THE SPECIFIC BINDING OF THE PLATELET-ACTIVATING-FACTOR (PAF) RECEPTORANTAGONIST WEB-2086 AND THE BENZODIAZEPINE FLUNITRAZEPAM TO RAT HEPATOCYTES

Thieno-triazolodiazepines WEB 2086 and BN 50739 have been described as the potent PAF receptor antagonists. Binding of radidlabeled [H-3]WEB 2086 has been widely employed to characterize PAF receptors in differ ent cells. In a search for a PAF receptor in isolated rat hepatocytes, we discovered that the binding of [H-3]WEB to rat hepatocytes was hig hly specific but had a relatively low affinity with a K-d of 113 nM an d B-max of 0.65 pmol/10(6) cells in freshly isolated cell suspension a nd K-d of 1.65 mu M and B-max of 2.0 pmol/plate in cultured hepatocyte s. No consistent specific binding of [H-3]PAF itself was found in the same cell preparations. The binding of [H-3]flunitrazepam in the prese nce of the peripheral type of benzodiazepine receptor antagonist Ro 5- 4864 was saturated and exhibited a K-i of 3.8 nM and B-max of 3.5 pmol /plate. The central type of benzodiazepine receptor antagonist clonaze pam also competed for the [H-3]flunitrazepam binding, however with a m uch lower affinity. Various antagonists inhibited the binding of [H-3] WEB 2086 with a rank order BN 50739 much greater than Ro 5-4864 greate r than or equal to clonazepam. Interestingly, bicuculline, a specific antagonist of GABA(A) recognition sites, also significantly reduced th e binding of [H-3]WEB 2086. The binding of [H-3]flunitrazepam was inhi bited with a rank potency BN 50739 much greater than WEB 2086. Taken t ogether, these findings suggest that the specific binding of PAF recep tor antagonists WEB 2086 and BN 50739 in rat hepatocytes does not invo lve PAF receptors and occurs via peripheral benzodiazepine and, possib ly GABA(A) receptor sites.