and in RAECs. Disruption of endothelial caveolae with methyl-β-cyclodextrin significantly decreased ACh-induced vasodilation in arteries from both groups. In gracilis arteries, endothelial membrane cholesterol was significantly decreased following 48 h of CH. In conclusion, CH results in a functional coupling between muscarinic receptors, TRPV4 and K_ca channels in gracilis arteries.