Increases in endothelial Ca2+ activate KCa channels and elicit EDHF-type arteriolar dilation via gap junctions

In skeletal muscle arterioles, the pathway leading to non-nitric oxide (NO), non-prostaglandin-mediated endothelium-derived hyperpolarizing factor (EDHF)-type dilations is not well characterized. To elucidate some of the steps in this process, simultaneous changes in endothelial intracellular Ca2+ concentration ([Ca2+]i) and the diameter of rat gracilis muscle arterioles (∼60 μm) to acetylcholine (ACh) were measured by fura 2 microfluorimetry (in the absence of NO and prostaglandins). ACh elicited rapid increases in endothelial [Ca2+]i (101 ± 7%), followed by substantial dilations (73 ± 2%, coupling time: 1.3 ± 0.2 s) that were prevented by endothelial loading of an intracellular Ca2+ chelator [1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′-tetraacetic acid]. Arteriolar dilations to ACh were also inhibited by intraluminal administration of the Ca2+-activated K+ (KCa) channel blockers charybdotoxin plus apamin or by palmitoleic acid, an uncoupler of myoendothelial gap junctions without affecting changes in endothelial [Ca2+]i. The presence of large conductance KCa channels on arteriolar endothelial cells was demonstrated with immunohistochemisty. We propose that in skeletal muscle arterioles, EDHF-type mediation is evoked by an increase in endothelial [Ca2+]i, which by activating endothelial KCa channels elicits hyperpolarization that is conducted via myoendothelial gap junctions to the smooth muscle resulting in decreases in [Ca2+]i and consequently dilation.

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Release of nitric oxide and prostaglandin H2 to acetylcholine in skeletal muscle venules

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1997.272.6.h2541 ◽

1997 ◽

Vol 272 (6) ◽

pp. H2541-H2546 ◽

Cited By ~ 4

Author(s):

G. Dornyei ◽

G. Kaley ◽

A. Koller

Keyword(s):

Nitric Oxide ◽

Skeletal Muscle ◽

Perfusion Pressure ◽

Thromboxane A2 ◽

No Synthase ◽

Gracilis Muscle ◽

Vasoactive Agents ◽

Before And After ◽

Higher Doses

The role of endothelium in regulating venular resistance is not well characterized. Thus we aimed to elucidate the endothelium-derived factors involved in the mediation of responses of rat gracilis muscle venules to acetylcholine (ACh) and other vasoactive agents. Changes in diameter of perfusion pressure (7.5 mmHg)- and norepinephrine (10(-6) M)-constricted venules (approximately 225 microns in diam) to cumulative doses of ACh (10(-9) to 10(-4) M) and sodium nitroprusside (SNP, 10(-9) to 10(-4) M), before and after endothelium removal or application of various inhibitors, were measured. Lower doses of ACh elicited dilations (up to 42.1 +/- 4.7%), whereas higher doses of ACh resulted in smaller dilations or even constrictions. Endothelium removal abolished both ACh-induced dilation and constriction. In the presence of indomethacin (2.8 x 10(-5) M), a cyclooxygenase blocker, or SQ-29548 (10(-6) M), a thromboxane A2-prostaglandin H2 (PGH2) receptor antagonist, higher doses of ACh caused further dilation (up to 72.7 +/- 7%) instead of constriction. Similarly, lower doses of arachidonic acid (10(-9) to 10(-6) M) elicited dilations that were diminished at higher doses. These reduced responses were, however, reversed to substantial dilation by SQ-29548. The nitric oxide (NO) synthase blocker, N omega-nitro-L-arginine (L-NNA, 10(-4) M), significantly reduced the dilation to ACh (from 30.6 +/- 5.5 to 5.4 +/- 1.4% at 10(-6) M ACh). In contrast, L-NNA did not affect dilation to SNP. Thus ACh elicits the release of both NO and PGH2 from the venular endothelium.

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Short-term daily exercise activity enhances endothelial NO synthesis in skeletal muscle arterioles of rats

Journal of Applied Physiology ◽

10.1152/jappl.1994.76.5.2241 ◽

1994 ◽

Vol 76 (5) ◽

pp. 2241-2247 ◽

Cited By ~ 120

Author(s):

D. Sun ◽

A. Huang ◽

A. Koller ◽

G. Kaley

Keyword(s):

