NONSEROTONERGIC H-3 KETANSERIN BINDING-SITES IN HUMAN PLATELETS - CHARACTERISTICS AND INTERACTION WITH CALCIUM-ANTAGONISTS

Here we report the identification of binding sites for 3H-ketanserin in human platelet membranes. At 4°C, 3H-ketanserin binding is saturable (Bmax=0.58 pmol/mg protein), rapid (equilibrium being attained within 20 min) and reversible. The kinetics of the association and dissociation curves are consistent with the existence of a single class of binding sites, as confirmed also by computer-assisted analysis of the saturation curve. Specific binding is increased by Ca2+ and Mg2+. 3H-ketanserin binding is inhibited by serotonin (Ki=48.5 μM), unlableled ketanserin (K1=3-15 nM), as well as by another antiserotonergic drug, methysergide (Ki-32.6 μM). However, other selective 5-HT2 ligands, such as ritanserin, spiperone and cyproheptadine fail to interact with 3H-ketanserin binding. On the contrary, tetrabenzine, a monoamine depleting agent, when preincubated at 30°C, did inhibit the specific binding completely. 3H-ketanserin specific binding is inhibited in a dose-dependent fashion by some calcium blocking agents, with different potencies: verapamil (Ki=2.25 μM), diltiazem (Ki=139 μM) and SIM6080, a new Ca2+-antagonist related to the phenylalkylamines (Ki=5.22 μM). Flunarizine inhibited 3H-ketanserin specific binding only at relatively high concentrations (IC50>100 μM), while nitrendipine did not show any inhibitory effect up to 20 μM. The present evidence indicates that all the sites labeled by 3H-ketanserin at 4°C might be coincident with the monoamino transporter identified in other systems, and that theymight play a role in the modulation of platelet aggregation exerted by some calcium blocking agents.