Differential autophosphorylation of CaM kinase II from phasic and tonic smooth muscle tissues

2002 ◽  
Vol 283 (5) ◽  
pp. C1399-C1413 ◽  
Author(s):  
Jillinda M. Lorenz ◽  
Marilyn H. Riddervold ◽  
Elizabeth A. H. Beckett ◽  
Salah A. Baker ◽  
Brian A. Perrino
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscles ◽  
Proximal Colon ◽  
Gastric Fundus ◽  
Calcium Oscillations ◽  
Cam Kinase Ii ◽  
Activity Levels ◽  
Dependent Manner ◽  
Cam Kinase ◽  
Autonomous Activity

Ca+/calmodulin-dependent protein kinase II (CaM kinase II) is regulated by calcium oscillations, autophosphorylation, and its subunit composition. All four subunit isoforms were detected in gastric fundus and proximal colon smooth muscles by RT-PCR, but only the γ and δ isoforms are expressed in myocytes. Relative γ and δ message levels were quantitated by real-time PCR. CaM kinase II protein and Ca2+/calmodulin-stimulated (total) activity levels are higher in proximal colon smooth muscle lysates than in fundus lysates, but Ca2+/calmodulin-independent (autonomous) activity is higher in fundus lysates. CaM kinase II in fundus lysates is relatively unresponsive to Ca2+/calmodulin. Alkaline phosphatase decreased CaM kinase II autonomous activity in fundus lysates and restored its responsiveness to Ca2+/calmodulin. Acetylcholine (ACh) increased autonomous CaM kinase II activity in fundus and proximal colon smooth muscles in a time- and dose-dependent manner. KN-93 enhanced ACh-induced fundus contractions but inhibited proximal colon contractions. The different properties of CaM kinase II from fundus and proximal colon smooth muscles suggest differential regulation of its autophosphorylation and activity in tonic and phasic gastrointestinal smooth muscles.

Roles of CaM kinase II and phospholamban in SNP-induced relaxation of murine gastric fundus smooth muscles

2006 ◽  
Vol 291 (2) ◽  
pp. C337-C347 ◽  
Author(s):  
Minkyung Kim ◽  
In Soo Han ◽  
Sang Don Koh ◽  
Brian A. Perrino
Keyword(s):  
Smooth Muscle ◽  
Soluble Guanylate Cyclase ◽  
Muscle Relaxation ◽  
Smooth Muscles ◽  
Cyclopiazonic Acid ◽  
Gastric Fundus ◽  
Smooth Muscle Relaxation ◽  
Cam Kinase Ii ◽  
Cam Kinase ◽  
No Donor

The mechanisms by which nitric oxide (NO) relaxes smooth muscles are unclear. The NO donor sodium nitroprusside (SNP) has been reported to increase the Ca2+ release frequency (Ca2+ sparks) through ryanodine receptors (RyRs) and activate spontaneous transient outward currents (STOCs), resulting in smooth muscle relaxation. Our findings that caffeine relaxes and hyperpolarizes murine gastric fundus smooth muscles and increases phospholamban (PLB) phosphorylation by Ca2+/calmodulin (CaM)-dependent protein kinase II (CaM kinase II) suggest that PLB phosphorylation by CaM kinase II participates in smooth muscle relaxation by increasing sarcoplasmic reticulum (SR) Ca2+ uptake and the frequencies of SR Ca2+ release events and STOCs. Thus, in the present study, we investigated the roles of CaM kinase II and PLB in SNP-induced relaxation of murine gastric fundus smooth muscles. SNP hyperpolarized and relaxed gastric fundus circular smooth muscles and activated CaM kinase II. SNP-induced CaM kinase II activation was prevented by KN-93. Ryanodine, tetracaine, 2-aminoethoxydiphenylborate, and cyclopiazonic acid inhibited SNP-induced fundus smooth muscle relaxation and CaM kinase II activation. The Ca2+-activated K+ channel blockers iberiotoxin and apamin inhibited SNP-induced hyperpolarization and relaxation. The soluble guanylate cyclase inhibitor 1 H-[1,2,4]oxadiazolo-[4,3-α]quinoxalin-1-one inhibited SNP-induced relaxation and CaM kinase II activation. The membrane-permeable cGMP analog 8-bromo-cGMP relaxed gastric fundus smooth muscles and activated CaM kinase II. SNP increased phosphorylation of PLB at Ser16 and Thr17. Thr17 phosphorylation of PLB was inhibited by cyclopiazonic acid and KN-93. Ser16 and Thr17 phosphorylation of PLB was sensitive to 1 H-[1,2,4]oxadiazolo-[4,3-α]quinoxalin-1-one. These results demonstrate a novel pathway linking the NO-soluble guanylyl cyclase-cGMP pathway, SR Ca2+ release, PLB, and CaM kinase II to relaxation in gastric fundus smooth muscles.