Nitric Oxide ◽

Skeletal Muscle ◽

Gracilis Muscle ◽

Myogenic Tone ◽

Control Group ◽

Free Solution ◽

Short Term ◽

Daily Exercise ◽

Exercise Activity ◽

Dose Dependent

We aimed to test the hypothesis that as a consequence of short-term daily bouts of exercise the control of arteriolar smooth muscle by endothelium is altered. Rats ran on a treadmill once a day, 5 days/wk, for 2–4 wk (with gradually increasing intensity, up to 26 min at 22 m/min at a 1% grade by the beginning of the 3rd wk and up to 38 min at 28 m/min at a 2% grade by the beginning of the 4th wk) while a control group remained sedentary (SED). Cannulated and pressurized arterioles of rat gracilis muscle developed spontaneous myogenic tone, which was slightly enhanced in exercised (EX) compared with SED rat arterioles. At 80 mmHg pressure, the passive (Ca(2+)-free solution) and active diameters of SED and EX rat arterioles were 105.4 +/- 3.8 and 55.1 +/- 2.3 microns and 107.1 +/- 3.4 and 50.2 +/- 2.2 microns, respectively. Dose-dependent dilations to sodium nitroprusside (10(-8)-10(-6) M) and constrictions to norepinephrine (10(-8)-10(-6) M) were not affected in EX arterioles, whereas dilations to adenosine (10(-6)-10(-4) M) were significantly reduced. In contrast, dose-dependent dilations to acetylcholine (ACh; 5 x 10(-9)-10(-7) M) and L-arginine [precursor of nitric oxide (NO); 10(-4)-10(-3) M] were significantly enhanced (by 33–78 and 57–75%, respectively) in arterioles of EX compared with those of SED rats. Responses of arterioles to sodium nitrite were not different in SED and EX groups.(ABSTRACT TRUNCATED AT 250 WORDS)

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EDHF mediates flow-induced dilation in skeletal muscle arterioles of female eNOS-KO mice

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.2001.280.6.h2462 ◽

2001 ◽

Vol 280 (6) ◽

pp. H2462-H2469 ◽

Cited By ~ 93

Author(s):

An Huang ◽

Dong Sun ◽

Mairead A. Carroll ◽

Houli Jiang ◽

Carolyn J. Smith ◽

...

Keyword(s):

Nitric Oxide ◽

Skeletal Muscle ◽

Endothelial Nitric Oxide Synthase ◽

Hexanoic Acid ◽

Gracilis Muscle ◽

Wild Type ◽

Basal Release ◽

Oxide Synthase ◽

Endothelial Nitric Oxide ◽

Cytochrome P 450

Vasodilation to increases in flow was studied in isolated gracilis muscle arterioles of female endothelial nitric oxide synthase (eNOS)-knockout (KO) and female wild-type (WT) mice. Dilation to flow (0–10 μl/min) was similar in the two groups, yet calculated wall shear stress was significantly greater in arterioles of eNOS-KO than in arterioles of WT mice. Indomethacin, which inhibited flow-induced dilation in vessels of WT mice by ∼40%, did not affect the responses of eNOS-KO mice, whereas miconazole and 6-(2-proparglyoxyphenyl)hexanoic acid (PPOH) abolished the responses. Basal release of epoxyeicosatrienonic acids from arterioles was inhibited by PPOH. Iberiotoxin eliminated flow-induced dilation in arterioles of eNOS-KO mice but had no effect on arterioles of WT mice. In WT mice, neither N ω-nitro-l-arginine methyl ester nor miconazole alone affected flow-induced dilation. Combination of both inhibitors inhibited the responses by ∼50%. 1 H-[1,2,4]oxadiazolo[4,3- a]quinoxalin-1-one (ODQ) alone inhibited flow-induced dilation by ∼49%. ODQ + indomethacin eliminated the responses. Thus, in arterioles of female WT mice, nitric oxide and prostaglandins mediate flow-induced dilation. When eNOS is inhibited, endothelium-derived hyperpolarizing factor substitutes for nitric oxide. In female eNOS-KO mice, metabolites of cytochrome P-450, via activation of large-conductance Ca2+-activated K+ channels of smooth muscle, mediate entirely the arteriolar dilation to flow.

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Mechanical strain-induced Ca2+waves are propagated via ATP release and purinergic receptor activation

AJP Cell Physiology ◽

10.1152/ajpcell.2000.279.2.c295 ◽

2000 ◽

Vol 279 (2) ◽

pp. C295-C307 ◽

Cited By ~ 104

Author(s):

H. Sauer ◽

J. Hescheler ◽

M. Wartenberg

Keyword(s):

