Platelet activating factor stimulates a receptor-coupled membrane GTPase in guinea pig eosinophils.

The platelet activating factor (PAF) receptor of the guinea pig peritoneal eosinophil was characterized by radioligand ([3H]WEB 2086) binding and measurement of PAF-stimulated membrane GTPase activity. Specific binding of [3H]WEB 2086 was rapid, reversible and saturable, with an equilibrium KD of 20.4 nM at 0 degrees C, and was displaced competitively by unlabelled PAF with a KI of 3.26 nM and a pseudo-Hill coefficient significantly less than unity (0.44). The affinity of PAF for these binding sites was reduced by the nonhydrolysable GTP analogue 5'-guanylylimidodiphosphate (GppNHp), suggesting the coupling of PAF receptors to intracellular effectors through a guanine nucleotide-binding protein (G protein). PAF stimulated a membrane-associated GTPase, indicating the formation of a G protein alpha subunit-GTP complex upon agonist occupation of the PAF receptor. The EC50 for PAF stimulation was 25.5 nM and the Hill coefficient was significantly less than unity (0.56), while the response to 1 microM PAF was antagonised by WEB 2086 with an IC50 of 128 nM and a slope factor not significantly different from unity (0.91), suggesting the coupling of multiple classes of PAF receptors to the G protein. The activation of GTPase by PAF was insensitive to inhibition by cholera toxin; basal GTPase activity was increased by pertussis toxin and no further stimulation was attainable with PAF.