Nonreversible loss of platelet aggregability induced by calcium deprivation

Platelets lose their ability to aggregate when deprived of divalent cations. This usually was studied by incubating human citrated platelet-rich plasma with EDTA or EGTA and then adding enough CaCl2 to combine with the chelating agent. Incubation for 5-7 min at 37 degrees C caused irreversible loss of the platelets' ability to adhere to glass and to aggregate with ADP, epinephrine, A23187, vasopressin, or serotonin or upon rewarming after chilling and markedly reduced aggregation with collagen or thrombin. Control samples incubated with saline, CaEDTA, or CaEGTA were not inhibited. Untreated platelets washed and incubated in solutions treated with resins that remove divalent cations lost their ability to aggregate in 30 min. More than about 0.26 mM Mg2+ partially protected the platelets. Unlike aggregation, ADP-induced shape change, clot retraction caused by thrombin or ADP plus reptilase, and thrombin-induced 14C-serotonin release were not inhibited after incubation. Aggregability was not restored by prolonged incubation with CaCl2, adding normal plasma, or washing the platelets. Its loss was temperature and pH dependent, occurring in 2 min at 43 degrees C but not in 7 min at 30 degrees C, and at pH 7.8 but much less at pH 7.2. The defect was not associated with an increase in platelet cyclic AMP, a decrease in metabolic ATP, or the presence of free ADP.