title: The inositol trisphosphate receptor antagonist 2-aminoethoxydiphenylborate (2-APB) blocks Ca2+ entry channels in human platelets: cautions for its use in studying Ca2+ influx.
abstract: It has been reported that store-mediated Ca2+ entry (SMCE) in human platelets is likely to be mediated by a secretion-like coupling mechanism. Recently, 2-aminoethoxydiphenylborate (2-APB) has been used in the investigation of SMCE. Here, the mechanism of action of 2-APB is investigated in human platelets. In a Ca2+-free medium (EGTA added), addition of 0.1 U/ml thrombin caused an elevation in [Ca2+]i. Preincubation with 100 microM 2-APB for 170s abolished the release of internal Ca2+. In platelets whose internal Ca2+ stores had been depleted by treatment with 200 nM thapsigargin, addition of extracellular Ca2+ caused an elevation in [Ca2+]i indicative of SMCE. Preincubation with 2-APB decreased SMCE by 95.5+/-1.1%. After activation of SMCE, addition of 2-APB rapidly decreased [Ca2+]i to basal levels; in contrast, the coupling between Trp1 and IP3RII, which has been shown to play an important role in SMCE in platelets, remained intact at the same time points. The rate of decrease of [Ca2+]i and the absence of measurable latency in the effect of 2-APB were comparable to the effects of La3+ (a cation channel blocker). These data suggest that 2-APB may act as a blocker of Ca2+ permeable plasma membrane channels. These data provide further information regarding the mechanism and site of action of 2-APB and highlight the necessity of careful interpretation of work performed using this molecule.
authors.affiliation: Department of Physiology, University of Cambridge, Cambridge, UK.
authors.forename: J M
authors.initials: JM
authors.lastname: Diver
authors.forename: S O
authors.initials: SO
authors.lastname: Sage
authors.forename: J A
authors.initials: JA
authors.lastname: Rosado
mesh_terms: D001692:Biological Transport; D001792:Blood Platelets / Q000378:metabolism*; D015153:Blotting, Western; D001896:Boron Compounds / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects* / Q000378:metabolism; D016257:Fura-2; D006801:Humans; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007811:Lanthanum / Q000378:metabolism; D011233:Precipitin Tests; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors* / Q000378:metabolism; D050052:TRPC Cation Channels; D019284:Thapsigargin / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology
chemical_list: D001896:Boron Compounds; D002121:Calcium Channel Blockers; D015220:Calcium Channels; C506926:ITPR1 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D050052:TRPC Cation Channels; C096351:transient receptor potential cation channel, subfamily C, member 1; D019284:Thapsigargin; D007811:Lanthanum; C109986:2-aminoethoxydiphenyl borate; D013917:Thrombin; D002118:Calcium; D016257:Fura-2