title: Inhibition of thrombin-induced Ca²⁺ influx in platelets by R59949, an inhibitor of diacylglycerol kinase.
abstract: OBJECTIVE
  The aim of this study was to determine whether diacylglycerol kinase (DGK) is involved in transplasmalemmal Ca²⁺ influx of platelets.
  
  
  METHODS
  Effects of R59949, an inhibitor of diacylglycerol kinase, on intracellular Ca²⁺ concentration ([Ca²⁺](i) ) and mRNA expression of DGK isozymes were investigated using washed human platelet suspensions.
  
  
  RESULTS
  Thrombin-induced increase in [Ca²⁺](i) was significantly inhibited by pretreatment of platelets with R59949, while thapsigargin-induced increase in [Ca²⁺](i) was comparable in platelets with and without R59949 pretreatment. Thapsigargin-induced increase in [Ca²⁺](i) was markedly attenuated in the presence of SKF-96365. In the presence of SKF-96365, thrombin-induced increase in [Ca²⁺](i) was significantly attenuated, and additional treatment with R59949 caused a further decrease in [Ca²⁺](i) . Pretreatment of platelets with 1-butanol significantly attenuated thrombin-induced increase in [Ca²⁺](i) , while thrombin-induced increase in [Ca²⁺](i) was augmented in the presence of propranolol. mRNA expression of DGK-α and DGK-γ, which are known to be inhibited by R59949, in platelets was confirmed by RT-PCR analysis.
  
  
  CONCLUSIONS
  R59949 inhibited a store-depletion-insensitive component of transplasmalemmal Ca²⁺ entry induced by thrombin, while store-operated Ca²⁺ entry was not affected by R59949. The results of this study suggest that phosphatidic acid is involved in thrombin-induced Ca²⁺ influx of platelets.
authors.affiliation: Department of Environmental and Preventive Medicine, Hyogo College of Medicine, Mukogawa-cho, Nishinomiya, Hyogo, Japan.
authors.forename: Mikio
authors.initials: M
authors.lastname: Marumo
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Nakano
authors.forename: Yuji
authors.initials: Y
authors.lastname: Takeda
authors.forename: Kaoru
authors.initials: K
authors.lastname: Goto
authors.forename: Ichiro
authors.initials: I
authors.lastname: Wakabayashi
mesh_terms: D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D000440:Butanols / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D019852:Diacylglycerol Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006490:Hemostatics / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D010712:Phosphatidic Acids / Q000378:metabolism; D010880:Piperidines / Q000494:pharmacology*; D010936:Plant Extracts / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects; D011433:Propranolol / Q000494:pharmacology; D052999:Quinazolinones / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053629:Thapsia / Q000737:chemistry; D019284:Thapsigargin / Q000494:pharmacology*; D013917:Thrombin / Q000494:pharmacology*
chemical_list: D000440:Butanols; D002121:Calcium Channel Blockers; D006490:Hemostatics; D007093:Imidazoles; D010712:Phosphatidic Acids; D010880:Piperidines; D010936:Plant Extracts; D052999:Quinazolinones; D012333:RNA, Messenger; C058544:R 59949; D019284:Thapsigargin; D011433:Propranolol; D019852:Diacylglycerol Kinase; D013917:Thrombin; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D002118:Calcium