Nitric oxide suppresses vascular voltage-gated T-type Ca2+ channels through cGMP/PKG signaling

2014 ◽  
Vol 306 (2) ◽  
pp. H279-H285 ◽  
Author(s):  
Osama F. Harraz ◽  
Suzanne E. Brett ◽  
Donald G. Welsh
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Channel Blocker ◽  
Inhibitory Effect ◽  
No Donors ◽  
Channel Current ◽  
Steady State Inactivation ◽  
Arterial Smooth Muscle Cells ◽  
Patch Clamp Electrophysiology ◽  
Ca2 Channels

Recent reports have noted that T-type Ca2+ channels (CaV3.x) are expressed in vascular smooth muscle and are potential targets of regulation. In this study, we examined whether and by what mechanism nitric oxide (NO), a key vasodilator, influences this conductance. Using patch-clamp electrophysiology and rat cerebral arterial smooth muscle cells, we monitored an inward Ba2+ current that was divisible into a nifedipine-sensitive and -insensitive component. The latter was abolished by T-type channel blocker and displayed classic T-type properties including faster activation and steady-state inactivation at hyperpolarized potentials. NO donors (sodium nitroprusside, S-nitroso- N-acetyl- dl-penicillamine), along with activators of protein kinase G (PKG) signaling, suppressed T-type currents. Inhibitors of guanylyl cyclase/PKG {1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and KT5823, respectively}, had no effect on basal currents; KT5823 did, however, mask T-type Ca2+ channel current inhibition by NO/PKG. Functional experiments confirmed an inhibitory effect for NO on the T-type contribution to cerebral arterial myogenic tone. Cumulatively, our findings support the view that T-type Ca2+ channels are a regulatory target of vasodilatory signaling pathways. This targeting will influence Ca2+ dynamics and consequent tone development in the cerebral circulation.

Nitric oxide inhibits Ca2+ mobilization through cADP-ribose signaling in coronary arterial smooth muscle cells

2000 ◽  
Vol 279 (3) ◽  
pp. H873-H881 ◽  
Author(s):  
Jiang-Zhou Yu ◽  
David X. Zhang ◽  
Ai-Ping Zou ◽  
William B. Campbell ◽  
Pin-Lan Li
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Inhibitory Effect ◽  
Cyclase Activity ◽  
Arterial Smooth Muscle ◽  
Adp Ribosyl Cyclase ◽  
Intracellular Ca ◽  
Arterial Smooth Muscle Cells

The present study was designed to determine whether the cADP-ribose-mediated Ca2+ signaling is involved in the inhibitory effect of nitric oxide (NO) on intracellular Ca2+ mobilization. With the use of fluorescent microscopic spectrometry, cADP-ribose-induced Ca2+ release from sarcoplasmic reticulum (SR) of bovine coronary arterial smooth muscle cells (CASMCs) was determined. In the α-toxin-permeabilized primary cultures of CASMCs, cADP-ribose (5 μM) produced a rapid Ca2+ release, which was completely blocked by pretreatment of cells with the cADP-ribose antagonist 8-bromo-cADP-ribose (8-Br-cADPR). In intact fura 2-loaded CASMCs, 80 mM KCl was added to depolarize the cells and increase intracellular Ca2+ concentration ([Ca2+]i). Sodium nitroprusside (SNP), an NO donor, produced a concentration-dependent inhibition of the KCl-induced increase in [Ca2+]i, but it had no effect on the U-46619-induced increase in [Ca2+]i. In the presence of 8-Br-cADPR (100 μM) and ryanodine (10 μM), the inhibitory effect of SNP was markedly attenuated. HPLC analyses showed that CASMCs expressed the ADP-ribosyl cyclase activity, and SNP (1–100 μM) significantly reduced the ADP-ribosyl cyclase activity in a concentration-dependent manner. The effect of SNP was completely blocked by addition of 10 μM oxygenated hemoglobin. We conclude that ADP-ribosyl cyclase is present in CASMCs, and NO may decrease [Ca2+]i by inhibition of cADP-ribose-induced Ca2+ mobilization.

Mechanisms of cyclic nucleotide-induced relaxation in canine tracheal smooth muscle

1995 ◽  
Vol 268 (3) ◽  
pp. L407-L413 ◽  
Author(s):  
I. McGrogan ◽  
S. Lu ◽  
S. Hipworth ◽  
L. Sormaz ◽  
R. Eng ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Cyclic Nucleotide ◽  
Cyclopiazonic Acid ◽  
Inhibitory Effect ◽  
Beta Agonist ◽  
Cyclic Monophosphate ◽  
Median Effective Concentration ◽  
Camp Levels ◽  
Ca2 Channels

The effects of exogeneous cyclopiazonic acid (CPA, 10 microM), a selective inhibitor of the sarcoplasmic reticulum (SR) Ca2+ adenosinetriphosphatase, on cyclic nucleotide-induced relaxations of canine airway smooth muscle were examined. Strips of tracheal muscle were precontracted with carbachol (50% median effective concentration, 0.1 microM) or with 60 mM KCl. The beta-agonist isoproterenol (ISO, 10 microM) relaxed the tissue by approximately 50%. The relaxation was reduced in the presence of CPA when L-type Ca2+ channels were available but not when these were blocked by 0.1 microM nifedipine. Forskolin (1.0 microM), an adenylate cyclase activator, was less effective at inhibiting the contraction than ISO, and addition of CPA did not block its inhibitory effect as effectively as when ISO was used. Radioimmunoassay indicated that both these agents raised adenosine 3',5'-cyclic monophosphate (cAMP) levels to the same degree. Very little relaxation of the precontracted smooth muscle was elicited by 3 mM 8-bromo-adenosine 3',5'-cyclic monophosphate (8-BrcAMP), and addition of CPA had no effect. Sodium nitroprusside (100 microM) and 8-bromo-guanosine 3',5'-cyclic monophosphate (10 mM) inhibited contraction to a greater degree than any agent that raised cAMP. These inhibitions were greatly reduced in the presence of CPA when L-type Ca2+ channels were available. We conclude that pumping of Ca2+ into SR plays a major role guanosine 3',5'-cyclic monophosphate-produced but not cAMP-induced relaxation; L-type Ca2+ channels must be available for the relaxant role of Ca2+ pumping into the SR to be expressed; and ISO-induced relaxation may not involve primarily elevation of the cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Nitric Oxide Inhibits L-Type Ca2+ Current in Glomus Cells of the Rabbit Carotid Body Via a cGMP-Independent Mechanism

1999 ◽  
Vol 81 (4) ◽  
pp. 1449-1457 ◽  
Author(s):  
Beth A. Summers ◽  
Jeffrey L. Overholt ◽  
Nanduri R. Prabhakar
Keyword(s):  
Nitric Oxide ◽  
Carotid Body ◽  
Channel Blocker ◽  
Adult Rabbit ◽  
Subsequent Formation ◽  
No Donors ◽  
Glomus Cells ◽  
Channel Proteins ◽  
Carotid Bodies ◽  
Independent Mechanism

Nitric oxide inhibits L-type Ca2+ current in glomus cells of the rabbit carotid body via a cGMP-independent mechanism. Previous studies have shown that nitric oxide (NO) inhibits carotid body sensory activity. To begin to understand the cellular mechanisms associated with the actions of NO in the carotid body, we monitored the effects of NO donors on the macroscopic Ca2+ current in glomus cells isolated from rabbit carotid bodies. Experiments were performed on freshly dissociated glomus cells from adult rabbit carotid bodies using the whole cell configuration of the patch-clamp technique. The NO donors sodium nitroprusside (SNP; 600 μM, n = 7) and spermine nitric oxide (SNO; 100 μM, n = 7) inhibited the Ca2+ current in glomus cells in a voltage-independent manner. These effects of NO donors were rapid in onset and peaked within 1 or 2 min. In contrast, the outward K+ current was unaffected by SNP (600 μM, n = 6), indicating that the inhibition by SNP was not a nonspecific membrane effect. 2-(4-carboxyphenyl)-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (carboxy-PTIO; 500 μM), an NO scavenger, prevented inhibition of the Ca2+ current by SNP ( n = 7), whereas neither superoxide dismutase (SOD; 2,000 U/ml, n = 4), a superoxide scavenger, nor sodium hydrosulfite (SHS; 1 mM, n = 7), a reducing agent, prevented inhibition of the Ca2+ current by SNP. However, SNP inhibition of the Ca2+ current was reversible in the presence of either SOD or SHS. These results suggest that NO itself inhibits Ca2+current in a reversible manner and that subsequent formation of peroxynitrites results in irreversible inhibition. SNP inhibition of the Ca2+ current was not affected by 30 μM LY 83,583 ( n = 7) nor was it mimicked by 600 μM 8-bromoguanosine 3′:5′-cyclic monophosphate (8-Br-cGMP; n = 6), suggesting that the effects of NO on the Ca2+ current are mediated, in part, via a cGMP-independent mechanism. N-ethylmaleimide (NEM; 2.5 mM, n= 6) prevented the inhibition of the Ca2+ current by SNP, indicating that SNP is acting via a modification of sulfhydryl groups on Ca2+ channel proteins. Norepinephrine (NE; 10 μM) further inhibited the Ca2+ current in the presence of NEM ( n = 7), implying that NEM did not nonspecifically eliminate Ca2+ current modulation. Nisoldipine, an L-type Ca2+ channel blocker (2 μM, n = 6), prevented the inhibition of Ca2+ current by SNP, whereas ω-conotoxin GVIA, an N-type Ca2+ channel blocker (1 μM, n = 9), did not prevent the inhibition of Ca2+ current by SNP. These results demonstrate that NO inhibits L-type Ca2+ channels in adult rabbit glomus cells, in part, due to a modification of calcium channel proteins. The inhibition might provide one plausible mechanism for efferent inhibition of carotid body activity by NO.

Two types of calcium channels in isolated smooth muscle cells from rat tail artery

1989 ◽  
Vol 256 (5) ◽  
pp. H1361-H1368 ◽  
Author(s):  
R. Wang ◽  
E. Karpinski ◽  
P. K. Pang
Keyword(s):  
Smooth Muscle ◽  
Steady State ◽  
Calcium Channels ◽  
Rat Tail Artery ◽  
Inward Current ◽  
Tail Artery ◽  
Channel Current ◽  
Steady State Inactivation ◽  
Channel Currents ◽  
Rat Tail

Whole cell patch-clamp recordings were carried out on smooth muscle cells from rat tail artery in short-term culture to verify the existence of and to characterize the calcium channels that are present. Two types of voltage-dependent calcium channels were identified in 55 of 63 cells studied. The T-type calcium channel was activated at -50 mV, and the peak inward current occurred at -10 mV, whereas the L-type channel was activated at -20 mV, and the peak inward current occurred at +10 or +20 mV. The T-type channel current inactivated quickly in contrast to the much slower inactivation of the L-channel current. The voltage dependence of steady-state inactivation of the two channels was similar to that reported for other vascular smooth muscle preparations. An internal solution containing Cs2-aspartate maintained the calcium-channel currents for at least 20 min with only a 5-10% decline. BAY K 8644 had no effect on T-channel currents, but the L-channel current was increased by at least a factor of two. In addition, BAY K 8644 shifted the activation threshold, the peak inward current, and the steady-state inactivation-activation curves of L-type channel currents in the direction of hyperpolarization.

scholarly journals Development of Photoinitiated Nitric Oxide Releasing Polymer Films for Controlled Drug Delivery

2009 ◽  
Vol 3 (2) ◽  
Author(s):  
D. M. Smeenge ◽  
M. J. Barron ◽  
M. T. Nielsen ◽  
J. Goldman ◽  
M. C. Frost
Keyword(s):  
Nitric Oxide ◽  
Drug Delivery ◽  
Smooth Muscle ◽  
Cell Cultures ◽  
Polymer Films ◽  
Surface Flux ◽  
Surface Fluxes ◽  
No Donors ◽  
Polymeric Systems ◽  
Smooth Muscle Cell Cultures

Nitric Oxide (NO) is small, free radical gas that has been shown to have a wide variety of physiological functions, including the ability to hinder tumor angiogenesis at high, but non lethal, concentrations [1]. Previous work suggests that if NO could be effectively delivered in vivo to tumors of patients currently undergoing chemotherapy treatments at the appropriate levels, less damaging chemotherapy treatments could be used against cancer [2]. This could increase the overall survivability of cancer patients, especially in those prone to the harmful effects of chemotherapy: children, elderly, and those of weak immune systems. If NO is especially successful at preventing and eliminating tumor growth, angiogenesis, and carcinogenesis the need for stressful chemotherapy treatments could be eliminated altogether. This project is focused on developing novel photosensitive NO donors that can be incorporated into polymeric systems and used in a fiber optic drug delivery system. Development of these NO-releasing polymers will allow continued investigation of NO's role in tumor death by precisely controlling the surface flux of NO that cells are exposed to. Generating specific surface fluxes of NO from polymer films has been demonstrated by using polymer films that contain photoinitiated NO donors [3], prepared by synthesizing S-nitrosothiol (RSNO) derivitized polymer fillers that are blended into hydrophobic polymers and cast into a film. These films generate and sustain a surface flux of NO based on the wavelength and intensity of light used [3]. Polymers releasing NO are more promising as an NO donor than simply injecting NO into samples because they allow for spatial and temporal control of NO delivery. The specific concentration of NO needed to produce desirable effects on tumor cells (i.e., apoptosis) is not known. Data will be presented that show the synthesis and NO-release properties of novel RSNOs based on the nitrosation of benzyl mercaptan thiols. Specifically, UV-Vis spectrum of benzyl mercaptan in toluene and S-nitrosobenzyl mercaptan after the addition of t-butyl nitrite will be presented. We are currently investigating the effects of varying NO-surface fluxes generated from photolytic NO donating polymer films on aortic smooth muscle cell cultures obtained from mice. Once we have established that we can quantitatively determine the effects of different levels of NO on the proliferation of smooth muscle cell cultures, work will begin to apply this methodology and these novel NO-releasing polymeric systems to begin investigating what durations and surface fluxes of NO are necessary to have tumorcidal effects on specific cancer cells.

Nitric oxide mediates the neural nonadrenergic, noncholinergic relaxation of pig tracheal smooth muscle

1992 ◽  
Vol 262 (4) ◽  
pp. L511-L514 ◽  
Author(s):  
M. S. Kannan ◽  
D. E. Johnson
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Electrical Field Stimulation ◽  
Selective Inhibition ◽  
Inhibitory Effect ◽  
Isometric Tension ◽  
Tracheal Smooth Muscle ◽  
Basal Tone ◽  
Dependent Inhibition ◽  
Nanc Relaxation

In pig tracheal smooth muscle, the isometric tension responses to electrical field stimulation (EFS) were studied after raising the tone with carbamylcholine chloride (carbachol). EFS induced frequency-dependent relaxations that were nonadrenergic, noncholinergic (NANC) in nature. Addition of NG-nitro-L-arginine (L-NOArg), an inhibitor of nitric oxide (NO) synthesis from L-arginine, resulted in concentration-dependent inhibition of the relaxation response to EFS. Pretreatment of the tissues with L-arginine (1 mM) prevented the inhibitory effect of L-NOArg on the EFS-induced relaxations at the frequencies studied. However, in the presence of D-arginine, EFS-induced relaxations were inhibited by L-NOArg. L-Arginine, D-arginine, and L-NOArg had no significant effects on basal tone of the muscle. In the presence of L-NOArg, vasoactive intestinal polypeptide (3 x 10(-7) M), the nicotinic agonist dimethylphenyl piperazinium bromide (100 microM), and isoproterenol (1 microM) relaxed carbachol-induced tone. The concentration-dependent selective inhibition of neural relaxation by L-NOArg and its reversal by L-arginine in a stereospecific manner are consistent with NO-mediated NANC relaxation of pig tracheal smooth muscle.

Adenosine receptor-mediated relaxation of rabbit airway smooth muscle: a role for nitric oxide

1997 ◽  
Vol 273 (3) ◽  
pp. L581-L587 ◽  
Author(s):  
S. Ali ◽  
W. J. Metzger ◽  
H. A. Olanrewaju ◽  
S. J. Mustafa
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Epithelial Cells ◽  
Airway Smooth Muscle ◽  
Adenosine Receptor ◽  
Inhibitory Effect ◽  
No Production ◽  
Relaxant Effect ◽  
Adenosine Receptor Agonists ◽  
Cgs 21680

In this study, we investigated the relaxant effect of adenosine receptor agonists on KCl-precontracted airway smooth muscle from rabbits and characterized the type of receptor involved in bronchorelaxation in the presence and absence of epithelium. We further defined the role of epithelium-derived relaxing factor, i.e., nitric oxide (NO), on these responses. In both epithelium-intact and -denuded tertiary airway rings from rabbits, the adenosine receptor agonists 2-[p-(2-carboxyethyl)]phenylethylamino-5-N'-ethylcarboxamidoadenos ine (CGS-21680), 5'-(N-ethyl-carboxamido)adenosine (NECA), 2-chloroadenosine (CAD), and (-)-N6-(2-phenylisopropyl)adenosine (R-PIA) relaxed airway smooth muscle with a potency order of CGS-21680 > NECA > CAD > R-PIA. A 98.5, 89.7, 73.2, and 64.7% relaxation was observed at 10(-5) M by CGS-21680, NECA, CAD, and R-PIA in the epithelium-intact bronchial rings, respectively. The 50% maximum effective concentration (EC50; x 10(-7) M) values for CGS-21680, NECA, CAD, and R-PIA were 2, 4, 9, and 80, respectively. Denuded rings, however, showed much less relaxant responses to various adenosine agonists compared with epithelium-intact rings. The adenosine receptor antagonist 8-(sulfophenyl)theophylline significantly attenuated the relaxant responses to all the agonists in the epithelium-intact and -denuded rings. The epithelium-dependent relaxant effect of the agonists in airway rings was inhibited by NG-monomethyl-L-arginine (L-NMMA; 30 microM). The EC50 (x 10(-6) M) values for CGS-21680, NECA, CAD, and R-PIA in the presence of inhibitor were 5.5, 8, 30, and 200, respectively. The L-NMMA produced an insignificant inhibitory effect in the epithelium-denuded rings. L-Arginine but not D-arginine (100 microM) reversed the inhibitory effect of L-NMMA on adenosine agonist-induced relaxation. In primary epithelial cells in culture, CGS-21680 (10(-5) M) induced a fourfold increase in NO production over the control. The CGS-21680-induced NO production in epithelial cells was significantly inhibited by NG-nitro-L-arginine methyl ester (L-NAME). Moreover, L-arginine reversed the inhibitory effect of L-NAME in the epithelial cells. The data suggest that adenosine relaxes rabbit airway smooth muscle through an A2 adenosine receptor and the epithelium serves as a source of NO.

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