CaM kinase II and phospholamban contribute to caffeine-induced relaxation of murine gastric fundus smooth muscle

2005 ◽  
Vol 288 (6) ◽  
pp. C1202-C1210 ◽  
Author(s):  
Minkyung Kim ◽  
Sang Yun Cho ◽  
In Soo Han ◽  
Sang Don Koh ◽  
Brian A. Perrino
Keyword(s):  
Smooth Muscle ◽  
Sarcoplasmic Reticulum ◽  
Channel Blocker ◽  
Muscle Tone ◽  
Smooth Muscles ◽  
Cyclopiazonic Acid ◽  
Gastric Fundus ◽  
Cam Kinase Ii ◽  
Cam Kinase ◽  
Camkii Activation

Caffeine has been shown to increase the Ca2+ release frequency (Ca2+ sparks) from the sarcoplasmic reticulum (SR) through ryanodine-sensitive stores and relax gastric fundus smooth muscle. Increased Ca2+ store refilling increases the frequency of Ca2+ release events and store refilling is enhanced by CaM kinase II (CaMKII) phosphorylation of phospholamban (PLB). These findings suggest that transient, localized Ca2+ release events from the SR may activate CaMKII and contribute to relaxation by enhancing store refilling due to PLB Thr17 phosphorylation. To investigate this possibility, we examined the effects of caffeine on CaMKII, muscle tone, and PLB phosphorylation in murine gastric fundus smooth muscle. Caffeine (1 mM) hyperpolarized and relaxed murine gastric fundus smooth muscle and activated CaMKII. Ryanodine, tetracaine, or cyclopiazonic acid each prevented CaMKII activation and significantly inhibited caffeine-induced relaxation. The large-conductance Ca2+-activated K+ channel blocker iberiotoxin, but not apamin, partially inhibited caffeine-induced relaxation. Caffeine-induced CaMKII activation increased PLB Thr17, but not PLB Ser16 phosphorylation. 3-Isobutyl-1-methylxanthine increased PLB Ser16 phosphorylation, but not PLB Thr17 phosphorylation. The CaMKII inhibitor KN-93 inhibited caffeine-induced relaxation and PLB Thr17 phosphorylation. These results show that caffeine-induced CaMKII activation and PLB phosphorylation play a role in the relaxation of gastric fundus smooth muscles.

Phospholamban knockout increases CaM kinase II activity and intracellular Ca2+ wave activity and alters contractile responses of murine gastric antrum

2008 ◽  
Vol 294 (2) ◽  
pp. C432-C441 ◽  
Author(s):  
Minkyung Kim ◽  
Grant W. Hennig ◽  
Terence K. Smith ◽  
Brian A. Perrino
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscles ◽  
Cyclopiazonic Acid ◽  
Gastric Fundus ◽  
Gastric Antrum ◽  
Cam Kinase Ii ◽  
Cam Kinase ◽  
Smooth Muscle Contractility ◽  
Intracellular Ca ◽  
Dependent Protein

