The rat pups were administered intraperitoneally 0.4 mg/kg ABT-491 which was dissolved in saline, either immediately before or after hypoxia.

Platelet activation and platelet–leukocyte interaction were assessed with the use of flow cytometry. Aliquots of blood were added to microcentrifuge tubes containing HEPES-Tyrodes buffer (5 mM HEPES, 137 mM NaCl, 2.7 mM NaHCO3, 0.36 mM NaH2PO4, 2 mM CaCl2, 4 mM MgCl2, and 5 mM glucose, pH 7.4) and fluorochrome-labeled ligands. A phycoerythrin (PE) Cc5 conjugated antibody to CD42 was used as an activation-independent marker of platelets. FITC-conjugated fibrinogen was added to permit assessment of activation of GP IIb-IIIa. When thrombin was used as an agonist, FITC-conjugated PAC-1 was used instead of FITC-fibrinogen, and the peptide GPRP was added to block polymerization of endogenous fibrinogen. A PE-conjugated antibody to CD62 was used to identify platelet surface expression of P-selectin. For assessment of platelet–leukocyte aggregates, a PE-conjugated antibody to CD14 was used to identify monocytes, and a FITC-conjugated antibody to CD61 was used as an activationindependent marker of platelets. For experiments with the receptor antagonists to PAF (ABT-491, 2.5 mkM), to P2Y12 (cangrelor, 10–1000 nM), and to P2Y1 (MRS-2179, 10–1000 mkM), whole blood anticoagulated with CTI (32 mkg/ml) was incubated at room temperature for 15 minutes in the presence of antagonist or control (vehicle) before addition of aliquots of blood to assay tubes.