Endothelial-derived hyperpolarization factor (EDHF) contributes to PlGF-induced dilation of mesenteric resistance arteries from pregnant rats

Abstract

The aim of this study was to investigate the cellular mechanism involved in the potent vasodilatory action of PlGF on mesenteric resistance arteries from pregnant rats. PlGF (3 nM) induced a vasodilation of 64 ± 3.8% that was completely abolished by endothelial denudation. Significant dilation (28 ± 4.0%) remained, however, in the presence of nitric oxide synthase and cyclooxygenase inhibition, and was associated with significant reductions in vascular smooth muscle cell calcium. Absence of dilation in potassium-depolarizing solution (30 mM) confirmed its dependence on endothelial-derived hyperpolarization factor. Subsequent studies established that vasodilation was abolished by pharmacologic inhibition of SK(Ca) (apamin) and BK(Ca) (iberiotoxin) but not IK(Ca) (tram-34) potassium channels. In summary, PlGF acts through the release of a combination of endothelium-derived relaxation factors. Based on the results of potassium channel blockade, we suggest that it induces endothelial hyperpolarization via SK(Ca) channel activation; this, in turn, leads to the release of a diffusible mediator that activates vascular smooth muscle BK(Ca) channels, hyperpolarization and vasodilation. This is the first study to identify the mechanism for PlGF/VEGFR-1 resistance artery dilation in the pregnant state, whose attenuation likely contributes to the systemic hypertension characteristic of pre- eclampsia.

Fig. 1

PlGF vasodilation is endothelium dependent, and significant residual dilation remains in the presence of NOS + COX inhibition: intact and endothelium-denuded MRA were preconstricted with phenylephrine prior to being exposed to PlGF (3 nM). Intact arteries were used in the presence of inhibitors of NOS (L-L = L-NAME + L-NNA; each at 100 μM) and cyclooxygenase (I = indomethacin at 10 μM). Data are reported as means ± SEM; n = 4; ∗ p < 0.01, ∗∗ p < 0.001.

Fig. 2

PlGF vasodilation is associated with a reduction in VSM calcium: original trace demonstrating reductions in SMC [Ca²⁺]_i and dilation of MRA in response to application of 3 nM PlGF or 1 μM ACh.

Fig. 3

Effects of repeated PlGF application on arterial VSM calcium and diameter responses: graph summarizing the vasodilatory (a) and SMC [Ca²⁺]_i responses (b) of mesenteric arteries to successive application of 3 nM PlGF. A time of 15 min was necessary to wash out the first application of PlGF prior to any readdition. The effects of 1 μM ACh are shown for comparison. Vasodilation is expressed as a percentage of maximal response obtained in papaverine and diltiazem. L-NAME, L-NNA and indomethacin were present throughout all experiments. Data are reported as mean ± SEM, n = 7. ∗ p < 0.05.

Fig. 4

Complete inhibition of PlGF vasodilation by potassium-induced depolarization: intact MRA were preconstricted with high-potassium (High K⁺ = 30 mM) depolarizing solution prior to being exposed to PlGF (3 nM). L-NAME, L-NNA and indomethacin (L-L-I.) were present throughout all experiments. Data are shown as means ± SEM; n = 4; ∗ p < 0.05.

Fig. 5

PlGF vasodilation is potassium channel dependent: intact, Phe-preconstricted MRA were pre-treated with L-NNA + L-NAME (L-L, 100 μM) and indomethacin (I, 10 μM) along with combinations of potassium channel inhibitors: apamin (Apa, for SK_Ca channels), charybdotoxin (ChTx, for IK_Ca and BK_Ca channels), iberiotoxin (IbTx, for Bk_Ca channels) and TRAM-34 (for IK_Ca channels), prior to the application of PlGF (3 nM). Data are reported as mean ± SEM; n = 4; ∗ p < 0.001.