Role of cGMP-dependent protein kinase in development of tolerance to nitric oxide in pulmonary veins of newborn lambs

2004 ◽  
Vol 286 (4) ◽  
pp. L786-L792 ◽  
Author(s):  
Yuansheng Gao ◽  
Srinivas Dhanakoti ◽  
Earleen M. Trevino ◽  
Xiaohua Wang ◽  
Fred C. Sander ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Pulmonary Veins ◽  
Nitric Oxide Donor ◽  
Continuous Exposure ◽  
Mrna Levels ◽  
Dependent Protein Kinase ◽  
Dependent Protein ◽  
A Cell ◽  
Development Of Tolerance

Continuous exposure to nitrovasodilators and nitric oxide induces tolerance to their vasodilator effects in vascular smooth muscle. This study was done to determine the role of cGMP-dependent protein kinase (PKG) in the development of tolerance to nitric oxide. Isolated fourth-generation pulmonary veins of newborn lambs were studied. Incubation of veins for 20 h with DETA NONOate (DETA NO; a stable nitric oxide donor) significantly reduced their relaxation response to the nitric oxide donor and to β-phenyl-1, N2-etheno-8-bromo-cGMP (8-Br-PET-cGMP, a cell-permeable cGMP analog). Incubation with DETA NO significantly reduced PKG activity and protein and mRNA levels in the vessels. These effects were prevented by 1H-(1,2,4)oxadiazolo(4,3- a)quinoxalin-1-one (an inhibitor of soluble guanylyl cyclase) and Rp-8-Br-PET-cGMPS (an inhibitor of PKG). A decrease in PKG protein and mRNA levels was also observed after continuous exposure to cGMP analogs. The PKG inhibitor abrogated these effects. The decrease in cGMP-mediated relaxation and in PKG activity caused by continuous exposure to DETA NO was not affected by KT-5720, an inhibitor of cAMP-dependent protein kinase. Prolonged exposure to 8-Br-cAMP (a cell-permeable cAMP analog) did not affect PKG protein level in the veins. These results suggest that continuous exposure to nitric oxide or cGMP downregulates PKG by a PKG-dependent mechanism. Such a negative feedback mechanism may contribute to the development of tolerance to nitric oxide in pulmonary veins of newborn lambs.

Guanylyl cyclase stimulatory coupling to KCachannels

2000 ◽  
Vol 279 (6) ◽  
pp. C1938-C1945 ◽  
Author(s):  
M. Nara ◽  
P. D. K. Dhulipala ◽  
G. J. Ji ◽  
U. R. Kamasani ◽  
Y.-X. Wang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Guanylyl Cyclase ◽  
Phosphorylation Site ◽  
Nitric Oxide Donor ◽  
Dependent Manner ◽  
Dependent Protein Kinase ◽  
Α Subunit ◽  
Dependent Protein ◽  
Spermine Nonoate

We coexpressed the human large-conductance, calcium-activated K (KCa) channel (α- and β-subunits) and rat atrial natriuretic peptide (ANP) receptor genes in Xenopus oocytes to examine the mechanism of guanylyl cyclase stimulatory coupling to the channel. Exposure of oocytes to ANP stimulated whole cell KCa currents by 21 ± 3% (at 60 mV), without altering current kinetics. Similarly, spermine NONOate, a nitric oxide donor, increased KCa currents (20 ± 4% at 60 mV) in oocytes expressing the channel subunits alone. Stimulation of KCacurrents by ANP was inhibited in a concentration-dependent manner by a peptide inhibitor of cGMP-dependent protein kinase (PKG). Receptor/channel stimulatory coupling was not completely abolished by mutating the cAMP-dependent protein kinase phosphorylation site on the α-subunit (S869; Nars M, Dhulipals PD, Wang YX, and Kotlikoff MI. J Biol Chem 273: 14920–14924, 1998) or by mutating a neighboring consensus PKG site (S855), but mutation of both residues virtually abolished coupling. Spermine NONOate also failed to stimulate channels expressed from the double mutant cRNAs. These data indicate that nitric oxide donors stimulate KCa channels through cGMP-dependent phosphorylation and that two serine residues (855 and 869) underlie this stimulatory coupling.

