TRPV4-dependent dilation of peripheral resistance arteries influences arterial pressure

Abstract

Transient receptor potential vanilloid 4 (TRPV4) channels have been implicated as mediators of calcium influx in both endothelial and vascular smooth muscle cells and are potentially important modulators of vascular tone. However, very little is known about the functional roles of TRPV4 in the resistance vasculature or how these channels influence hemodynamic properties. In the present study, we examined arterial vasomotor activity in vitro and recorded blood pressure dynamics in vivo using TRPV4 knockout (KO) mice. Acetylcholine-induced hyperpolarization and vasodilation were reduced by approximately 75% in mesenteric resistance arteries from TRPV4 KO versus wild-type (WT) mice. Furthermore, 11,12-epoxyeicosatrienoic acid (EET), a putative endothelium-derived hyperpolarizing factor, activated a TRPV4-like cation current and hyperpolarized the membrane of vascular smooth muscle cells, resulting in the dilation of mesenteric arteries from WT mice. In contrast, 11,12-EET had no effect on membrane potential, diameter, or ionic currents in the mesenteric arteries from TRPV4 KO mice. A disruption of the endothelium reduced 11,12-EET-induced hyperpolarization and vasodilatation by approximately 50%. A similar inhibition of these responses was observed following the block of endothelial (small and intermediate conductance) or smooth muscle (large conductance) K(+) channels, suggesting a link between 11,12-EET activity, TRPV4, and K(+) channels in endothelial and smooth muscle cells. Finally, we found that hypertension induced by the inhibition of nitric oxide synthase was greater in TRPV4 KO compared with WT mice. These results support the conclusion that both endothelial and smooth muscle TRPV4 channels are critically involved in the vasodilation of mesenteric arteries in response to endothelial-derived factors and suggest that in vivo this mechanism opposes the effects of hypertensive stimuli.

Fig. 1.

ACh-induced hyperpolarization and dilation are suppressed in mesenteric arteries from transient receptor potential vanilloid 4 knockout (TRPV4 KO) vs. wild-type (WT) mice. Arteries were pretreated with N^ω-nitro-l-arginine (l-NNA, 100 μM) and indomethacin (10 μM), constricted to 50% of maximum using phenylephrine (PE) and then exposed to a single dose of ACh (3 μM). Initial diameters and membrane potentials were 157 ± 3 μm and −32 ± 2 mV, respectively, for WT arteries and 178 ± 15 μm and −32 ± 1 mV, respectively, for KO arteries. *P < 0.05 vs. WT; n = 7 WT and 6 TRPV4 KO mice.

Fig. 2.

11,12-Epoxyeicosatrienoic acid (11,12-EET)-induced vasodilation and smooth muscle hyperpolarization are absent in mesenteric arteries from TRPV4 KO mice. Top: vasodilation (%reversal of PE-induced constriction) in response to 11,12-EET (3 μM). Bottom: change in smooth muscle membrane potential (in mV) in response to 11,12-EET (3 μM). +E and −E, data from arteries with and without endothelium, respectively. Significance levels are as indicated. NS, no significant difference. Initial diameters and membrane potentials were 143 ± 16 μm and −32 ± 2 mV for WT + E arteries, 137 ± 10 μm and −34 ± 1 mV for WT − E arteries, 181 ± 17 μm and −31 ± 1 mV for KO + E arteries, and 135 ± 10 μm and −30 ± 1 mV for KO − E arteries, respectively; n = 5 for each group.

Fig. 3.

11,12-EET-activated whole cell currents are absent in vascular smooth muscle cells from TRPV4 KO mice. A: examples of whole cell currents activated by 11,12-EET (3 μM) in mesenteric arterial smooth muscle cells isolated from WT and TRPV4 KO mice. I, current; V, voltage. B: summary data for 11,12-EET-activated currents (difference currents) in WT (n = 5) and TRPV4 KO (n = 3) mouse mesenteric artery myocytes. *P < 0.05 vs. WT.

Fig. 4.

11,12-EET-induced hyperpolarizations are inhibited by combined apamin and 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34), or IBTX alone, and are abolished in the presence of all 3 K⁺ channel blockers. The channel blockers were added both to the arterial lumen and the bath solution. Initial diameters and membrane potentials were, respectively, 127 ± 11 μm and −30 ± 1 mV for control arteries, 152 ± 9 μm and −30 ± 1 mV for arteries treated with apamin (0.3 μM) and TRAM-34 (1 μM), 127 ± 7 μm and −31 ± 1 mV for arteries treated with iberiotoxin (IBTX, 100 nM), and 122 ± 7 μm and −31 ± 1 mV for arteries treated with apamin, TRAM-34, and IBTX. *P < 0.05 vs. control; †P < 0.05 vs. control, apamin + TRAM-34, and IBTX; n = 4 mice for each condition.

Fig. 5.

Nitric oxide synthase inhibition-induced hypertension is greater in TRPV4 KO vs. WT mice. Mean arterial pressure in TRPV4 KO (n = 5–8) and WT (n = 7 to 8) mice before and after the nitric oxide synthase inhibitor l-NNA was added to drinking water. *P ≤ 0.05 vs. WT.