Abstract
Background and Purpose: Arterial diameter is dictated by the contractile state of the vascular smooth muscle cells (VSMCs), which is modulated by direct and indirect inputs from endothelial cells (ECs). Modulators of KCNQ-encoded kV7 channels have considerable impact on arterial diameter and these channels are known to be expressed in VSMCs but not yet defined in ECs. However, expression of kV7 channels in ECs would add an extra level of vascular control. This study aims to characterize the expression and function of KV7 channels within rat mesenteric artery ECs.Experimental Approach: In rat mesenteric artery, KCNQ transcript and KV7 channel protein expression were determined via RT-qPCR, immunocytochemistry, immunohistochemistry and immunoelectron microscopy. Wire myography was used to determine vascular reactivity.Key Results: KCNQ transcript was identified in isolated ECs and VSMCs. KV7.1, KV7.4 and KV7.5 protein expression was determined in both isolated EC and VSMC and in whole vessels. Removal of ECs attenuated vasorelaxation to two structurally different KV7.2-5 activators S-1 and ML213. KIR2 blockers ML133, and BaCl2 also attenuated S-1 or ML213-mediated vasorelaxation in an endothelium-dependent process. KV7 inhibition attenuated receptor-dependent nitric oxide (NO)-mediated vasorelaxation to carbachol, but had no impact on relaxation to the NO donor, SNP.Conclusion and Implications: In rat mesenteric artery ECs, KV7.4 and KV7.5 channels are expressed, functionally interact with endothelial KIR2.x channels and contribute to endogenous eNOS-mediated relaxation. This study identifies KV7 channels as novel functional channels within rat mesenteric ECs and suggests that these channels are involved in NO release from the endothelium of these vessels.
Highlights
KCNQ-encoded KV7 channels are key regulators of arterial reactivity
This study shows that KV7 channels are expressed in rat mesenteric artery endothelial cell (EC) and contribute to both KV7 activator-mediated relaxation via a potential functional interaction with KIR2 channels and endothelial nitric oxide (NO) synthase-dependent axis of carbachol (CCh)-mediated vasorelaxation
Cell isolation efficiency is demonstrated by a reduction in EC marker Pecam within vascular smooth muscle cell (VSMC) cell lysates when compared to ECs (P ≤ 0.05) and a reduction in VSMCs marker α-smooth muscle actin-2 (Acta2) in EC cell lysates when compared to VSMCs (P = 0.065, Figures 1B,C)
Summary
KCNQ-encoded KV7 channels are key regulators of arterial reactivity. Within the vasculature, of the five KCNQ subtypes, KCNQ4 is predominantly expressed, followed by KCNQ5 > KCNQ1; with little to no contribution from KCNQ2/3 (Ohya et al, 2003; Yeung et al, 2007; Ng et al, 2011). In human and rodent blood vessels KV7 channels contribute to resting tone (Ohya et al, 2003; Yeung et al, 2007; Mackie et al, 2008; Ng et al, 2011), whereby their blockers such as linopirdine or XE991 produce contractions or enhance vasoconstrictor responses. Vascular KV7 channel studies have focused predominantly on vascular smooth muscle cells (VSMCs) or whole arteries, and as a result it is currently unclear whether endothelial cells (ECs) express KV7 channels and if so, what their functional role may be. Arterial diameter is dictated by the contractile state of the vascular smooth muscle cells (VSMCs), which is modulated by direct and indirect inputs from endothelial cells (ECs).
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