Nitric oxide donor SIN-1 inhibits mammalian cardiac calcium current through cGMP-dependent protein kinase

1995 ◽  
Vol 268 (1) ◽  
pp. C45-C54 ◽  
Author(s):  
G. M. Wahler ◽  
S. J. Dollinger
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Calcium Current ◽  
Inhibitory Effect ◽  
Cardiac Cells ◽  
Nitric Oxide Donor ◽  
Dependent Protein Kinase ◽  
Cyclic Monophosphate ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

The effect of the nitric oxide (NO) donor SIN-1 (3-morpholino-sydnonimine) on the calcium current (ICa) was examined in guinea pig ventricular myocytes. SIN-1 had little effect on basal ICa. After moderate stimulation of ICa with 10 nM isoproterenol (ISO), 10 microM SIN-1 caused either stimulation or inhibition of ICa; 100 microM SIN-1 consistently caused inhibition. SIN-1 (1-100 microM) inhibited ICa equally following considerable enhancement of ICa by either 1 microM ISO or 100 microM 3-isobutyl-1-methylxanthine, a nonspecific phosphodiesterase (PDE) inhibitor. SIN-1 (100 microM) also inhibited ICa equally following enhancement by either 10 microM pipette adenosine 3',5'-cyclic monophosphate (cAMP) or hydrolysis-resistant 8-bromo-cAMP. Thus the inhibitory effect of SIN-1 appears independent of PDEs. Addition of LY-83583 (a blocker of guanylate cyclase) to the pipette or superfusion with KT-5823 [a blocker of the guanosine 3',5'-cyclic monophosphate (cGMP)-dependent protein kinase] suppressed the inhibitory effect of SIN-1. We conclude that NO is an important modulator of beta-adrenergic effects on ICa and that the mechanism of NO inhibition of ICa in mammalian cardiac cells involves the cGMP-dependent protein kinase.

scholarly journals Nitrosyl-cobinamide (NO-Cbi), a new nitric oxide donor, improves wound healing through cGMP/cGMP-dependent protein kinase

2013 ◽  
Vol 25 (12) ◽  
pp. 2374-2382 ◽  
Author(s):  
Ryan Spitler ◽  
Raphaela Schwappacher ◽  
Tao Wu ◽  
Xiangduo Kong ◽  
Kyoko Yokomori ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Wound Healing ◽  
Protein Kinase ◽  
Nitric Oxide Donor ◽  
Dependent Protein Kinase ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

scholarly journals Anti-echinococcal effect of verapamil involving the regulation of the calcium/calmodulin-dependent protein kinase II response in vitro and in a murine infection model

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Hai-Jun Gao ◽  
Xu-Dong Sun ◽  
Yan-Ping Luo ◽  
Hua-Sheng Pang ◽  
Xing-Ming Ma ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Mrna Expression ◽  
Echinococcus Granulosus ◽  
Calcium Content ◽  
Mrna Levels ◽  
Dependent Protein Kinase ◽  
Calcium Calmodulin Dependent ◽  
Dependent Protein

Abstract Background Echinococcosis, which is caused by the larvae of cestodes of the genus Echinococcus, is a parasitic zoonosis that poses a serious threat to the health of humans and animals globally. Albendazole is the drug of choice for the treatment of echinococcosis, but it is difficult to meet clinical goals with this chemotherapy due to its low cure rate and associated side effects after its long-term use. Hence, novel anti-parasitic targets and effective treatment alternatives are urgently needed. A previous study showed that verapamil (Vepm) can suppress the growth of Echinococcus granulosus larvae; however, the mechanism of this effect remains unclear. The aim of the present study was to gain insight into the anti-echinococcal effect of Vepm on Echinococcus with a particular focus on the regulatory effect of Vepm on calcium/calmodulin-dependent protein kinase II (Ca2+/CaM-CaMKII) in infected mice. Methods The anti-echinococcal effects of Vepm on Echinococcus granulosus protoscoleces (PSC) in vitro and Echinococcus multilocularis metacestodes in infected mice were assessed. The morphological alterations in Echinococcus spp. induced by Vepm were observed by scanning electron microscopy (SEM), and the changes in calcium content in both the parasite and mouse serum and liver were measured by SEM-energy dispersive spectrometry, inductively coupled plasma mass spectrometry and alizarin red staining. Additionally, the changes in the protein and mRNA levels of CaM and CaMKII in infected mice, and in the mRNA levels of CaMKII in E. granulosus PSC, were evaluated after treatment with Vepm by immunohistochemistry and/or real-time quantitative polymerase chain reaction. Results In vitro, E. granulosus PSC could be killed by Vepm at a concentration of 0.5 μg/ml or higher within 8 days. Under these conditions, the ultrastructure of PSC was damaged, and this damage was accompanied by obvious calcium loss and downregulation of CaMKII mRNA expression. In vivo, the weight and the calcium content of E. multilocularis metacestodes from mice were reduced after treatment with 40 mg/kg Vepm, and an elevation of the calcium content in the sera and livers of infected mice was observed. In addition, downregulation of CaM and CaMKII protein and mRNA expression in the livers of mice infected with E. multilocularis metacestodes was found after treatment with Vepm. Conclusions Vepm exerted a parasiticidal effect against Echinococcus both in vitro and in vivo through downregulating the expression of Ca2+/CaM-CaMKII, which was over-activated by parasitic infection. The results suggest that Ca2+/CaM-CaMKII may be a novel drug target, and that Vepm is a potential anti-echinococcal drug for the future control of echinococcosis.