Phospholamban (PLB) inhibits the sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA), and this inhibition is relieved by Ca2+ calmodulin-dependent protein kinase II (CaM kinase II) phosphorylation. We previously reported significant differences in contractility, SR Ca2+ release, and CaM kinase II activity in gastric fundus smooth muscles as a result of PLB phosphorylation by CaM kinase II. In this study, we used PLB-knockout (PLB-KO) mice to directly examine the effect of PLB absence on contractility, CaM kinase II activity, and intracellular Ca2+ waves in gastric antrum smooth muscles. The frequencies and amplitudes of spontaneous phasic contractions were elevated in antrum smooth muscle strips from PLB-KO mice. Bethanecol increased the amplitudes of phasic contractions in antrum smooth muscles from both control and PLB-KO mice. Caffeine decreased and cyclopiazonic acid (CPA) increased the basal tone of antrum smooth muscle strips from PLB-KO mice, but the effects were less pronounced compared with control strips. The CaM kinase II inhibitor KN-93 was less effective at inhibiting caffeine-induced relaxation in antrum smooth muscle strips from PLB-KO mice. CaM kinase II autonomous activity was elevated, and not further increased by caffeine, in antrum smooth muscles from PLB-KO mice. Similarly, the intracellular Ca2+ wave frequency was elevated, and not further increased by caffeine, in antrum smooth muscles from PLB-KO mice. These findings suggest that PLB is an important modulator of gastric antrum smooth muscle contractility by modulation of SR Ca2+ release and CaM kinase II activity.

CaM kinase II activation and phospholamban phosphorylation by SNP in murine gastric antrum smooth muscles

2007 ◽  
Vol 292 (4) ◽  
pp. G1045-G1054 ◽  
Author(s):  
Minkyung Kim ◽  
Brian A. Perrino
Keyword(s):  
Protein Kinase ◽  
Muscle Relaxation ◽  
Smooth Muscles ◽  
Cyclopiazonic Acid ◽  
Gastric Fundus ◽  
Gastric Antrum ◽  
Smooth Muscle Relaxation ◽  
Cam Kinase Ii ◽  
Cam Kinase ◽  
Intracellular Ca

Elevations in the intracellular Ca2+ concentration activate the serine/threonine protein kinase Ca2+/calmodulin-dependent protein kinase II (CaM kinase II). We tested the hypothesis that increased sarco(endo)plasmic reticulum Ca2+-ATPase activity by phospholamban (PLB) phosphorylation contributes to smooth muscle relaxation by elevating the sarcoplasmic reticulum (SR) Ca2+ load and increasing the frequency of Ca2+ release events from the SR. We have previously shown that caffeine or sodium nitroprusside (SNP) relaxes murine gastric fundus smooth muscles and increases PLB phosphorylation by CaM kinase II. These findings suggest that an increased SR Ca2+ load increases the frequency of Ca2+ transients from the SR and results in PLB phosphorylation by CaM kinase II, contributing to caffeine- or SNP-induced relaxation. The aim of the present study was to investigate the effects of SNP on CaM kinase II and PLB phosphorylation in gastric antrum smooth muscles. SNP or 8-bromo-cGMP decreased the basal tone and amplitudes of spontaneous phasic contractions and activated CaM kinase II. SNP-induced relaxation and CaM kinase II activation were blocked by [1,2,4]oxadizolo-[4,3α]quinoxaline-1-one (ODQ) and inhibited by cyclopiazonic acid (CPA) or KN-93. SNP also increased PLBSer16 and PLBThr17 phosphorylation. Both PLBSer16 and Thr17 phosphorylation were ODQ sensitive. However, only PLBThr17 phosphorylation was inhibited by CPA or KN-93. These results suggest that CaM kinase II activation and PLB phosphorylation participate in the relaxant effect of SNP on murine gastric antrum smooth muscles through a nitric oxide/guanylyl cyclase/cGMP pathway.

Endothelium-dependent hyperpolarization elicited by adenine compounds in rabbit carotid artery

1991 ◽  
Vol 260 (4) ◽  
pp. H1037-H1042 ◽  
Author(s):  
G. Chen ◽  
H. Suzuki
Keyword(s):  
Smooth Muscle ◽  
Carotid Artery ◽  
Direct Action ◽  
Smooth Muscles ◽  
Muscle Membrane ◽  
Dependent Manner ◽  
Rabbit Carotid Artery ◽  
Mechanical Removal ◽  
Smooth Muscle Membrane ◽  
Adventitial Cells

