Dataset Information

Nitric Oxide Signals Through IRAG to Inhibit TRPM4 Channels and Dilate Cerebral Arteries.

ABSTRACT: Nitric oxide (NO) relaxes vascular smooth muscle cells (SMCs) and dilates blood vessels by increasing intracellular levels of cyclic guanosine monophosphate (cGMP), which stimulates the activity of cGMP-dependent protein kinase (PKG). However, the vasodilator mechanisms downstream of PKG remain incompletely understood. Here, we found that transient receptor potential melastatin 4 (TRPM4) cation channels, which are activated by Ca²⁺ released from the sarcoplasmic reticulum (SR) through inositol triphosphate receptors (IP₃Rs) under native conditions, are essential for SMC membrane depolarization and vasoconstriction. We hypothesized that signaling via the NO/cGMP/PKG pathway causes vasodilation by inhibiting TRPM4. We found that TRPM4 currents activated by stretching the plasma membrane or directly activating IP₃Rs were suppressed by exogenous NO or a membrane-permeable cGMP analog, the latter of which also impaired IP₃R-mediated release of Ca²⁺ from the SR. The effects of NO on TRPM4 activity were blocked by inhibition of soluble guanylyl cyclase or PKG. Notably, upon phosphorylation by PKG, IRAG (IP₃R-associated PKG substrate) inhibited IP₃R-mediated Ca²⁺ release, and knockdown of IRAG expression diminished NO-mediated inhibition of TRPM4 activity and vasodilation. Using superresolution microscopy, we found that IRAG, PKG, and IP₃Rs form a nanoscale signaling complex on the SR of SMCs. We conclude that NO/cGMP/PKG signaling through IRAG inhibits IP₃R-dependent activation of TRPM4 channels in SMCs to dilate arteries.

Significance statement

Nitric oxide is a gaseous vasodilator produced by endothelial cells that is essential for cardiovascular function. Although NO-mediated signaling pathways have been intensively studied, the mechanisms by which they relax SMCs to dilate blood vessels remain incompletely understood. In this study, we show that NO causes vasodilation by inhibiting the activity of Ca²⁺-dependent TRPM4 cation channels. Probing further, we found that NO does not act directly on TRPM4 but instead initiates a signaling cascade that inhibits its activation by blocking the release of Ca²⁺ from the SR. Thus, our findings reveal the essential molecular pathways of NO-induced vasodilation-a fundamental unresolved concept in cardiovascular physiology.

SUBMITTER: Ali S

PROVIDER: S-EPMC8557268 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Nitric Oxide Signals Through IRAG to Inhibit TRPM4 Channels and Dilate Cerebral Arteries.

Ali Sher S Solano Alfredo Sanchez AS Gonzales Albert L AL Thakore Pratish P Krishnan Vivek V Yamasaki Evan E Earley Scott S

Function (Oxford, England) 20211009 6

Nitric oxide (NO) relaxes vascular smooth muscle cells (SMCs) and dilates blood vessels by increasing intracellular levels of cyclic guanosine monophosphate (cGMP), which stimulates the activity of cGMP-dependent protein kinase (PKG). However, the vasodilator mechanisms downstream of PKG remain incompletely understood. Here, we found that transient receptor potential melastatin 4 (TRPM4) cation channels, which are activated by Ca<sup>2+</sup> released from the sarcoplasmic reticulum (SR) through ...[more]

PMID: 34734188

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Project description:Activation of the apelin/APJ receptor signaling system causes endothelium-dependent and nitric oxide (NO)-dependent relaxation in several peripheral arteries. The effects of apelin in cerebral arteries are unknown; however, apelin inhibits voltage-dependent increases in large-conductance, calcium-activated K channel (BKCa) currents in cerebral artery smooth muscle cells. Because NO-induced relaxation of cerebral arteries is mediated, in part, by activation of BKCa channels, the goals of this study were to determine the net effect of apelin in cerebral arteries, as well as test the hypothesis that the actions of apelin in cerebral arteries are secondary to stimulation of APJ receptors. Immunoblot and quantitative reverse transcription polymerase chain reaction analyses detected APJ receptors in cerebral arteries of male Sprague-Dawley rats, and immunofluorescence studies using confocal microscopy confirmed APJ receptor localization in smooth muscle cells. In myograph studies, apelin itself had no direct vasomotor effect but inhibited relaxations to the NO-donor, diethylamine NONOate, and to the endothelium-dependent vasodilator, bradykinin. These effects of apelin were mimicked by the selective BKCa-channel blocker, iberiotoxin, and suppressed by the APJ receptor antagonist, F13A. Apelin also inhibited relaxations evoked by the BKCa-channel openers, NS1619 and BMS 191011, but had no effect on relaxation to levcromakalim, a selective KATP-channel opener. Apelin had no effect on diethylamine NONOate-induced or bradykinin-induced increases in cyclic guanosine monophosphate levels. Patch clamp recordings demonstrated that apelin and iberiotoxin each suppressed the increase in BKCa currents induced by DEA and NS1619 in freshly isolated cerebral artery smooth muscle cells. The results demonstrate that apelin inhibits NO-induced relaxation of cerebral arteries through a mechanism involving activation of APJ receptors and inhibition of BKCa channels in cerebral arterial smooth muscle cells.

Project description:RationalePressure-induced arterial depolarization and constriction (the myogenic response) is a smooth muscle cell (myocyte)-specific mechanism that controls regional organ blood flow and systemic blood pressure. Several different nonselective cation channels contribute to pressure-induced depolarization, but signaling mechanisms involved are unclear. Similarly uncertain is the contribution of anion channels to the myogenic response and physiological functions and mechanisms of regulation of recently discovered transmembrane 16A (TMEM16A), also termed Anoctamin 1, chloride (Cl(-)) channels in arterial myocytes.ObjectiveTo investigate the hypothesis that myocyte TMEM16A channels control membrane potential and contractility and contribute to the myogenic response in cerebral arteries.Methods and resultsCell swelling induced by hyposmotic bath solution stimulated Cl(-) currents in arterial myocytes that were blocked by TMEM16A channel inhibitory antibodies, RNAi-mediated selective TMEM16A channel knockdown, removal of extracellular calcium (Ca(2+)), replacement of intracellular EGTA with BAPTA, a fast Ca(2+) chelator, and Gd(3+) and SKF-96365, nonselective cation channel blockers. In contrast, nimodipine, a voltage-dependent Ca(2+) channel inhibitor, or thapsigargin, which depletes intracellular Ca(2+) stores, did not alter swelling-activated TMEM16A currents. Pressure-induced (-40 mm Hg) membrane stretch activated ion channels in arterial myocyte cell-attached patches that were inhibited by TMEM16A antibodies and were of similar amplitude to recombinant TMEM16A channels. TMEM16A knockdown reduced intravascular pressure-induced depolarization and vasoconstriction but did not alter depolarization-induced (60 mmol/L K(+)) vasoconstriction.ConclusionsMembrane stretch activates arterial myocyte TMEM16A channels, leading to membrane depolarization and vasoconstriction. Data also provide a mechanism by which a local Ca(2+) signal generated by nonselective cation channels stimulates TMEM16A channels to induce myogenic constriction.

Dataset Information

Nitric Oxide Signals Through IRAG to Inhibit TRPM4 Channels and Dilate Cerebral Arteries.

Significance statement

Publications

Nitric Oxide Signals Through IRAG to Inhibit TRPM4 Channels and Dilate Cerebral Arteries.

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