Nitric oxide and cGMP protein kinase activity in aged ventricular myocytes

2001 ◽  
Vol 281 (6) ◽  
pp. H2304-H2309 ◽  
Author(s):  
Qihang Zhang ◽  
Bruno Molino ◽  
Lin Yan ◽  
Todd Haim ◽  
Yakir Vaks ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Kinase Inhibitor ◽  
Ventricular Myocytes ◽  
Nitric Oxide Donor ◽  
Protein Kinase Activity ◽  
Edge Detector ◽  
Negative Effects ◽  
Dependent Protein

We tested the hypothesis that nitric oxide-induced negative functional effects through cGMP would be reduced in aged cardiac myocytes. Maximum rate of shortening ( R max) and percent shortening of ventricular myocytes from young (6 mo) and old (3 y) rabbits were studied using a video edge detector. cGMP-dependent phosphorylation was examined by electrophoresis and autoradiography. Myocytes received a nitric oxide donor S-nitroso- N-acetyl-penicillamine (SNAP, 10−7, 10−6, and 10−5 M) followed by KT-5823 (10−6 M), a cGMP protein kinase inhibitor. Baseline function was similar in young and old myocytes (89.1 ± 4.5 young vs. 86.4 ± 8.3 μm/s old R max, 5.6 ± 0.3 vs. 5.2 ± 0.7%shortening). SNAP (10−5 M) decreased R max in both young (25%, n = 6) and old myocytes (24%, n = 7). SNAP also reduced percent shortening by 28% in young and 23% in old myocytes. The negative effects of SNAP were partially reversed by KT-5823 only in young myocytes. Multiple proteins were phosphorylated by cGMP, and KT-5823 could reduce this effect. The degree of phosphorylation was significantly less in old myocytes. These results suggest that the functional response of ventricular myocytes to nitric oxide was preserved during aging. However, the importance of cGMP-dependent protein phosphorylation was decreased, indicating a shift to other pathways.

scholarly journals Coordination between Calcium/Calmodulin-Dependent Protein Kinase II and Neuronal Nitric Oxide Synthase in Neurons

2020 ◽  
Vol 21 (21) ◽  
pp. 7997
Author(s):  
Shoma Araki ◽  
Koji Osuka ◽  
Tsuyoshi Takata ◽  
Yukihiro Tsuchiya ◽  
Yasuo Watanabe
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Cell Injury ◽  
Dependent Protein Kinase ◽  
Calcium Calmodulin Dependent ◽  
Pathophysiological Role ◽  
Protein Kinase Ii ◽  
Mutual Regulation ◽  
Dependent Protein ◽  
Oxide Synthase

Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) is highly abundant in the brain and exhibits broad substrate specificity, thereby it is thought to participate in the regulation of neuronal death and survival. Nitric oxide (NO), produced by neuronal NO synthase (nNOS), is an important neurotransmitter and plays a role in neuronal activity including learning and memory processes. However, high levels of NO can contribute to excitotoxicity following a stroke and neurodegenerative disease. Aside from NO, nNOS also generates superoxide which is involved in both cell injury and signaling. CaMKII is known to activate and translocate from the cytoplasm to the post-synaptic density in response to neuronal activation where nNOS is predominantly located. Phosphorylation of nNOS at Ser847 by CaMKII decreases NO generation and increases superoxide generation. Conversely, NO-induced S-nitrosylation of CaMKII at Cys6 is a prominent determinant of the CaMKII inhibition in ATP competitive fashion. Thus, the “cross-talk” between CaMKII and NO/superoxide may represent important signal transduction pathways in brain. In this review, we introduce the molecular mechanism of and pathophysiological role of mutual regulation between CaMKII and nNOS in neurons.

