Aim: 

Platelet stimulation with the physiological agonist thrombin results in elevation in cytoplasmic free Ca²⁺ concentration ([Ca²⁺]_c) due to Ca²⁺ release from intracellular stores and entry from the extracellular medium. Two different intracellular Ca²⁺ stores have been described in human platelets, the dense tubular system (DTS) and the lysosomal-like acidic stores (López et al, 2005). Here we provide evidence for the contribution of the acidic stores in thrombin-induced platelet aggregation.

Methods: 

The percentage, rate and lag-time of aggregation in washed platelets were monitored using a Chronolog (Havertown, Pa, USA) aggregometer at 37 °C under stirring at 1200 rpm.

Results: 

Platelet aggregation induced by thrombin is reduced in a Ca²⁺ free medium. Depletion of the acidic Ca²⁺ compartments by treatment with the SERCA3 selective inhibitor 2,5-di-(tert-butyl)-1,4-hydroquinone (TBHQ) significantly attenuated thrombin-evoked platelet aggregation (p<0.05; Student's t test). In the presence of TBHQ, platelet aggregation induced by the PAR-1 and PAR-4 agonist peptides, SFLLRN and AYPGKF, respectively, was significantly reduced (p<0.05). In cells with depleted acidic stores, activation of GPIb-IX-V by thrombin resulted in reduced or no platelet aggregation in a medium containing 1 mM Ca²⁺or in a Ca²⁺-free medium, respectively.

Conclusion: 

Our results indicate that Ca²⁺ accumulation in the acidic Ca²⁺ pools is required for platelet aggregation induced by activation of the G-coupled PAR-1 and PAR-4 thrombin receptors and, by the occupation of the leucine-rich glycoprotein GPIb-IX-V and provide evidence supporting a functional role of the lysosomal-like acidic Ca²⁺ stores in human platelets.

References: López J.J., Camello-Almaraz C., Pariente J.A., Salido G.M., Rosado J.A. 2005. Ca²⁺ accumulation into acidic organelles mediated by Ca²⁺- and vacuolar H⁺-ATPases in human platelets. Biochem J. 390, 243-252.

Supported by BFU2007-60104.