Depletion of intracellular Ca2+ stores activates a Ca(2+)-selective channel in vascular endothelium

1994 ◽  
Vol 267 (4) ◽  
pp. C920-C925 ◽  
Author(s):  
L. Vaca ◽  
D. L. Kunze
Keyword(s):  
Vascular Endothelium ◽  
Reversal Potential ◽  
Inward Rectification ◽  
Ion Permeability ◽  
Channel Activity ◽  
Permeability Ratio ◽  
Extracellular Solution ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Selective Channel

The present study was designed to identify the channel responsible for Ca2+ influx after depletion of intracellular Ca2+ stores. Different maneuvers that deplete intracellular Ca2+ stores activated a Ca(2+)-selective channel. Superfusion of single bovine aortic endothelial cells with 50 nmol/l bradykinin, 10 mumol/l ATP, or 10 mumol/l 2,5-di(tert-butyl)-1,4-benzohydroquinone produced activation of channels of the same amplitude in cell-attached patches. Channel activity declined within the first minute after patch excision. The channel showed strong inward rectification and a reversal potential of 0 mV in symmetrical sodium sulfate (Na2SO4) solution. Under these conditions, the conductance was 5 pS in the inward direction. Addition of 10 mmol/l Ca2+ to the extracellular solution shifted the reversal potential to +30 +/- 5 mV, and the conductance for inward current was 11 pS. The reversal potential was used to calculate an ion permeability ratio of Ca2+/Na+ > 10:1.

scholarly journals Prostacyclin and beta-adrenergic catecholamines inhibit arachidonate release and PGI2 synthesis by vascular endothelium

Blood ◽  
1981 ◽  
Vol 58 (3) ◽  
pp. 514-517 ◽  
Author(s):  
B Adler ◽  
MA Jr Gimbrone ◽  
AI Schafer ◽  
RI Handin
Keyword(s):  
Endothelial Cells ◽  
Cyclic Amp ◽  
Vascular Endothelium ◽  
Calcium Ionophore ◽  
Phosphodiesterase Activity ◽  
Dibutyryl Cyclic Amp ◽  
Membrane Phospholipids ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Arachidonate Release

Abstract We have investigated the mechanism by which cyclic AMP inhibits PGI2 synthesis in cultured bovine aortic endothelial cells. Inhibition of cyclic AMP phosphodiesterase activity by 3-isobutyl-1-methylxanthine (IBMX) blocks calcium ionophore-induced PGI2 production by 62%. The addition of 3 mM dibutyryl cyclic AMP, alone with IBMX, increases the inhibition to 96%. Release o 3H-arachidonate from membrane phospholipids was inhibited 25% by dibutyryl cyclic AMP, 48% by IBMX, and 76% by isoproterenol plus IBMX. Inhibition by isoproterenol was reversed by 10 micro M propranolol. Release of 3H-arachidonate was also reduced 75% by a combination of 10 micro M PGI2 and 3 mM IBMX. We conclude that hormones like isoproterenol and PGI2 may regulate endothelial cell PGI2 biosynthesis by increasing intracellular cyclic AMP, which then inhibits release of endogenous arachidonate from membrane phospholipids.

scholarly journals Prostacyclin and beta-adrenergic catecholamines inhibit arachidonate release and PGI2 synthesis by vascular endothelium

Blood ◽  
1981 ◽  
Vol 58 (3) ◽  
pp. 514-517
Author(s):  
B Adler ◽  
MA Jr Gimbrone ◽  
AI Schafer ◽  
RI Handin
Keyword(s):  
Endothelial Cells ◽  
Cyclic Amp ◽  
Vascular Endothelium ◽  
Calcium Ionophore ◽  
Phosphodiesterase Activity ◽  
Dibutyryl Cyclic Amp ◽  
Membrane Phospholipids ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Arachidonate Release

We have investigated the mechanism by which cyclic AMP inhibits PGI2 synthesis in cultured bovine aortic endothelial cells. Inhibition of cyclic AMP phosphodiesterase activity by 3-isobutyl-1-methylxanthine (IBMX) blocks calcium ionophore-induced PGI2 production by 62%. The addition of 3 mM dibutyryl cyclic AMP, alone with IBMX, increases the inhibition to 96%. Release o 3H-arachidonate from membrane phospholipids was inhibited 25% by dibutyryl cyclic AMP, 48% by IBMX, and 76% by isoproterenol plus IBMX. Inhibition by isoproterenol was reversed by 10 micro M propranolol. Release of 3H-arachidonate was also reduced 75% by a combination of 10 micro M PGI2 and 3 mM IBMX. We conclude that hormones like isoproterenol and PGI2 may regulate endothelial cell PGI2 biosynthesis by increasing intracellular cyclic AMP, which then inhibits release of endogenous arachidonate from membrane phospholipids.

IP3-activated Ca2+ channels in the plasma membrane of cultured vascular endothelial cells

1995 ◽  
Vol 269 (3) ◽  
pp. C733-C738 ◽  
Author(s):  
L. Vaca ◽  
D. L. Kunze
Keyword(s):  
Endothelial Cells ◽  
Plasma Membrane ◽  
Single Channel ◽  
Vascular Endothelial Cells ◽  
Channel Activity ◽  
Open Probability ◽  
Aortic Endothelial Cells ◽  
Control Value ◽  
Selective Channel ◽  
Inside Out

Although it is clear that D-myo-inositol 1,4,5-trisphosphate (IP3) plays an important role in the activation of Ca2+ influx, the mechanisms by which this occurs remain controversial. In an attempt to determine the role of IP3 in the activation of Ca2+ influx, patch-clamp single-channel experiments in the cell-attached, inside-out, and outside-out configurations were performed on cultured bovine aortic endothelial cells (BAEC). The results presented indicate that both IP3 and intracellular Ca2+ can modulate the activity of a Ca(2+)-selective channel found in the plasma membrane of these cells. Addition of 10 microM IP3 increased channel open probability (P(o)) from a control value of 0.12 +/- 0.05 to 0.7 +/- 0.13 at a constant intracellular Ca2+ of 1 nM in excised inside-out patches. D-Myo-inositol 1,3,4,5-tetrakisphosphate at 50 microM was ineffective in altering channel P(o). Channel activity declined after approximately 2 min in the continuous presence of IP3. Three to four minutes after addition of IP3, channel P(o) was reduced from 0.7 +/- 0.2 to 0.2 +/- 0.1, indicating that an additional regulator might be required to maintain channel activity in excised patches. The channel was reversibly blocked by application of 1 microgram/ml heparin to the intracellular side of inside-out patches. This Ca(2+)-selective channel is indistinguishable from the depletion-activated Ca2+ channel we have previously described in BAEC.

Bradykinin-activated calcium influx pathway in bovine aortic endothelial cells

1992 ◽  
Vol 262 (4) ◽  
pp. H942-H948 ◽  
Author(s):  
D. Mendelowitz ◽  
K. Bacal ◽  
D. L. Kunze
Keyword(s):  
Endothelial Cells ◽  
Membrane Potential ◽  
Normal Saline ◽  
Voltage Pulse ◽  
Calcium Influx ◽  
Reversal Potential ◽  
Voltage Response ◽  
Aortic Endothelial Cells ◽  
Inward Current ◽  
Bovine Aortic Endothelial Cells

Clusters of electrically coupled endothelial cells were used to characterize a bradykinin (BK)-activated Ca2+ influx pathway. Spatial voltage control of clusters containing three to eight cells, evaluated as the ratio of the voltage response in one cell to a voltage pulse in the most distant cell of the cluster, was 0.96 at a holding potential of 0 mV in normal saline bath and 0.88 in the presence of BK. BK activated an inward current that was carried by either Na+ or Ca2+ when the membrane potential was held at -60 mV. Current was activated within 3 s of application of BK and peaked within 1 min. With Ca2+ as the permeable extracellular ion the current was stable for 1-3 min and then declined over a period of 5-8 min in the continued presence of BK. However, when Na+ carried the current it was sustained over a 10-min test period. The reversal potential of the BK-activated current was near 0 mV, suggesting activation of a nonspecific cation channel(s). The inward current at -60 mV averaged 13 +/- 4.5 pA (n = 9)/cell in Ca2+ and 12.2 +/- 9.3 pA (n = 5)/cell in Na+. Both Na+ and Ca2+ currents were blocked by 200 microM lanthanum.

Angiotensin II activation of Ca(2+)-permeant nonselective cation channels in rat adrenal glomerulosa cells

1996 ◽  
Vol 271 (5) ◽  
pp. C1705-C1715 ◽  
Author(s):  
D. P. Lotshaw ◽  
F. Li
Keyword(s):  
Angiotensin Ii ◽  
Channel Gating ◽  
Reversal Potential ◽  
Inward Rectification ◽  
Ang Ii ◽  
Permeability Ratio ◽  
Open Probability ◽  
Glomerulosa Cells ◽  
Single Channel Currents ◽  
Inside Out

A Ca(2+)-permeant, nonselective cation channel was observed in cell-attached and inside-out membrane patches from rat adrenal glomerulosa cells maintained in primary cell culture. In cell-attached patches under near physiological ionic conditions, single-channel currents exhibited a reversal potential near -10 mV, inward rectification, a nearly linear slope conductance between 0 and -80 mV of 17.4 pS, and voltage-dependent block at potentials more negative than -80 mV. Channels exhibiting identical conductance and gating properties were observed in inside-out patches; however, channel gating ran down within minutes in this configuation. In the inside-out configuration, channel gating did not require cytosolic Ca2+ (Ca2+ < 10(-9) M), and inward rectification was relieved by removal of intracellular Mg2+. Relative ionic permeability was calculated using reversal potential measurements from inside-out patches under bi-ionic conditions. The channel discriminated poorly among monovalent cations (PLi > PK > PCs > PNa) and was not significantly permeable to anions. The channel was permeable to Ca2+, exhibiting a relative permeability ratio of 0.29 PCa/PNa) when measured with 110 mM Ca2+ on the intracellular face and a permeability ratio of 4.38 (PCa/PNa) with 110 mM Ca2+ on the extracellular face. Channel gating behavior was episodic with open times ranging from milliseconds to tens of seconds and closed times lasting up to several minutes or longer. Channel gating appeared to be relatively voltage independent except that mean channel open time and open probability were reduced by membrane hyperpolarization. In cell-attached patches, bath application of 1 nM angiotensin II (ANG II) increased the channel open probability, primarily affecting channels exhibiting a low open probability, primarily affecting channels exhibiting a low open probability before stimulation. With the use of nystatin perforated-patch current clamp to measure membrane potential, ANG II was observed to induce large transient membrane depolarizations, consistent with activation of an inward current. We hypothesize that this channel is an important component of ANG II-induced membrane depolarization and Ca2+ influx during stimulation of aldosterone secretion.

Ornithine Decarboxylase Activity in Cultured Endothelial Cells Stimulated by Serum, Thrombin and Serotonin

1978 ◽  
Vol 39 (02) ◽  
pp. 496-503 ◽  
Author(s):  
P A D’Amore ◽  
H B Hechtman ◽  
D Shepro
Keyword(s):  
Endothelial Cells ◽  
Ornithine Decarboxylase ◽  
Decarboxylase Activity ◽  
Aortic Endothelial Cells ◽  
Ornithine Decarboxylase Activity ◽  
Rate Limiting Step ◽  
Bovine Aortic Endothelial Cells ◽  
Limiting Step ◽  
Rate Limiting ◽  
Serum Serotonin

SummaryOrnithine decarboxylase (ODC) activity, the rate-limiting step in the synthesis of polyamines, can be demonstrated in cultured, bovine, aortic endothelial cells (EC). Serum, serotonin and thrombin produce a rise in ODC activity. The serotonin-induced ODC activity is significantly blocked by imipramine (10-5 M) or Lilly 11 0140 (10-6M). Preincubation of EC with these blockers together almost completely depresses the 5-HT-stimulated ODC activity. These observations suggest a manner by which platelets may maintain EC structural and metabolic soundness.

Effects of Dipyrone on Prostaglandin Production by Human Platelets and Cultured Bovine Aortic Endothelial Cells

1983 ◽  
Vol 49 (02) ◽  
pp. 132-137 ◽  
Author(s):  
A Eldor ◽  
G Polliack ◽  
I Vlodavsky ◽  
M Levy
Keyword(s):  
Endothelial Cells ◽  
Arachidonic Acid ◽  
Competitive Inhibition ◽  
Inhibitory Effect ◽  
Human Platelets ◽  
Ionophore A23187 ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

SummaryDipyrone and its metabolites 4-methylaminoantipyrine, 4-aminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoan- tipyrine inhibited the formation of thromboxane A2 (TXA2) during in vitro platelet aggregation induced by ADP, epinephrine, collagen, ionophore A23187 and arachidonic acid. Inhibition occurred after a short incubation (30–40 sec) and depended on the concentration of the drug or its metabolites and the aggregating agents. The minimal inhibitory concentration of dipyrone needed to completely block aggregation varied between individual donors, and related directly to the inherent capacity of their platelets to synthesize TXA2.Incubation of dipyrone with cultured bovine aortic endothelial cells resulted in a time and dose dependent inhibition of the release of prostacyclin (PGI2) into the culture medium. However, inhibition was abolished when the drug was removed from the culture, or when the cells were stimulated to produce PGI2 with either arachidonic acid or ionophore A23187.These results indicate that dipyrone exerts its inhibitory effect on prostaglandins synthesis by platelets or endothelial cells through a competitive inhibition of the cyclooxygenase system.

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