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This Article Full Text Full Text (PDF) All Versions of this Article: 570/2/309    most recent jphysiol.2005.100800v1 Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ben-Amor, N. Articles by Rosado, J. A. Search for Related Content PubMed PubMed Citation Articles by Ben-Amor, N. Articles by Rosado, J. A. Related Collections Cellular

¹ Unité de Recherche de Biochimie, Institute Superieur de Biotechnologie, Monastir, Tunisia
² Department of Physiology, University of Extremadura, 10071 Cáceres, Spain

A major pathway for Ca²⁺ entry in non-excitable cells is activated following depletion of intracellular Ca²⁺ stores. A de novo conformational coupling between elements in the plasma membrane (PM) and Ca²⁺ stores has been proposed as the most likely mechanism to activate this capacitative Ca²⁺ entry (CCE) in several cell types, including platelets. Here we report that a cytochrome P450 metabolite, 5,6-EET, might be a component of the de novo conformational coupling in human platelets. In these cells, 5,6-EET induces divalent cation entry without having any detectable effect on Ca²⁺ store depletion. 5,6-EET-induced Ca²⁺ entry was sensitive to the CCE blockers 2-APB, lanthanum, SKF-96365 and nickel and impaired by incubation with anti-hTRPC1 antibody. Ca²⁺ entry stimulated by low concentrations of thapsigargin, which selectively depletes the dense tubular system and induces EET production, was impaired by the cytochrome P450 inhibitor 17-ODYA, which has no effect on CCE mediated by depletion of the acidic stores using 2,5-di-(tert-butyl)-1,4-hydroquinone. We have found that 5,6-EET-induced Ca²⁺ entry requires basal levels of H₂O₂, which might maintain a redox state favourable for this event. Finally, our results indicate that 5,6-EET induces the activation of tyrosine kinase proteins and the reorganization of the actin cytoskeleton, which might provide a support for the transport of portions of the Ca²⁺ store towards the PM to facilitate de novo coupling between IP₃R type II and hTRPC1 detected by coimmunoprecipitation. We propose that the involvement of 5,6-EET in TG-induced coupling between IP₃R type II and hTRPC1 and subsequently CCE is compatible with the de novo conformational coupling in human platelets.

(Received 26 October 2005; accepted after revision 17 November 2005; first published online 24 November 2005)
Corresponding author J. A. Rosado: Department of Physiology, University of Extremadura, Cáceres 10071, Spain. Email: jarosado{at}unex.es

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