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

Abstract

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 SKca and IKca channels to activation of the endothelial BKca 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 Kca 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 Kca channels in gracilis arteries.

Involvement of Alternative Calcium Conductance on Human Myometrial Contractile and Pharmacological Properties

“…However, in the present study, there was no coordinated activation between oxytocin and the TRPV4 agonist GSK1016790A. Our current view is that this TRPV4 agonist likely activates an inward Ca 2+ flux, as previously demonstrated on vascular smooth muscle cells [9,27], which in turn would directly or indirectly activate an outward K + current through BKCa channels, located nearby. Activation of this specific K + conductance is responsible for membrane hyperpolarization and concomitant Ca 2+ channel inactivation resulting in contractile quiescence.…”

“…Ca 2+ entry (through Nifedipine-sensitive Ca 2+ channels) or Ca 2+ release (through IP 3 -receptors) would trigger strong and efficient phasic contractions [6], while divalent cationic currents, through TRPV proteins and/or Ryanodine-receptors, would regulate relaxation rate and cell membrane polarization (due to the activation of BKCa channels), hence contractile frequency. While these assertions remain speculative since no corresponding measurements were performed in the current study, the likelihood of fine-tuning through activation of K + conductance is nonetheless very high in smooth muscle cells [4,27,31]. Accordingly, IbTX was used herein to block the K + conductance through BKCa channels, which prevented membrane hyperpolarization, thus allowing activation or reactivation of L-type Ca 2+ channels as previously proposed in USMC [4,5,30].…”

“…On the other hand, in smooth muscle cells, Ca 2+ location and affinity represent differentiating parameters [6]. The co-localization of TRPV4 and BKCa channels in the plasma membrane has previously been demonstrated in vascular smooth muscle cells [9,12,27], although this coupling between TRPV4 and BKCa channels has been posulated to likely involve the activation of ryanodine receptorcalcium release channels [12,27]. Of note, ryanodine receptors type 3 has previously been shown not to contribute to the contraction in mouse myometrium [33].…”

“…The putative role of another calcium conductance supported by TRPV4 channels was also assessed, the latter having previously been shown to be involved in the control of other smooth muscle reactivity, mainly from the vascular bed [8,12,13,27]. The pharmacology of such conductance has moreover yielded the synthesis of a new agonist and antagonist [28].…”

“…Moreover, during vaginal delivery, there is no evidence that GKCa activation would be directly under the control of TRPV4 activation. In vivo, during parturition under physiological conditions, the activation of BKCa channels is likely under the control of submembrane Ca 2+ concentrations, phosphorylation levels and membrane-voltage [4,5,27]. The pharmacological activation of TRPV4 by selective agonists could therefore represent anovel strategy to enhance the K + conductance of USMC plasma membranes.…”