Thromboxane-insensitive dog platelets have impaired activation of phospholipase C due to receptor-linked G protein dysfunction.
Human platelet thromboxane A2/prostaglandin H2 (TXA2/PGH2) receptors are linked to phosphoinositide-specific phospholipase C (PI-PLC) via a G protein tentatively identified as a member of the Gq class. In contrast, platelet thrombin receptors appear to activate PI-PLC via other unidentified G proteins. Platelets from most dogs are TXA2 insensitive (TXA2-); i.e., they do not aggregate irreversibly or secrete although they bind TXA2, but they respond normally to thrombin. In contrast, a minority of dogs have TXA2-sensitive (TXA2+) platelets that are responsive to TXA2. To determine the mechanism responsible for TXA2- platelets, we evaluated receptor activation of PI-PLC. Equilibrium binding of TXA2/PGH2 receptor agonists, [125I]BOP and [3H]U46619, and antagonist, [3H]SQ29,548, revealed comparable high-affinity binding to TXA2-, TXA2+, and human platelets. U46619-induced PI-PLC activation was impaired in TXA2- platelets as evidenced by reduced (a) phosphorylation of the 47-kD substrate of protein kinase C, (b) phosphatidic acid (PA) formation, (c) rise in cytosolic calcium concentration, and (d) inositol 1,4,5 trisphosphate (IP3) formation, while thrombin-induced PI-PLC activation was not impaired. GTPase activity stimulated by U46619, but not by thrombin, was markedly reduced in TXA2- platelets. Antisera to Gq class alpha subunits abolished U46619-induced GTPase activity in TXA2-, TXA2+, and human platelets. Direct G protein stimulation by GTP gamma S yielded significantly less PA and IP3 in TXA2- platelets. Immunotransfer blotting revealed comparable quantities of Gq class alpha-subunits in all three platelet types. Thus, TXA2- dog platelets have impaired PI-PLC activation in response to TXA2/PGH2 receptor agonists secondary to G protein dysfunction, presumably involving a member of the Gq class.
Johnson
G J
GJ
Department of Medicine, Veterans Administration Medical Center, Minneapolis, Minnesota 55417.
Leis
L A
LA
Dunlop
P C
PC
D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D001643:Bridged Bicyclo Compounds / Q000494:pharmacology; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002118:Calcium / Q000378:metabolism; D004285:Dogs; D004789:Enzyme Activation; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D010712:Phosphatidic Acids / Q000378:metabolism; D010766:Phosphorylation; D010974:Platelet Aggregation / Q000187:drug effects; D011450:Prostaglandin Endoperoxides, Synthetic / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism; D017482:Receptors, Thromboxane / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism; D013928:Thromboxane A2 / Q000378:metabolism* / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*
D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; D010712:Phosphatidic Acids; D011450:Prostaglandin Endoperoxides, Synthetic; D017482:Receptors, Thromboxane; C061750:7-(3-(3-hydroxy-4-(4'-iodophenoxy)-1-butenyl)-7-oxabicyclo(2.2.1)heptan-2-yl)-5-heptenoic acid; D012701:Serotonin; D013928:Thromboxane A2; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D015544:Inositol 1,4,5-Trisphosphate; D011494:Protein Kinases; D010738:Type C Phospholipases; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D002118:Calcium