Dual transmembrane signalling mechanisms in eosinophils : evidence for two functionally distinct receptors for platelet-activating factor

The effect of PAF on eosinophil activation was investigated. TxB2 release required a lower concentration of PAF (ED50 = 17.2 nM) whereas for superoxide anion (O2) production doses in excess of 1 μM (ED50 = 31.7 μM) were needed. The PAF-induced O2 release occurred in the absence of increased [Ca2+]i whereas the production of TxB2 paralleled the magnitude of the [Ca2+]i increase. Pretreatment of the eosinophils with pertussis toxin (PTX) reduced both the PAF-induced release of TxB2 and the PAF-induced rise in [Ca2+]i. However, PTX failed to inhibit PAF-induced O2 generation. Experiments with the microsomal (100,000 g) fraction from these cells demonstrated that PTX pretreatment had ADP-ribosylated a 41-kD protein in the membrane, confirming that GTP binding proteins are present in eosinophil membranes. Scatchard plot analysis of radioligand binding to eosinophil membranes indicated the presence or two binding sites with an apparent Kd of 0.33 and 11.5 nM, respectively. The results suggest that two distinct forms of the PAF receptor can be identified in eosinophil membranes.