Depletion of intracellular Ca2+ stores sensitizes the flow-induced Ca2+ influx in rat endothelial cells.
Hemodynamic shear stress elicits a rise in endothelial [Ca2+]i, which may serve as a key second messenger to regulate many flow-associated physiological and biochemical processes. In the present study, we used Mn2+ quenching of fluorescent dye Fluo3 as an assay to investigate the Ca2+ influx of rat aortic endothelial cells in response to flow. We found that the Ca2+ signaling in response to flow could be greatly influenced by the status of intracellular Ca2+ stores. Depletion of intracellular Ca2+ stores by thapsigargin (4 micromol/L) or cyclopiazonic acid (10 micromol/L) drastically sensitized the Ca2+ influx in response to flow. Ca2+-mobilizing agonist bradykinin (100 nmol/L) or ATP (100 micromol/L) had similar sensitizing effect. The effect of bradykinin or ATP was blocked by Xestospongin C and U73122, suggesting that the sensitization was related to the IP3-mediated store depletion. On the other hand, the Mn2+ quenching in response to flow was greatly reduced by ochratoxin A (100 nmol/L), an agent that could increase the filling state of intracellular Ca2+ stores. In addition, we found that depletion-sensitized Ca2+ influx in response to flow was mediated by a PKG-inhibitable cation channel and that the influx was affected by membrane potential and K+ channel activity. In conclusion, the present study argues for a critical role of intracellular Ca2+ status in determining the Ca2+ signaling in response to flow and it provides a general mechanistic explanation for the stimulatory role of blood-borne agonists on flow-induced Ca2+ influx.
Kwan
Hiu-Yee
HY
Department of Physiology, Faculty of Medicine, Chinese University of Hong Kong, Hong Kong, China.
Leung
Pan-Cheung
PC
Huang
Yu
Y
Yao
Xiaoqiang
X
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000494:pharmacology; D000814:Aniline Compounds; D000818:Animals; D001920:Bradykinin / Q000037:antagonists & inhibitors / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000493:pharmacokinetics; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007424:Intracellular Fluid / Q000378:metabolism*; D047028:Macrocyclic Compounds; D008297:Male; D008345:Manganese / Q000493:pharmacokinetics; D009793:Ochratoxins / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D020442:Poloxamer; D015221:Potassium Channels / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors; D013050:Spectrometry, Fluorescence; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors; D014966:Xanthenes
D000814:Aniline Compounds; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D053496:Inositol 1,4,5-Trisphosphate Receptors; D047028:Macrocyclic Compounds; D009793:Ochratoxins; D010080:Oxazoles; D015221:Potassium Channels; D018160:Receptors, Cytoplasmic and Nuclear; D014966:Xanthenes; C108644:xestospongin A; D020442:Poloxamer; C025589:ochratoxin A; C059715:Fluo-3; D008345:Manganese; D000255:Adenosine Triphosphate; D017869:Cyclic GMP-Dependent Protein Kinases; D010738:Type C Phospholipases; D001920:Bradykinin; D002118:Calcium