Reciprocal regulation of capacitative and non-capacitative Ca2+ entry in A7r5 vascular smooth muscle cells: only the latter operates during receptor activation

Abstract

In A7r5 vascular smooth muscle cells, Arg8-vasopressin (AVP) stimulates phospholipase C leading to activation of two distinct Ca2+ entry pathways. The capacitative Ca2+ entry (CCE) pathway is activated by depletion of Ca2+ stores, is permeable to Mn2+, Ba2+ and Ca2+, and is selectively blocked by Gd3+ (1μM). A7r5 cells also express a non-capacitative Ca2+ entry (NCCE) pathway, which is activated by arachidonic acid that is released by the sequential activities of phospholipase C and diacylglycerol lipase. This pathway is permeable to Sr2+, Ba2+ and Ca2+ and selectively blocked by (R,S)-(3,4-dihydro-6,7-dimethoxy-isochinolin-1-yl)-2-phenyl-N,N-di[2-(2,3,4-trimethoxyphenyl)ethyl]acetamid mesylate ('LOE-908'). We use these selective tools to show that AVP, via the same signalling pathway that leads to activation of NCCE, also inhibits CCE and that the inhibition is not due to depolarization of the plasma membrane. Using the selective inhibitors to resolve the contributions of each Ca2+ entry pathway during stimulation with AVP, we establish that reciprocal regulation of CCE and NCCE by arachidonic acid ensures that only NCCE is active in the presence of AVP, whereas CCE is active only after its removal. NCCE and CCE are therefore activated in a strict temporal sequence: NCCE first and then CCE. Because Ca2+ passing through different Ca2+ entry pathways can selectively regulate different responses, reciprocal regulation of CCE and NCCE may allow a stimulus to first evoke a response and then recruit actively a different response when the stimulus is removed.

Long Lasting Inhibition of Adenylyl Cyclase Selectively Mediated by Inositol 1,4,5-Trisphosphate-evoked Calcium Release

“…2ϩ Entry-Depolarization (by the addition of HBS containing 53 mM KCl in place of NaCl) stimulated an increase in [Ca 2ϩ ] i which was prevented by 100 nM nimodipine or 10 M verapamil and potentiated by 100 nM BAYK8644, confirming the involvement of L-type Ca 2ϩ channels (39,49) (Fig. 5A).…”

“…Our initial intention had been to explore the relative effectiveness of Ca 2ϩ entry via CCE and NCCE pathways in causing inhibition of AC, but we began our experiments (and then continued) using a variant of the cell line in which AVP neither inhibits CCE nor activates NCCE (39). In the cells used for this study, therefore, all AVP-evoked Ca 2ϩ entry is mediated by CCE.…”

“…In A7r5 cells, Arg 8 -vasopressin (AVP), via V 1A receptors (37), stimulates phospholipase C and thereby both release of Ca 2ϩ from intracellular stores and Ca 2ϩ entry across the plasma membrane. The latter involves a complex interplay between different Ca 2ϩ channels: AVP promotes opening of L-type voltage-gated Ca 2ϩ channels (38), and it can reciprocally regulate CCE and a noncapacitative Ca 2ϩ entry (NCCE) pathway (39,40). Given the importance of Ca 2ϩ -cAMP interactions in vascular smooth muscle, the evidence that not all Ca 2ϩ signals are equally effective in regulating AC (7), and evidence that AVP inhibits AC in A7r5 cells (33), we decided to examine whether inhibition of AC in these cells was regulated selectively by distinct Ca 2ϩ entry pathways.…”

Arachidonic Acid–Induced Ca2+ Entry Is Involved in Early Steps of Tumor Angiogenesis

“…A reciprocal regulation of AA-activated NSOCE and SOCE has been reported by several authors (36,37,69,70). It has been suggested that AA coordinates the contribution of different receptor-regulated pathways such that each is activated in a specific temporal sequence.…”

“…It has been suggested that AA coordinates the contribution of different receptor-regulated pathways such that each is activated in a specific temporal sequence. By activating NSOCE, and simultaneously inhibiting SOCE, AA ensures that, when the extracellular stimulus is present, only the NSOCE pathway contributes to Ca 2+ entry whereas SOCE contributes only after removal of the stimulus (37). The mechanisms underlying this phenomenon are still unclear.…”