Gap Junctions ◽

Purinergic Receptor ◽

Mechanical Strain ◽

Atp Release ◽

Anion Channel ◽

Receptor Activation ◽

Disulfonic Acid ◽

Channel Blockers ◽

Junctional Coupling ◽

Intracellular Ca

Mechanical strain applied to prostate cancer cells induced an intracellular Ca2+ (Cai 2+) wave spreading with a velocity of 15 μm/s. Cai 2+ waves were not dependent on extracellular Ca2+ and membrane potential because propagation was unaffected in high-K+ and Ca2+-free solution. Waves did not depend on the cytoskeleton or gap junctions because cytochalasin B and nocodazole, which disrupt microfilaments and microtubules, respectively, and 1-heptanol, which uncouples gap junctions, were without effects. Fluorescence recovery after photobleaching experiments revealed an absence of gap junctional coupling. Cai 2+ waves were inhibited by the purinergic receptor antagonists basilen blue and suramin; by pretreatment with ATP, UTP, ADP, UDP, 2-methylthio-ATP, and benzoylbenzoyl-ATP; after depletion of ATP by 2-deoxyglucose; and after ATP scavenging by apyrase. Waves were abolished by the anion channel inhibitors 5-nitro-2-(3-phenylpropylamino)benzoic acid, tamoxifen, 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid, niflumic acid, and gadolinium. ATP release following strain was significantly inhibited by anion channel blockers. Hence, ATP is secreted via mechanosensitive anion channels and activates purinergic receptors on the same cell or neighboring cells in an autocrine and paracrine manner, thus leading to Cai 2+ wave propagation.

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Characterizing voltage-dependent Ca2+ channels coupled to VIP release and NO synthesis in enteric synaptosomes

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00400.2001 ◽

2002 ◽

Vol 283 (5) ◽

pp. G1027-G1034 ◽

Cited By ~ 4

Author(s):

M. Kurjak ◽

A. Sennefelder ◽

M. Aigner ◽

V. Schusdziarra ◽

H. D. Allescher

Keyword(s):

Nitric Oxide ◽

No Synthase ◽

Channel Blockers ◽

Voltage Dependent ◽

Intracellular Ca ◽

P Type ◽

Ca2 Channels

In enteric synaptosomes of the rat, the role of voltage-dependent Ca2+channels in K+-induced VIP release and nitric oxide (NO) synthesis was investigated. Basal VIP release was 39 ± 4 pg/mg, and cofactor-substituted NO synthase activity was 7.0 ± 0.8 fmol · mg−1 · min−1. K+ depolarization (65 mM) stimulated VIP release Ca2+ dependently (basal, 100%; K+, 172.2 ± 16.2%; P < 0.05, n = 5). K+-stimulated VIP release was reduced by blockers of the P-type (ω-agatoxin-IVA, 3 × 10−8 M) and N-type (ω-conotoxin-GVIA, 10−6 M) Ca2+ channels by ∼50 and 25%, respectively, but not by blockers of the L-type (isradipine, 10−8 M), Q-type (ω-conotoxin-MVIIC, 10−6 M), or T-type (Ni2+, 10−6 M) Ca2+ channels. In contrast, NO synthesis was suppressed by ω-agatoxin-IVA, ω-conotoxin-GVIA, and isradipine by ∼79, 70, and 70%, respectively, whereas Ni2+ and ω-conotoxin-MVIIC had no effect. These findings are suggestive of a coupling of depolarization-induced VIP release primarily to the P- and N-type Ca2+ channels, whereas NO synthesis is presumably dependent on Ca2+ influx not only via the P- and N- but also via the L-type Ca2+ channel. In contrast, none of the Ca2+ channel blockers affected VIP release evoked by exogenous NO, suggesting that NO induces VIP secretion by a different mechanism, presumably involving intracellular Ca2+ stores.

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Increased expression of calreticulin is linked to ANG IV-mediated activation of lung endothelial NOS

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1999.277.4.l794 ◽

1999 ◽

Vol 277 (4) ◽

pp. L794-L801 ◽

Cited By ~ 7

Author(s):

Jawaharlal M. Patel ◽

Yong D. Li ◽

Jianliang Zhang ◽

Craig H. Gelband ◽

Mohan K. Raizada ◽

...

Keyword(s):

Nitric Oxide ◽

Endothelial Cells ◽

Catalytic Activity ◽

Endothelial Cell ◽

Ang Ii ◽

Tetraacetic Acid ◽

Free Medium ◽

Intracellular Ca ◽

Ang Iv ◽

Oxide Synthase

This study demonstrates that ANG IV-induced activation of lung endothelial cell nitric oxide synthase (ecNOS) is mediated through mobilization of Ca2+ concentration and by increased expression and release of the Ca2+ binding protein calreticulin in pulmonary artery endothelial cells (PAEC). In Ca2+-free medium and in the presence of the ANG II AT1 and AT2 receptor antagonists losartan and PD-123319 (1 μM each), respectively, ANG IV (5, 50, and 500 nM) significantly increased intracellular Ca2+ release in PAEC ( P < 0.05 for all concentrations). In contrast, ANG IV-mediated activation of ecNOS was abolished by the intracellular Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid-AM. ANG IV stimulation resulted in significantly increased expression of calreticulin in cells as well as release of calreticulin into the medium of cells as early as 2 h after ANG IV stimulation ( P < 0.05). Catalytic activity of purified ecNOS in the absence of calmodulin was increased in a concentration-dependent fashion by calreticulin. Immunocoprecipitation studies revealed that ecNOS and calreticulin were coprecipitated in ANG IV-stimulated PAEC. These results demonstrate that ANG IV-mediated activation of ecNOS is regulated by intracellular Ca2+ mobilization and by increased expression of calreticulin, which appears to involve interaction of ecNOS and calreticulin proteins in PAEC.

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Capacitative Ca2+ entry in enteric glia induced by thapsigargin and extracellular ATP

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1998.275.3.g550 ◽

1998 ◽

Vol 275 (3) ◽

pp. G550-G555 ◽

Cited By ~ 11

Author(s):

George A. Sarosi ◽

Douglas C. Barnhart ◽

Douglas J. Turner ◽

Michael W. Mulholland

Keyword(s):

Nitric Oxide ◽

Similar Degree ◽

Channel Blockers ◽

Nitric Oxide Synthase Inhibitor ◽

Enteric Glia ◽

Enteric Glial Cells ◽

Intracellular Ca ◽

Specific Antagonist ◽

Synthase Inhibitor ◽

Oxide Synthase

Mobilization of intracellular Ca2+ stores is coupled to Ca2+ influx across the plasma membrane, a process termed capacitative Ca2+ entry. Capacitative Ca2+ entry was examined in cultured guinea pig enteric glia exposed to 100 μM ATP, an inositol trisphosphate-mediated Ca2+-mobilizing agonist, and to 1 μM thapsigargin, an inhibitor of microsomal Ca2+ ATPase. Both agents caused mobilization of intracellular Ca2+stores followed by influx of extracellular Ca2+. This capacitative Ca2+ influx was inhibited by Ni2+ (88 ± 1%) and by La3+ (87 ± 1%) but was not affected by L- or N-type Ca2+channel blockers. Pretreatment of glia with 100 nM phorbol 12-myristate 13-acetate for 24 h decreased capacitative Ca2+ entry by 48 ± 2%. Chelerythrine (0.1–10 μM), a specific antagonist of protein kinase C (PKC), dose dependently inhibited capacitative Ca2+ entry. The nitric oxide synthase inhibitor N G-nitro-l-arginine (1 mM) decreased Ca2+ influx by 42 ± 1%. Capacitative Ca2+ entry was inhibited to a similar degree by the guanylate cyclase inhibitor (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one). Capacitative Ca2+ entry occurs in enteric glial cells via lanthanum-inhibitable channels through a process regulated by PKC and nitric oxide.

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Localization of sperm intracellular Ca2+ keeps fertilizability in the newt vas deferens

Reproduction ◽

10.1530/rep-19-0252 ◽

2020 ◽

Vol 159 (3) ◽

pp. 339-349

Author(s):

Nanae Makino ◽

Nozomi Sato ◽

Eriko Takayama-Watanabe ◽

Akihiko Watanabe

Keyword(s):

Sperm Motility ◽

Salt Solution ◽

Acrosome Reaction ◽

Vas Deferens ◽

Sperm Quality ◽

Ethylene Glycol Tetraacetic Acid ◽

Channel Blockers ◽

Tetraacetic Acid ◽

Intracellular Ca ◽

Principal Piece

Sperm intracellular Ca2+ is crucial for the induction of sperm-egg interaction, but little is known about the significance of Ca2+ maintenance prior to induction. In sperm of the newt Cynops pyrrhogaster, intracellular Ca2+ is localized to the midpiece during storage in the vas deferens, while extracellular Ca2+ is influxed in modified Steinberg’s salt solution to promote a spontaneous acrosome reaction related to the decline of sperm quality. In the present study, sperm from the vas deferens were loaded with the Ca2+ indicator Fluo8H, and changes in Ca2+ localization in modified Steinberg’s salt solution were examined. Calcium ions expanded from the cytoplasmic area of the midpiece to the entire tail in most sperm during a 1-h incubation and localized to the principal piece in some sperm within 24 h. Similar changes in Ca2+ localization were observed in reconstructed vas deferens solution that included ions and pH at equivalent levels to those in the vas deferens fluid. Sperm with Ca2+ localization in the entire tail or the principal piece weakened or lost responsiveness to sperm motility-initiating substances, which trigger sperm motility for fertilization, but responded to a trigger for acrosome reaction. The change in Ca2+ localization was delayed and transiently reversed by ethylene glycol tetraacetic acid or a mixture of Ca2+ channel blockers including Ni2+ and diltiazem. These results suggest that C. pyrrhogaster sperm localize intracellular Ca2+ to the midpiece through Ca2+ transport in the vas deferens to allow for responses to sperm motility-initiating substances.

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