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Reactive oxygen species-mediated TRPC6 protein activation in vascular myocytes, a mechanism for vasoconstrictor-regulated vascular tone.

ABSTRACT: Both TRPC6 and reactive oxygen species (ROS) play an important role in regulating vascular function. However, their interplay has not been explored. The present study examined whether activation of TRPC6 in vascular smooth muscle cells (VSMCs) by ROS was a physiological mechanism for regulating vascular tone by vasoconstrictors. In A7r5 cells, arginine vasopressin (AVP) evoked a striking Ca(2+) entry response that was significantly attenuated by either knocking down TRPC6 using siRNA or inhibition of NADPH oxidases with apocynin or diphenyleneiodonium. Inhibition of TRPC6 or ROS production also decreased AVP-stimulated membrane currents. In primary cultured aortic VSMCs, catalase and diphenyleneiodonium significantly suppressed AVP- and angiotensin II-induced whole cell currents and Ca(2+) entry, respectively. In freshly isolated and endothelium-denuded thoracic aortas, hyperforin (an activator of TRPC6), but not its vehicle, induced dose- and time-dependent constriction in TRPC6 wide type (WT) mice. This response was not observed in TRPC6 knock-out (KO) mice. Consistent with the ex vivo study, hyperforin stimulated a robust Ca(2+) entry in the aortic VSMCs from WT mice but not from KO mice. Phenylephrine induced a dose-dependent contraction of WT aortic segments, and this response was inhibited by catalase. Moreover, H(2)O(2) itself evoked Ca(2+) influx and inward currents in A7r5 cells, and these responses were significantly attenuated by either inhibition of TRPC6 or blocking vesicle trafficking. H(2)O(2) also induced inward currents in primary VSMCs from WT but not from TRPC6 KO mice. Additionally, H(2)O(2) stimulated a dose-dependent constriction of the aortas from WT mice but not from the vessels of KO mice. Furthermore, TIRFM showed that H(2)O(2) triggered membrane trafficking of TRPC6 in A7r5 cells. These results suggest a new signaling pathway of ROS-TRPC6 in controlling vessel contraction by vasoconstrictors.

SUBMITTER: Ding Y

PROVIDER: S-EPMC3173128 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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Reactive oxygen species-mediated TRPC6 protein activation in vascular myocytes, a mechanism for vasoconstrictor-regulated vascular tone.

Ding Yanfeng Y Winters Ali A Ding Min M Graham Sarabeth S Akopova Irina I Muallem Shmuel S Wang Yanxia Y Hong Jeong Hee JH Gryczynski Zygmunt Z Yang Shao-Hua SH Birnbaumer Lutz L Ma Rong R

The Journal of biological chemistry 20110715 36

Both TRPC6 and reactive oxygen species (ROS) play an important role in regulating vascular function. However, their interplay has not been explored. The present study examined whether activation of TRPC6 in vascular smooth muscle cells (VSMCs) by ROS was a physiological mechanism for regulating vascular tone by vasoconstrictors. In A7r5 cells, arginine vasopressin (AVP) evoked a striking Ca(2+) entry response that was significantly attenuated by either knocking down TRPC6 using siRNA or inhibiti ...[more]

PMID: 21768109

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Project description:BackgroundOxidative stress is causally linked to the progression of heart failure, and mitochondria are critical sources of reactive oxygen species in failing myocardium. We previously observed that in heart failure, elevated cytosolic Na(+) ([Na(+)](i)) reduces mitochondrial Ca(2+) ([Ca(2+)](m)) by accelerating Ca(2+) efflux via the mitochondrial Na(+)/Ca(2+) exchanger. Because the regeneration of antioxidative enzymes requires NADPH, which is indirectly regenerated by the Krebs cycle, and Krebs cycle dehydrogenases are activated by [Ca(2+)](m), we speculated that in failing myocytes, elevated [Na(+)](i) promotes oxidative stress.Methods and resultsWe used a patch-clamp-based approach to simultaneously monitor cytosolic and mitochondrial Ca(2+) and, alternatively, mitochondrial H(2)O(2) together with NAD(P)H in guinea pig cardiac myocytes. Cells were depolarized in a voltage-clamp mode (3 Hz), and a transition of workload was induced by beta-adrenergic stimulation. During this transition, NAD(P)H initially oxidized but recovered when [Ca(2+)](m) increased. The transient oxidation of NAD(P)H was closely associated with an increase in mitochondrial H(2)O(2) formation. This reactive oxygen species formation was potentiated when mitochondrial Ca(2+) uptake was blocked (by Ru360) or Ca(2+) efflux was accelerated (by elevation of [Na(+)](i)). In failing myocytes, H(2)O(2) formation was increased, which was prevented by reducing mitochondrial Ca(2+) efflux via the mitochondrial Na(+)/Ca(2+) exchanger.ConclusionsBesides matching energy supply and demand, mitochondrial Ca(2+) uptake critically regulates mitochondrial reactive oxygen species production. In heart failure, elevated [Na(+)](i) promotes reactive oxygen species formation by reducing mitochondrial Ca(2+) uptake. This novel mechanism, by which defects in ion homeostasis induce oxidative stress, represents a potential drug target to reduce reactive oxygen species production in the failing heart.

Dataset Information

Reactive oxygen species-mediated TRPC6 protein activation in vascular myocytes, a mechanism for vasoconstrictor-regulated vascular tone.

Publications

Reactive oxygen species-mediated TRPC6 protein activation in vascular myocytes, a mechanism for vasoconstrictor-regulated vascular tone.

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