scholarly journals Cardiomyocyte depolarization triggers NOS-dependent NO transient after calcium release, reducing the subsequent calcium transient

2021 ◽  
Vol 116 (1) ◽  
Author(s):  
Matias Mosqueira ◽  
Roland Konietzny ◽  
Carolin Andresen ◽  
Chao Wang ◽  
Rainer H.A. Fink
Keyword(s):  
Calcium Release ◽  
Soluble Guanylate Cyclase ◽  
Calcium Transient ◽  
Inhibitory Effect ◽  
Transient Signal ◽  
No Production ◽  
Millisecond Time Scale ◽  
No Signaling ◽  
Novel Strategy ◽  
Cardiac Excitation

AbstractCardiac excitation–contraction coupling and metabolic and signaling activities are centrally modulated by nitric oxide (NO), which is produced by one of three NO synthases (NOSs). Despite the significant role of NO in cardiac Ca2+ homeostasis regulation under different pathophysiological conditions, such as Duchenne muscular dystrophy (DMD), no precise method describes the production, source or effect of NO through two NO signaling pathways: soluble guanylate cyclase-protein kinase G (NO-sGC-PKG) and S-nitrosylation (SNO). Using a novel strategy involving isolated murine cardiomyocytes loaded with a copper-based dye highly specific for NO, we observed a single transient NO production signal after each electrical stimulation event. The NO transient signal started 67.5 ms after the beginning of Rhod-2 Ca2+ transient signal and lasted for approximately 430 ms. Specific NOS isoform blockers or NO scavengers significantly inhibited the NO transient, suggesting that wild-type (WT) cardiomyocytes produce nNOS-dependent NO transients. Conversely, NO transient in mdx cardiomyocyte, a mouse model of DMD, was dependent on inducible NOS (iNOS) and endothelial (eNOS). In a consecutive stimulation protocol, the nNOS-dependent NO transient in WT cardiomyocytes significantly reduced the next Ca2+ transient via NO-sGC-PKG. In mdx cardiomyocytes, this inhibitory effect was iNOS- and eNOS-dependent and occurred through the SNO pathway. Basal NO production was nNOS- and iNOS-dependent in WT cardiomyocytes and eNOS- and iNOS-dependent in mdx cardiomyocytes. These results showed cardiomyocyte produces NO isoform-dependent transients upon membrane depolarization at the millisecond time scale activating a specific signaling pathway to negatively modulate the subsequent Ca2+ transient.

α2-Adrenergic-mediated tubular NO production inhibits thick ascending limb chloride absorption

2001 ◽  
Vol 281 (4) ◽  
pp. F679-F686 ◽  
Author(s):  
Craig F. Plato ◽  
Jeffrey L. Garvin
Keyword(s):  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
Chloride Transport ◽  
Receptor Activation ◽  
Inhibitory Effect ◽  
No Production ◽  
Thick Ascending Limb ◽  
Chloride Absorption ◽  
Ly 83583

Stimulation of α2-adrenergic receptors inhibits transport in various nephron segments, and the thick ascending limb of the loop of Henle (THAL) expresses α2-receptors. We hypothesized that selective α2-receptor activation decreases NaCl absorption by cortical THALs through activation of NOS and increased production of NO. We found that the α2-receptor agonist clonidine (10 nM) decreased chloride flux ( J Cl) from 119.5 ± 15.9 to 67.4 ± 13.8 pmol · mm−1 · min−1 (43% reduction; P < 0.02), whereas removal of clonidine from the bath increased J Cl by 20%. When NOS activity was inhibited by pretreatment with 5 mM N G-nitro-l-arginine methyl ester, the inhibitory effects of clonidine on THAL J Clwere prevented (81.7 ± 10.8 vs. 71.6 ± 6.9 pmol · mm−1 · min−1). Similarly, when the NOS substrate l-arginine was deleted from the bath, addition of clonidine did not decrease THAL J Cl from control (106.9 ± 11.6 vs. 132.2 ± 21.3 pmol · mm−1 · min−1). When we blocked the α2-receptors with rauwolscine (1 μM), we found that the inhibitory effect of 10 nM clonidine on THAL J Cl was abolished, verifying that α2, rather than I1, receptors mediate the effects of clonidine in the THAL. We investigated the mechanism of NOS activation and found that intracellular calcium concentration did not increase in response to clonidine, whereas pretreatment with 150 nM wortmannin abolished the clonidine-mediated inhibition of THAL J Cl, indicating activation of phosphatidylinositol 3-kinase and the Akt pathway. We found that pretreatment of THALs with 10 μM LY-83583, an inhibitor of soluble guanylate cyclase, blocked clonidine-mediated inhibition of THAL J Cl. In conclusion, α2-receptor stimulation decreases THAL J Cl by increasing NO release and stimulating guanylate cyclase. These data suggest that α2-receptors act as physiological regulators of THAL NO synthesis, thus inhibiting chloride transport and participating in the natriuretic and diuretic effects of clonidine in vivo.

scholarly journals Hydrogen sulfide, an enhancer of vascular nitric oxide signaling: mechanisms and implications

2017 ◽  
Vol 312 (1) ◽  
pp. C3-C15 ◽  
Author(s):  
Csaba Szabo
Keyword(s):  
Nitric Oxide ◽  
Hydrogen Sulfide ◽  
Soluble Guanylate Cyclase ◽  
Specific Activity ◽  
Cardiovascular Effects ◽  
Biologically Active ◽  
Reduced Form ◽  
No Production ◽  
Nitric Oxide Signaling ◽  
No Signaling

Nitric oxide (NO) vascular signaling has long been considered an independent, self-sufficient pathway. However, recent data indicate that the novel gaseous mediator, hydrogen sulfide (H2S), serves as an essential enhancer of vascular NO signaling. The current article overviews the multiple levels at which this enhancement takes place. The first level of interaction relates to the formation of biologically active hybrid S/N species and the H2S-induced stimulation of NO release from its various stable “pools” (e.g., nitrite). The next interactions occur on the level of endothelial calcium mobilization and PI3K/Akt signaling, increasing the specific activity of endothelial NO synthase (eNOS). The next level of interaction occurs on eNOS itself; H2S directly interacts with the enzyme: sulfhydration of critical cysteines stabilizes it in its physiological, dimeric state, thereby optimizing eNOS-derived NO production and minimizing superoxide formation. Yet another level of interaction, further downstream, occurs at the level of soluble guanylate cyclase (sGC): H2S stabilizes sGC in its NO-responsive, physiological, reduced form. Further downstream, H2S inhibits the vascular cGMP phosphodiesterase (PDE5), thereby prolonging the biological half-life of cGMP. Finally, H2S-derived polysulfides directly activate cGMP-dependent protein kinase (PKG). Taken together, H2S emerges an essential endogenous enhancer of vascular NO signaling, contributing to vasorelaxation and angiogenesis. The functional importance of the H2S/NO cooperative interactions is highlighted by the fact that H2S loses many of its beneficial cardiovascular effects when eNOS is inactive.

scholarly journals The Role of Orai1 in Regulating Sarcoplasmic Calcium Release, Mitochondrial Morphology and Function in Myostatin Deficient Skeletal Muscle

2020 ◽  
Vol 11 ◽  
Author(s):  
Mónika Sztretye ◽  
Zoltán Singlár ◽  
Norbert Balogh ◽  
Gréta Kis ◽  
Péter Szentesi ◽  
...  
Keyword(s):  
Neuromuscular Junction ◽  
Muscle Weakness ◽  
Calcium Release ◽  
Calcium Transient ◽  
Inhibitory Effect ◽  
Mitochondrial Morphology ◽  
Loss Of Function ◽  
Myostatin Gene ◽  
Store Operated Calcium Entry ◽  
And Function

In mice a naturally occurring 12-bp deletion in the myostatin gene is considered responsible for the compact phenotype (MstnCmpt–dl1Abc, Cmpt) labeled by a tremendous increase in body weight along with signs of muscle weakness, easier fatigability, decreased Orai1 expression and store operated calcium entry (SOCE). Here, on the one hand, Cmpt fibers were reconstructed with venus-Orai1 but this failed to restore SOCE. On the other hand, the endogenous Orai1 was silenced in fibers from wild type C57Bl6 mice which resulted in ∼70% of Orai1 being silenced in whole muscle homogenates as confirmed by Western blot, accompanied by an inhibitory effect on the voltage dependence of SR calcium release that manifested in a slight shift toward more positive potential values. This maneuver completely hampered SOCE. Our observations are consistent with the idea that Orai1 channels are present in distinct pools responsible for either a rapid refilling of the SR terminal cisternae connected to each voltage-activated calcium transient, or a slow SOCE associated with an overall depletion of calcium in the SR lumen. Furthermore, when Cmpt cells were loaded with the mitochondrial membrane potential sensitive dye TMRE, fiber segments with depolarized mitochondria were identified covering on average 26.5 ± 1.5% of the fiber area. These defective areas were located around the neuromuscular junction and displayed significantly smaller calcium transients. The ultrastructural analysis of the Cmpt fibers revealed changes in the mitochondrial morphology. In addition, the mitochondrial calcium uptake during repetitive stimulation was higher in the Cmpt fibers. Our results favor the idea that reduced function and/or expression of SOCE partners (in this study Orai1) and mitochondrial defects could play an important role in muscle weakness and degeneration associated with certain pathologies, perhaps including loss of function of the neuromuscular junction and aging.

Inhibitory effect of isolated constituents from sclerotia of Poria cocos on LPS-induced NO production

Planta Medica ◽  
2016 ◽  
Vol 81 (S 01) ◽  
pp. S1-S381
Author(s):  
SR Lee ◽  
S Lee ◽  
HJ Eom ◽  
HR Kang ◽  
JS Yu ◽  
...  
Keyword(s):  
Inhibitory Effect ◽  
No Production ◽  
Poria Cocos

Hypothermia-induced cardioprotection using extended ischemia and early reperfusion cooling

2007 ◽  
Vol 292 (4) ◽  
pp. H1995-H2003 ◽  
Author(s):  
Zuo-Hui Shao ◽  
Wei-Tien Chang ◽  
Kim Chai Chan ◽  
Kim R. Wojcik ◽  
Chin-Wang Hsu ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Therapeutic Hypothermia ◽  
Optimal Timing ◽  
No Production ◽  
Early Reperfusion ◽  
No Signaling ◽  
Nos Inhibitor ◽  
Oxide Synthase ◽  
Pkc Activation

Optimal timing of therapeutic hypothermia for cardiac ischemia is unknown. Our prior work suggests that ischemia with rapid reperfusion (I/R) in cardiomyocytes can be more damaging than prolonged ischemia alone. Also, these cardiomyocytes demonstrate protein kinase C (PKC) activation and nitric oxide (NO) signaling that confer protection against I/R injury. Thus we hypothesized that hypothermia will protect most using extended ischemia and early reperfusion cooling and is mediated via PKC and NO synthase (NOS). Chick cardiomyocytes were exposed to an established model of 1-h ischemia/3-h reperfusion, and the same field of initially contracting cells was monitored for viability and NO generation. Normothermic I/R resulted in 49.7 ± 3.4% cell death. Hypothermia induction to 25°C was most protective (14.3 ± 0.6% death, P < 0.001 vs. I/R control) when instituted during extended ischemia and early reperfusion, compared with induction after reperfusion (22.4 ± 2.9% death). Protection was completely lost if onset of cooling was delayed by 15 min of reperfusion (45.0 ± 8.2% death). Extended ischemia/early reperfusion cooling was associated with increased and sustained NO generation at reperfusion and decreased caspase-3 activation. The NOS inhibitor Nω-nitro-l-arginine methyl ester (200 μM) reversed these changes and abrogated hypothermia protection. In addition, the PKCε inhibitor myr-PKCε v1-2 (5 μM) also reversed NO production and hypothermia protection. In conclusion, therapeutic hypothermia initiated during extended ischemia/early reperfusion optimally protects cardiomyocytes from I/R injury. Such protection appears to be mediated by increased NO generation via activation of protein kinase Cε; nitric oxide synthase.

scholarly journals Characterization of the inositol 1,4,5-trisphosphate-induced calcium release from permeabilized endocrine cells and its inhibition by decavanadate and p-hydroxymercuribenzoate

1989 ◽  
Vol 262 (1) ◽  
pp. 83-89 ◽  
Author(s):  
K J Föhr ◽  
J Scott ◽  
G Ahnert-Hilger ◽  
M Gratzl
Keyword(s):  
Endocrine Cells ◽  
Calcium Release ◽  
Cell Types ◽  
Inhibitory Effect ◽  
Maximal Effect ◽  
Thiol Compounds ◽  
Inositol 1,4,5 Trisphosphate ◽  
Dependent Inhibition ◽  
Pheochromocytoma Pc12 ◽  
First Time

The inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ compartment of endocrine cells was studied with alpha-toxin- and digitonin-permeabilized rat insulinoma (RINA2) and rat pheochromocytoma (PC12) cells. The Ca2+ uptake was ATP-dependent, and submicromolar concentrations of IP3 specifically released the stored Ca2+. Half-maximal Ca2+ release was observed with 0.25-0.5 mumol of IP3/l, and the amount of Ca2+ released due to IP3 could be enhanced by additional loading of the Ca2+ compartment. Consecutive additions of the same concentration of IP3 for 1-2 h always released the same amount of Ca2+ without desensitization, providing an ideal basis to further characterize the IP3-induced Ca2+ release. Here we describe for the first time a reversible inhibitory effect of decavanadate on the IP3-induced Ca2+ release. Among the vanadium species tested (decavanadate, oligovanadate and monovanadate), only decavanadate was inhibitory, with a half-maximal effect at 5 mumol/l in both cell types. The effect of decavanadate could be overcome by increasing the amount of sequestered Ca2+ or added IP3. Decavanadate did not affect the ATP-driven Ca2+ uptake but oligovanadate was inhibitory on Ca2+ uptake. p-Hydroxymercuribenzoate (pHMB) at concentrations between 10 and 30 mumol/l also inhibited the Ca2+ release due to IP3. Thiol compounds such as dithiothreitol (DTT; 1 mmol/l) added before pHMB removed all its inhibitory effect on the IP3-induced Ca2+ release, whereas the inhibition caused by decavanadate was unaffected by DTT. Thus, the decavanadate-dependent inhibition functions by a distinctly different mechanism than pHMB and could serve as a specific tool to analyse various aspects of the IP3-induced Ca2+ release within endocrine cells.

scholarly journals Induction and stability of human Th17 cells require endogenous NOS2 and cGMP-dependent NO signaling

2013 ◽  
Vol 210 (7) ◽  
pp. 1433-1445 ◽  
Author(s):  
Nataša Obermajer ◽  
Jeffrey L. Wong ◽  
Robert P. Edwards ◽  
Kong Chen ◽  
Melanie Scott ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
T Cells ◽  
Cd4 T Cells ◽  
Th17 Cells ◽  
De Novo ◽  
Inflammatory Diseases ◽  
Suppressor Cells ◽  
Guanosine Monophosphate ◽  
No Production ◽  
No Signaling

Nitric oxide (NO) is a ubiquitous mediator of inflammation and immunity, involved in the pathogenesis and control of infectious diseases, autoimmunity, and cancer. We observed that the expression of nitric oxide synthase-2 (NOS2/iNOS) positively correlates with Th17 responses in patients with ovarian cancer (OvCa). Although high concentrations of exogenous NO indiscriminately suppress the proliferation and differentiation of Th1, Th2, and Th17 cells, the physiological NO concentrations produced by patients’ myeloid-derived suppressor cells (MDSCs) support the development of RORγt(Rorc)+IL-23R+IL-17+ Th17 cells. Moreover, the development of Th17 cells from naive-, memory-, or tumor-infiltrating CD4+ T cells, driven by IL-1β/IL-6/IL-23/NO-producing MDSCs or by recombinant cytokines (IL-1β/IL-6/IL-23), is associated with the induction of endogenous NOS2 and NO production, and critically depends on NOS2 activity and the canonical cyclic guanosine monophosphate (cGMP)–cGMP-dependent protein kinase (cGK) pathway of NO signaling within CD4+ T cells. Inhibition of NOS2 or cGMP–cGK signaling abolishes the de novo induction of Th17 cells and selectively suppresses IL-17 production by established Th17 cells isolated from OvCa patients. Our data indicate that, apart from its previously recognized role as an effector mediator of Th17-associated inflammation, NO is also critically required for the induction and stability of human Th17 responses, providing new targets to manipulate Th17 responses in cancer, autoimmunity, and inflammatory diseases.

scholarly journals Endogenous flow-induced nitric oxide reduces superoxide-stimulated Na/H exchange activity via PKG in thick ascending limbs

2015 ◽  
Vol 308 (5) ◽  
pp. F444-F449 ◽  
Author(s):  
Nancy J. Hong ◽  
Jeffrey L. Garvin
Keyword(s):  
Nitric Oxide ◽  
Soluble Guanylate Cyclase ◽  
Inhibitory Effect ◽  
Dependent Protein Kinase ◽  
Two Factors ◽  
Endogenous Nitric Oxide ◽  
Dependent Protein ◽  
Acid Load ◽  
Luminal Flow ◽  
Ly 83583

Luminal flow stimulates endogenous nitric oxide (NO) and superoxide (O2−) production by renal thick ascending limbs (TALs). The delicate balance between these two factors regulates Na transport in TALs; NO enhances natriuresis, whereas O2− augments Na absorption. Endogenous, flow-stimulated O2− enhances Na/H exchange (NHE). Flow-stimulated NO reduces flow-induced O2−, a process mediated by cGMP-dependent protein kinase (PKG). However, whether flow-stimulated, endogenously-produced NO diminishes O2−-stimulated NHE activity and the signaling pathway involved are unknown. We hypothesized that flow-induced NO reduces the stimulation of NHE activity caused by flow-induced O2− via PKG in TALs. Intracellular pH recovery after an acid load was measured as an indicator of NHE activity in isolated, perfused rat TALs. l-Arginine, the NO synthase substrate, decreased NHE activity by 34 ± 5% ( n = 5; P < 0.04). The O2− scavenger tempol decreased NHE activity by 46 ± 8% ( n = 6; P < 0.004) in the absence of NO. In the presence of l-arginine, the inhibitory effect of tempol on NHE activity was reduced to −19 ± 6% ( n = 6; P < 0.03). The soluble guanylate cyclase inhibitor LY-83583 blocked the effect of l-arginine thus restoring tempol's effect on NHE activity to −42 ± 4% ( n = 6; P < 0.0005). The PKG inhibitor KT-5823 also inhibited l-arginine's effect on tempol-reduced NHE activity (−43 ± 5%; n = 5; P < 0.03). We conclude that flow-induced NO reduces the stimulatory effect of endogenous, flow-induced O2− on NHE activity in TALs via an increase in cGMP and PKG activation.

scholarly journals Terbutaline, forskolin and cAMP reduce secretion of aqueous humour in the isolated bovine eye

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0244253
Author(s):  
Mohammad Shahidullah ◽  
William Stuart Wilson ◽  
Kazi Rafiq ◽  
Mahmudul Hasan Sikder ◽  
Jannatul Ferdous ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Protein Kinase A ◽  
Aqueous Humour ◽  
Calcium Release ◽  
Drug Effects ◽  
Inhibitory Effect ◽  
Plateau Phase ◽  
Intracellular Ca ◽  
Kinase A

In order to elucidate involvement of cyclic AMP and intracellular Ca2+,[Ca2+]i, in the modulation of aqueous humour formation (AHF), we studied the effects of terbutaline, forskolin and 8-Br-cAMP in the isolated bovine eye. We also studied the interaction of cAMP on calcium signaling in cultured ciliary epithelial (CE) cells. Drug effects on AHF were measured by fluorescein dilution. Drug effects on [Ca2+]i were studied by the fura-2 fluorescence ratio technique. Terbutaline (100 nmol-100 M), forskolin (30 nM-100 M) or 8-Br-cAMP (100 nM– 10 μM), administered in the arterial perfusate produced significant reductions in AHF. The AH reducing effect of terbutaline was blocked by a selective inhibitor of protein kinase A (KT-5720). ATP (100 M) caused a rapid, transient (peak) increase in [Ca2+]i followed by a sustained plateau phase lasting more than 5 minutes. Preincubation of the cells (6 min) with terbutaline, forskolin or 8-Br-cAMP significantly reduced the peak calcium response to ATP. The sustained plateau phase of the response, on the other hand, was augmented by each of the agents. KT-5720 partially reversed the inhibitory effect of terbutaline on the peak and totally inhibited its effect on the plateau phase. These data indicate: (a) that AHF in the bovine eye can be manipulated through cyclic AMP, operating via protein kinase A, (b) that protein kinase A can affect [Ca2+]i homeostasis, (c) that calcium release from the intracellular store, not the entry, affects AHF, and (d) that interaction of [Ca2+]i with cAMP plays a role in modulating AH secretion.

Abstract 17415: Mitochondrial Ca2+/Calmodulin-Dependent Kinase II Inhibition Changes the Calcium Homeostasis in the Endothelium and Decreases the Vascular Relaxation in Mesenteric Arteries

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Celio Damacena De Angelis ◽  
Daniel W Nuno ◽  
Olha Koval ◽  
Kathryn G Lamping ◽  
Isabella M Grumbach
Keyword(s):  
Calcium Release ◽  
Mesenteric Arteries ◽  
Lower Sensitivity ◽  
No Production ◽  
Vascular Relaxation ◽  
Fura 2 ◽  
The Right ◽  
Oxide Synthase ◽  
Mesenteric Resistance Artery ◽  
Endothelial Nitric Oxide

Introduction: The Ca2+/Calmodulin-dependent Kinase II (CaMKII) is present in mitochondria and cytosol. In mitochondria, it regulates the mitochondrial Ca 2+ uptake via the mitochondrial Ca2+ uniporter. Since endothelial nitric oxide synthase activity is regulated by intracellular [Ca2+], we hypothesized that it affects cytosolic Ca2+, NO production and ACh-dependent vasodilation. Hypothesis: Inhibition of mitochondrial CaMKII in endothelium increases the cytosolic [Ca2+], and decreases vasorelaxation by Acetylcholine. Methods: CaMKII in mitochondria was inhibited through expression of the mitochondria-targeted CaMKII inhibitor peptide (mito-CaMKIIN) in a novel transgenic mouse model (endo-mtCaMKIIN) in endothelial cells only or delivered by adenoviral transduction (Ad-mtCaMKIIN) in human Aortic Endothelium cells (HAEC). In HAEC, cytosolic Ca2+ levels (by FURA-2 AM), eNOS activation and NOx levels were measured. Results: The basal Ca2+ levels were higher in the cytosol of mitoCaMKIIN cells (1.08 ± 0.02 Fura-2 ratio normalized by control, p<0.05). Thapsigargin-induced ER Ca 2+ release was significantly higher with mitoCaMKIIN (AUC 0.252 ± 0.027 versus 0.112 ± 0.01275, p<0.05), whereas cytosolic Ca 2+ levels after ACh were reduced (AUC 0.191 ± 0.025 versus 0.435 ± 0.054). Higher levels of phosphorylation of eNOS at Ser1177 and Thr495 sites were seen at baseline. The concentration-response curve of vascular relaxation to acetylcholine and SNP shifted to the right (p<0.05) in mesenteric resistance artery of mitoCaMKIIN mice. Conclusions: The inhibition of mitochondrial CaMKII in the endothelium increases the cytosolic levels, endoplasmic reticulum storage of calcium and eNOS phosphorylation. However, there are lower calcium release and lower sensitivity to acetylcholine and SNP.

Sign in / Sign up

Export Citation Format

Share Document