Absence of (-) [3H]desmethoxyverapamil binding sites on human platelets and lack of evidence for voltage-dependent calcium channels.
The two major pathways for Ca2+ entry into cells are potential-sensitive channels and receptor-operated channels. The main object of this investigation was to identify which mechanism regulates Ca2+ entry into human platelets. Platelet stimulation with thrombin, adenosine diphosphate, platelet activating factor and arachidonic acid resulted in a concentration-dependent 2.5-3-fold increase in cytoplasmic free calcium concentration over the basal levels (140 +/- 32 nM or 104 +/- 21 respectively) as measured with the fluorescent dyes Quin-2 and Fura-2. Adrenaline and collagen had no effect in promoting intracellular Ca2+ increase as measured with Quin-2 and little effect when measured with Fura-2. Incubation of Quin-2-loaded platelets with the calcium antagonists verapamil and diltiazem, which are known to inhibit Ca2+ entry from voltage-gated channels in many types of cells, over the concentration range 10(-8) - 10(-4) M did not alter significantly either the resting or the cytoplasmic free Ca2+ after stimulation of platelets by several agonists. Moreover, the calcium antagonists exhibited little or no effect on aggregation and 5-hydroxytryptamine secretion induced by platelet activating factor, adenosine diphosphate, collagen or arachidonic acid in whole blood, platelet-rich plasma or washed platelets when employed at concentration ranges as above. Similar results were obtained in washed thrombin-stimulated platelets. High doses of verapamil (but not diltiazem) inhibited platelet aggregation and secretion in response to adrenaline. Direct radioligand binding studies with (-)[3H]desmethoxyverapamil showed that platelet membranes have no receptors for this drug, suggesting that Ca2+ entry occurs in human platelets via a pathway different from potential-sensitive Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)
D000328:Adult; D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000494:pharmacology; D001777:Blood Coagulation / Q000187:drug effects; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels; D004594:Electrophysiology; D006801:Humans; D066298:In Vitro Techniques; D007473:Ion Channels / Q000187:drug effects*; D010974:Platelet Aggregation / Q000187:drug effects; D051381:Rats; D011978:Receptors, Nicotinic / Q000097:blood*; D012701:Serotonin / Q000378:metabolism; D014700:Verapamil / Q000494:pharmacology*
D001095:Arachidonic Acids; D015220:Calcium Channels; D007473:Ion Channels; D011978:Receptors, Nicotinic; D016718:Arachidonic Acid; D012701:Serotonin; D014700:Verapamil; D002118:Calcium