Electrical responses of the membrane of intimal and adventitial smooth muscle cells of the rabbit carotid artery to ATP, ADP, AMP, and adenosine were recorded. In intimal cells, these compounds hyperpolarized the membrane. Mechanical removal of the endothelium altered the responses to ATP and ADP to one of a transient depolarization, with no alteration of the response to AMP and adenosine. In the adventitial cells, ATP and ADP produced a transient depolarization, whereas AMP and adenosine produced a sustained hyperpolarization, irrespective of the presence or absence of the endothelium. In tissues with an intact endothelium, 5'-adenylylimidodiphosphate tetralithium salt and alpha,beta-methylene ATP (mATP) transiently depolarized the membrane in these smooth muscles. In case of stabilization with mATP, only hyperpolarization was generated by ATP, in an endothelium-dependent manner. Our interpretation of these observations is that 1) ATP and ADP depolarize smooth muscle membrane by a direct action and hyperpolarize the membrane indirectly through the release of endothelium-derived hyperpolarizing factor, 2) AMP and adenosine hyperpolarize the membrane, independently of the endothelium, and 3) ATP receptors present on the endothelial cell membrane differ from those on smooth muscle membrane.

scholarly journals Molecular Mechanisms of Large-Conductance Ca2+-Activated Potassium Channel Activation by Ginseng Gintonin

2013 ◽  
Vol 2013 ◽  
pp. 1-14 ◽  
Author(s):  
S. H. Choi ◽  
B. H. Lee ◽  
S. H. Hwang ◽  
H. J. Kim ◽  
S. M. Lee ◽  
...  
Keyword(s):  
Blood Vessel ◽  
Calcium Binding ◽  
Molecular Mechanisms ◽  
Cam Kinase Ii ◽  
Phosphorylation Sites ◽  
Dependent Manner ◽  
Cam Kinase ◽  
Concentration Dependent Manner ◽  
Channel Activation ◽  
Lpa Receptor

Gintonin is a unique lysophosphatidic acid (LPA) receptor ligand found inPanax ginseng. Gintonin induces transient [Ca2+]ithrough G protein-coupled LPA receptors. Large-conductance Ca2+-activated K+(BKCa) channels are expressed in blood vessels and neurons and play important roles in blood vessel relaxation and attenuation of neuronal excitability. BKCachannels are activated by transient [Ca2+]iand are regulated by various Ca2+-dependent kinases. We investigated the molecular mechanisms of BKCachannel activation by gintonin. BKCachannels are heterologously expressed inXenopus oocytes. Gintonin treatment induced BKCachannel activation in oocytes expressing the BKCachannelαsubunit in a concentration-dependent manner (EC50= 0.71 ± 0.08 µg/mL). Gintonin-mediated BKCachannel activation was blocked by a PKC inhibitor, calphostin, and by the calmodulin inhibitor, calmidazolium. Site-directed mutations in BKCachannels targeting CaM kinase II or PKC phosphorylation sites but not PKA phosphorylation sites attenuated gintonin action. Mutations in the Ca2+bowl and the regulator of K+conductance (RCK) site also blocked gintonin action. These results indicate that gintonin-mediated BKCachannel activations are achieved through LPA1 receptor-phospholipase C-IP3-Ca2+-PKC-calmodulin-CaM kinase II pathways and calcium binding to the Ca2+bowl and RCK domain. Gintonin could be a novel contributor against blood vessel constriction and over-excitation of neurons.

Electromechanical effects of endothelin on ferret bronchial and tracheal smooth muscle

1990 ◽  
Vol 68 (1) ◽  
pp. 417-420 ◽  
Author(s):  
H. K. Lee ◽  
G. D. Leikauf ◽  
N. Sperelakis
Keyword(s):  
Smooth Muscle ◽  
Slow Wave ◽  
Smooth Muscles ◽  
Muscle Cells ◽  
Wave Activity ◽  
Slow Wave Activity ◽  
Resting Potential ◽  
Base Line ◽  
Dependent Manner ◽  
Bronchial Cells

