A ROLE FOR THE NA+/CA2+ EXCHANGER IN STORE-OPERATED CALCIUM ENTRY IN HUMAN PLATELETS
Abstract number: P-T-252
Harper1 A.G.S., Sage1 S.O.
1Department of Physiology, Development and Neuroscience, University of Cambridge, Cambridge, United Kingdom
How-to-cite Harper AGS, Sage SO. A ROLE FOR THE NA+/CA2+ EXCHANGER IN STORE-OPERATED CALCIUM ENTRY IN HUMAN PLATELETS. J Thromb Haemost 2007; 5 Supplement 2: P-T-252
Introduction: We have previously demonstrated a role for actin cytoskeleton reorganisation in store-operated calcium entry (SOCE) in human platelets and interpreted this as evidence for a de novo conformational coupling step involving hTRPC1 and the type II IP3 receptor in SOCE activation. Here we reassess this model in light of evidence that the actin cytoskeleton supports the operation of Na+/Ca2+ exchangers (NCXs) in reverse mode.
Methods: Human platelets were loaded with Fura-2 to monitor changes in cytosolic Ca2+ concentration and Mn2+ entry, or loaded with SBFI to monitor cytosolic Na+ concentration. Data are presented as mean S.E.M. Significance was determined using Student's paired t-test.
Results: The actin polymerization inhibitor cytochalasin D (Cyt D) reduced Ca2+ entry into thapsigargin (TG)-treated platelets to 44.8 9.7% of control (n = 3, p > 0.05) but was without effect on the entry of Mn2+ (104.9 18.2% of control; n = 10, p > 0.05) or Na+ (107.2 18.7% of control; n = 6, p > 0.05). Jasplakinolide (JP), which induces actin polymerization, inhibited TG-evoked Ca2+ entry as previously described (40.5 8.0% of control; n = 2) but did not inhibit TG-evoked Mn2+ (110.5 7.5% of control, n = 5, p > 0.05) or Na+ (174.3 22.7% of control; n = 6; p < 0.05) entry. An anti-hTRPC1 antibody inhibited TG-evoked entry of all three cations, indicating that they all permeate an hTRPC1-containing store-operated channel (SOC). These results indicate that the reorganization of the actin cytoskeleton is not involved in SOC activation. The NCX inhibitors, KB-R7943 or SN-6, caused a dose-dependent inhibition of Ca2+ but not Mn2+ or Na+ entry into TG-treated platelets. The effects of KB-R7943 was not additive with that of Cyt D, suggesting a common point of action.
Conclusions: These results indicate a role for two Ca2+ permeable pathways activated following Ca2+ store depletion in human platelets: A Ca2+-permeable, hTRPC1-containing SOC and reverse Na+/Ca2+ exchange.
Supported by the British Heart Foundation.
To cite this abstract use the following format:
Journal of Thrombosis and Haemostasis 2007; Volume 5, Supplement 2: abstract number
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