1 Agonists increase endothelial cell intracellular Ca 2+ , in part, by capacitative entry, which is triggered by the ®lling 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 a ects 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 e ects 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 mM) inhibited Ca 2+ entry induced by both agonists (ATP 1 mM, bradykinin 0.1 mM) and receptor-independent mechanisms (thapsigargin 1 mM, ionomycin 0.5 and 5 mM). 2APB did not diminish endothelial cell ATP-induced production of IP 3 nor e ect 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 una ected. 4 Similarly to 2APB, xestospongin C (3 ± 10 mM) inhibited ATP-induced Ca 2+ release and capacitative Ca 2+ entry. Further, xestospongin C inhibited capacitative Ca 2+ entry induced by thapsigargin (1 mM) and ionomycin (0.5 mM). 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.