The effects of endothelin (ET) on transmembrane potential and isometric force were studied in ferret bronchial and tracheal smooth muscles. At rest, the muscle cells were electrically and mechanically quiescent. The mean resting potential for the bronchial cells was -70 +/- 1 mV (n = 25 cells/8 ferrets), and that of the tracheal cells was -60 +/- 1 mV (n = 7 cells/2 ferrets). ET depolarized and contracted both types of muscle cells in a concentration-dependent manner. At 1 nM ET, the bronchial muscle cells were significantly depolarized with concomitant force generation. In contrast, greater than 30 nM ET was required for the tracheal muscle cells to respond. The bronchial cells were further depolarized by 10 and 100 nM ET with electrical slow-wave activity present. The calcium channel antagonist verapamil substantially inhibited the contractions produced by 100 nM ET and abolished the slow-wave activity without affecting the base-line depolarization. Pretreatment of the bronchial muscle with 30 microM indomethacin did not affect the ET-induced contraction. These results suggest that ET modulates airway smooth muscle tone by direct activation and/or depolarization-induced activation of sarcolemmal calcium channels.

scholarly journals Effects of Ursolic Acid on Contractile Activity of Gastric Smooth Muscles

2015 ◽  
Vol 10 (4) ◽  
pp. 1934578X1501000 ◽  
Author(s):  
Natalia Prissadova ◽  
Petko Bozov ◽  
Kiril Marinkov ◽  
Hristo Badakov ◽  
Atanas Kristev
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Ursolic Acid ◽  
Slow Wave ◽  
Contractile Activity ◽  
Smooth Muscles ◽  
Dependent Manner ◽  
Krebs Solution ◽  
Concentration Dependent Manner ◽  
Gastric Smooth Muscle

Ursolic acid (UA) in concentrations of 1×10−7 mol/L - 5×10−5 mol/L induced relaxation in gastric smooth muscle (SM) tissues, in a concentration-dependent manner. The relaxation did not change membrane potential and slow wave contraction patterns. A significant decrease in amplitude and frequency of spike-potentials was observed. UA-induced reactivity was removed when SM preparations were treated with nifedipine (1×10−6 mol/L). Ca2+- induced contractions of the depolarized SM preparations (42 mmol/L K+; Ca2+- free Krebs solution) were substantially reduced in the presence of UA. It was determined that, in certain concentrations, UA influenced L – type Ca2+ channels, and reduced the Ca2+ influx.

scholarly journals Inhibition of smooth-muscle myosin-light-chain phosphatase by Ruthenium Red

2000 ◽  
Vol 349 (3) ◽  
pp. 797-804 ◽  
Author(s):  
Aki YAMADA ◽  
Osamu SATO ◽  
Minoru WATANABE ◽  
Michael P. WALSH ◽  
Yasuo OGAWA ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Smooth Muscles ◽  
Direct Interaction ◽  
Smooth Muscle Contraction ◽  
Ruthenium Red ◽  
Dependent Manner ◽  
Myosin Light Chain Phosphatase ◽  
Concentration Dependent Manner

Ruthenium Red (RuR) is widely used as an inhibitor of ryanodine receptor Ca2+ release channels, but has additional effects such as the induction of Ca2+ sensitization of contraction of permeabilized smooth muscles. To address the mechanism underlying this process, we examined the effects of RuR on contractility in permeabilized guinea-pig ileum and on the activity of myosin-light-chain phosphatase (MP). RuR increased the force at submaximal [Ca2+] (pCa 6.3) approx. 4-fold. This effect was not observed after thiophosphorylation of MP. RuR also seemed capable of preventing the thiophosphorylation of MP, suggesting a direct interaction of RuR with MP. Consistent with this possibility, smooth-muscle MP was inhibited by RuR in a concentration-dependent manner (IC50 23µM). Exogenous calmodulin significantly increased RuR-induced contraction at pCa 6.3 but had little effect on contraction induced by microcystin at this [Ca2+]. Ca2+-independent contraction was induced by RuR (EC50 843µM) and by microcystin (EC50 59nM) but the maximal force induced by RuR was smaller than that induced by microcystin. The addition of 300µM RuR enhanced the contraction induced by 30nM microcystin but markedly decreased that induced by 1µM microcystin. Such a dual action of RuR on microcystin-induced effects was not observed in experiments using purified MP. We conclude that the RuR-induced Ca2+ sensitization of smooth-muscle contraction is due to the direct inhibition of MP by RuR.

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