Capacitative Ca(2+) entry in vascular endothelial cells is mediated via pathways sensitive to 2 aminoethoxydiphenyl borate and xestospongin C.
1. Agonists increase endothelial cell intracellular Ca(2+), in part, by capacitative entry, which is triggered by the filling state of intracellular Ca(2+) stores. It has been suggested that depletion of endoplasmic reticulum (ER) Ca(2+) stores either leads to a physical coupling between the ER and a plasma membrane channel, or results in production of an intracellular messenger which affects the gating of membrane channels. As an axis involving the IP(3) receptor has been implicated in a physical coupling mechanism the aim of this study was to examine the effects of the putative IP(3) receptor antagonists/modulators, 2 aminoethoxydiphenyl borate (2APB) and xestospongin C, on endothelial cell Ca(2+) entry. 2. Studies were conducted in fura 2 loaded cultured bovine aortic endothelial cells and endothelial cells isolated from rat heart. 3. 2APB (30 - 300 microM) inhibited Ca(2+) entry induced by both agonists (ATP 1 microM, bradykinin 0.1 microM) and receptor-independent mechanisms (thapsigargin 1 microM, ionomycin 0.5 and 5 microM). 2APB did not diminish endothelial cell ATP-induced production of IP(3) nor effect in vitro binding of [(3)H]-IP(3) to an adrenal cortex binding protein. Capacitative Ca(2+) entry was also blocked by disruption of the actin cytoskeleton with cytochalasin (100 nM) while the initial Ca(2+) release phase was unaffected. 4. Similarly to 2APB, xestospongin C (3 - 10 microM) inhibited ATP-induced Ca(2+) release and capacitative Ca(2+) entry. Further, xestospongin C inhibited capacitative Ca(2+) entry induced by thapsigargin (1 microM) and ionomycin (0.5 microM). 5. The data are consistent with a mechanism of capacitative Ca(2+) entry in vascular endothelial cells which requires (a) IP(3) receptor binding and/or an event distal to the activation of the ER receptor and (b) a spatial relationship, dictated by the cytoskeleton, between Ca(2+) release and entry pathways.
Bishara
Nour B
NB
Microvascular Biology Group, School of Medical Sciences, RMIT University, Bundoora, Victoria 3083, Australia.
Murphy
Timothy V
TV
Hill
Michael A
MA
D000199:Actins / Q000187:drug effects / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D001896:Boron Compounds / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D002417:Cattle; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D053496:Inositol 1,4,5-Trisphosphate Receptors; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D047028:Macrocyclic Compounds; D010080:Oxazoles / Q000494:pharmacology*; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013237:Stereoisomerism; D019284:Thapsigargin / Q000494:pharmacology
D000199:Actins; D001896:Boron Compounds; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007476:Ionophores; D047028:Macrocyclic Compounds; D010080:Oxazoles; D018160:Receptors, Cytoplasmic and Nuclear; C108644:xestospongin A; D015759:Ionomycin; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium