The characteristics of receptors for platelet-activating factor (PAF) on rabbit neutrophils are investigated in this report. The presence of PAF-specific binding to rabbit neutrophils was confirmed using radiolabeled ligand binding assays and a rabbit peritoneal neutrophil membrane preparation. Binding of PAF to the neutrophil membranes was reversible and reached equilibrium within 30 min. Scatchard analysis of PAF-specific binding to the rabbit neutrophil membranes revealed a dissociation constant (K_d) for PAF of 0.41 ± 0.045 nM and a B_max of 0.32 ± 0.11 pmol of PAF receptor/mg of protein. The order of potencies of PAF receptor antagonists to inhibit the binding of ³H-PAF to rabbit peritoneal neutrophil membranes was determined. For the competition assays, 100 μg of neutrophil or platelet membrane protein, 0.18 nM³H-PAF, and varying amounts of PAF antagonist were incubated at room temperature for 1 hr. PAF receptor antagonists tested were ONO-6240, brotizolam, kadsurenone, WEB-2086, L-652,731, BN-52021, CV-3988, triazolam, alprazolam, and verapamil. The orders of potencies of these PAF receptor antagonists were similar for inhibition of ³H-PAF binding to rabbit peritoneal neutrophil and platelet membranes (correlation coefficient, r = 0.97). PAF had a significantly higher affinity for rabbit neutrophil membranes (K_d = 0.41 ± 0.045 nM), as compared with its affinity for rabbit platelet membranes (K_d = 0.87 ± 0.092 nM). In addition, sodium was found to inhibit ³H-PAF specific binding to rabbit platelet membranes and not to affect ³H-PAF binding to neutrophil membranes. These data indicate that, although PAF receptors on rabbit platelets and neutrophils exhibit similar orders of potencies of PAF receptor antagonists to inhibit the binding of ³H-PAF, the disparity in K_d of PAF for the receptors and the effect of NaCl on the binding of ³H-PAF reveal subtle differences between the cell types.