Modes of inhibitory action of 4 beta-phorbol 12-myristate 13-acetate in thrombin-stimulated arachidonic acid release in intact and permeabilized platelets.
The tumor-promoting phorbol ester 4 beta-phorbol 12-myristate 13-acetate (PMA) inhibited thrombin-stimulated arachidonic acid (AA) release in rabbit and human platelets. PMA was effective over the same concentration range that activates protein kinase C in intact rabbit platelets: IC50 vs thrombin = 0.5 nM, greater than 90% inhibition at 10 nM. Suppression of thrombin-stimulated AA release was evident within 5 min of pretreatment with 1 nM PMA. A non-tumor-promoting phorbol ester, 4-O-methyl PMA, showed a very weak ability to inhibit AA release. Thrombin-stimulated serotonin secretion was progressively inhibited by PMA pretreatment in platelets, while PMA was a stimulus for secretion at higher concentrations. 1-(5-Isoquinolinylsulfonyl)-2-methyl-piperazine (H-7), a selective inhibitor of protein kinase C, blocked PMA-induced inhibition of AA release. Furthermore, H-7 enhanced the effect of thrombin on AA release. PMA pretreatment reduced the inhibitory effect of thrombin on forskolin-stimulated cAMP accumulation, but had no effect on nonstimulated cAMP metabolism in the presence of thrombin. PMA did not inhibit AA release caused by A23187 or melittin. In digitonin-permeabilized platelets, thrombin plus guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)-stimulated AA release, but not GTP gamma S- and AIF4(-)-stimulated AA release, was abolished by PMA pretreatment. These results suggest that activation of protein kinase C may exert negative feedback on the receptor-mediated activation of phospholipase A2. A possible uncoupling of thrombin receptor to GTP-binding protein leading to activation of phospholipase A2 by PMA pretreatment is discussed.
Murayama
T
T
Department of Pharmacology, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Kajiyama
Y
Y
Takahashi
A
A
Nomura
Y
Y
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000535:Aluminum / Q000494:pharmacology; D000077410:Aluminum Chloride; D017607:Aluminum Compounds; D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000097:blood*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002463:Cell Membrane Permeability; D002712:Chlorides / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000097:blood; D006801:Humans; D066298:In Vitro Techniques; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D010879:Piperazines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors; D011817:Rabbits; D012701:Serotonin / Q000097:blood; D012969:Sodium Fluoride / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013917:Thrombin / Q000494:pharmacology*
D017607:Aluminum Compounds; D001095:Arachidonic Acids; D002712:Chlorides; D007546:Isoquinolines; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D005576:Colforsin; D016718:Arachidonic Acid; D012701:Serotonin; D000077410:Aluminum Chloride; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D012969:Sodium Fluoride; D000535:Aluminum; D000242:Cyclic AMP; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate