Platelet aggregation induced by alpha 2-adrenoceptor and protein kinase C activation. A novel synergism.
Adrenaline or UK 14304 (a specific alpha 2-adrenoceptor agonist) and phorbol ester (phorbol 12,13-dibutyrate; PdBu) or bioactive diacylglycerols (sn-1,2-dioctanoylglycerol; DiC8) synergistically induced platelet aggregation and ATP secretion. The effect on aggregation was more pronounced than the effect on secretion, and it was observed in aspirinized, platelet-rich plasma or suspensions of washed aspirinized platelets containing ADP scavengers. No prior shape change was found. In the presence of adrenaline, DiC8 induced reversible aggregation and PdBu evoked irreversible aggregation that correlated with the different kinetics of DiC8- and PdBu-induced protein kinase C activation. Adrenaline and UK 14304 did not induce or enhance phosphorylation induced by DiC8 or PdBu of myosin light chain (20 kDa), the substrate of protein kinase C (47 kDa), or a 38 kDa protein. Immunoprecipitation studies using a Gcommon alpha antiserum or a Gi alpha antiserum showed that Gi alpha is not phosphorylated after exposure of platelets to PdBu or PdBu plus adrenaline. Adrenaline, PdBu or adrenaline plus PdBu did not cause stimulation of phospholipase C as reflected in production of [32P]phosphatidic acid. Adrenaline caused a small increase of Ca2+ in the platelet cytosol of platelets loaded with Indo-1; this effect was also observed in the absence of extracellular Ca2+. However, under conditions of maximal aggregation induced by adrenaline plus PdBu, no increase of cytosolic Ca2+ was observed. Platelet aggregation induced by PdBu plus adrenaline was not inhibited by a high intracellular concentration of the calcium chelator Quin-2. These experiments indicate that alpha 2-adrenoceptor agonists, known to interact with Gi, and protein kinase C activators synergistically induced platelet aggregation through a novel mechanism. The synergism occurs distally to Gi protein activation and protein kinase C-dependent protein phosphorylation and does not involve phospholipase C activation or Ca2+ mobilization.
Siess
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Cell Biology Department, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Lapetina
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EG
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000097:blood; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology; D000068438:Brimonidine Tartrate; D002118:Calcium / Q000097:blood; D004075:Diglycerides / Q000494:pharmacology; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D004837:Epinephrine / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007163:Immunosorbent Techniques; D037342:Pertussis Toxin; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D011493:Protein Kinase C / Q000097:blood*; D011810:Quinoxalines / Q000494:pharmacology; D011942:Receptors, Adrenergic, alpha / Q000502:physiology*; D010738:Type C Phospholipases / Q000097:blood; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000316:Adrenergic alpha-Agonists; D004075:Diglycerides; D011810:Quinoxalines; D011942:Receptors, Adrenergic, alpha; D010566:Virulence Factors, Bordetella; C044295:1,2-dioctanoylglycerol; D000246:Adenosine Diphosphate Ribose; D015240:Phorbol 12,13-Dibutyrate; D000068438:Brimonidine Tartrate; D000255:Adenosine Triphosphate; D037342:Pertussis Toxin; D011493:Protein Kinase C; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D002118:Calcium; D004837:Epinephrine