Ca2+/calmodulin-dependent protein kinase II activation and regulation of adrenal glomerulosa Ca2+ signaling

1995 ◽  
Vol 269 (6) ◽  
pp. F751-F760 ◽  
Author(s):  
R. J. Fern ◽  
M. S. Hahm ◽  
H. K. Lu ◽  
L. P. Liu ◽  
F. S. Gorelick ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Kinase Activity ◽  
Low Voltage ◽  
Extracellular Potassium ◽  
Dependent Protein Kinase ◽  
Calmodulin Antagonist ◽  
Cell Extracts ◽  
Protein Kinase Ii ◽  
Dependent Protein ◽  
A Cell

We recently reported that elevations in the intracellular Ca2+ concentration ([Ca2+]i) enhance low-voltage-activated, T-type, Ca2+ channel activity via Ca2+/calmodulin-dependent protein kinase II (CaMKII). Here, we document CaMKII activity in bovine adrenal glomerulosa (AG) cells and assess the importance of CaMKII in depolarization-induced Ca2+ signaling. AG cell extracts exhibited kinase activity toward a CaMKII-selective peptide substrate that was dependent on both Ca2+ [half-maximal concentration for Ca2+ activation (K0.5) = 1.5 microM] and calmodulin (K0.5 = 46 nM) and was sensitive to a calmodulin antagonist and a CaMKII peptide inhibitor. On cell treatment with elevated extracellular potassium (10-60 mM) or angiotensin II, Ca(2+)-independent CaMKII activity increased to 133-205% of basal activity. Ca(2+)-independent kinase activity in agonist-stimulated extracts was inhibited by the CaMKII inhibitor peptide, 1(-)[N,O-bis(1,5- isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN-62), a cell-permeable inhibitor of CaMKII, reduced the agonist-induced stimulation of Ca(2+)-independent CaMKII activity. KN-62 also diminished depolarization-induced increases in [Ca2+]i without affecting the membrane potential. These observations suggest that CaMKII is activated in situ by aldosterone secretagogues and augments Ca2+ signaling through voltage-gated Ca2+ channels.

Effect of oxygen on cyclic GMP-dependent protein kinase-mediated relaxation in ovine fetal pulmonary arteries and veins

2003 ◽  
Vol 285 (3) ◽  
pp. L611-L618 ◽  
Author(s):  
Yuansheng Gao ◽  
Srinivas Dhanakoti ◽  
Earleen M. Trevino ◽  
Fred C. Sander ◽  
Ada M. Portugal ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Cyclic Gmp ◽  
Pulmonary Arteries ◽  
Perinatal Period ◽  
Mrna Levels ◽  
Dependent Protein Kinase ◽  
Dependent Protein ◽  
Near Term ◽  
Effect Of Oxygen ◽  
Arteries And Veins

Cyclic GMP-dependent protein kinase (PKG) plays an important role in regulating pulmonary vasomotor tone in the perinatal period. In this study, we tested the hypothesis that a change in oxygen tension affects PKG-mediated pulmonary vasodilation. Isolated intrapulmonary arteries and veins of near-term fetal lambs were first incubated for 4 h under hypoxic and normoxic conditions (Po2 of 30 and 140 mmHg, respectively) and then contracted with endothelin-1. 8-Bromoguanosine 3′,5′-cyclic monophosphate (8-BrcGMP), a cell membrane-permeable analog of cGMP, induced a greater relaxation in vessels incubated in normoxia than in hypoxia. β-Phenyl-1, N2-etheno-8-bromoguanosine-3′,5′-cyclic monophosphorothioate, Rp isomer ( Rp-8-Br-PET-cGMPS), a selective inhibitor of PKG, attenuated relaxation induced by 8-BrcGMP (10-4 and 3 × 10-4 M). In the presence of Rp-8-Br-PET-cGMPS, the differential responses to 8-BrcGMP between hypoxia and normoxia treatment were abolished in veins but not in arteries. cGMP-stimulated PKG activity was present in arteries but not in veins after 4 h of hypoxia. Both vessel types showed significant increase in cGMP-stimulated PKG activity after 4 h of normoxia. PKG protein (Western blot analysis) and PKG mRNA levels (quantitative RT-PCR) were greater in veins but not in arteries after 4-h exposure to normoxia vs. hypoxia. These results demonstrate that oxygen augments cGMP-mediated vasodilation of fetal pulmonary arteries and veins. Furthermore, the effect of oxygen on response of the veins to cGMP is due to an increase in the activity, protein level, and mRNA of PKG.

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