title: Immunolocalization of P2Y1 and TPalpha receptors in platelets showed a major pool associated with the membranes of alpha -granules and the open canalicular system.
abstract: P2Y(1) and thromboxane-prostanoid-alpha (TPalpha) receptors on platelets belong to the G-protein-coupled 7-transmembrane domain family. They transmit signals for shape change, mobilization of calcium, and platelet aggregation. Immunogold labeling with a monoclonal antibody (MoAb) to the amino-terminal domain of P2Y(1) and a polyclonal antibody to the C-terminal domain of TPalpha revealed that while present at the platelet surface, both receptors were abundantly represented inside the platelet. Specifically, receptors were found in membranes of alpha-granules and elements of the open-canalicular system. A similar organization was found in mature megakaryocytes. Activation of platelets by adenosine diphosphate (ADP) and the thromboxane A(2) (TXA(2)) analog, I-BOP [1S-(1 alpha,2 beta(5Z),3 alpha-(1E,3S)4 alpha)-7-(3-(3- hydroxy-4-(p-iodophenoxy)-1-butenyl)-7-oxabicyclo(2.2.1)hept-2-yl)-5-heptenoic acid], increased the labeling of both P2Y(1) and TPalpha at the surface and in intracellular pools, suggesting that activation resulted in greater antibody accessibility to the receptor. A return to a platelet discoid shape and to basal values of labeling accompanied receptor desensitization. Platelets lacking the P2Y(12) ADP receptor normally expressed P2Y(1) and TPalpha, both before and after activation. Studies with the anti-ligand-induced binding site (anti-LIBS) MoAb, AP-6, confirmed that stored fibrinogen associated with internal pools of alpha(IIb)beta(3) at the start of secretion in a microenvironment containing agonist receptors. Pharmacologic antagonism of ADP or TXA(2) receptors in antithrombotic therapy may need to take into account blockade of internal receptor pools.
authors.affiliation: Centre National de la Recherche Scientifique (CNRS), Hôpital Cardiologique, Pessac, France. paquita.nurden@cnrshl.u-bordeaux2.fr
authors.forename: Paquita
authors.initials: P
authors.lastname: Nurden
authors.forename: Christel
authors.initials: C
authors.lastname: Poujol
authors.forename: Joelle
authors.initials: J
authors.lastname: Winckler
authors.forename: Robert
authors.initials: R
authors.lastname: Combrié
authors.forename: Nathalie
authors.initials: N
authors.lastname: Pousseau
authors.forename: Pamela B
authors.initials: PB
authors.lastname: Conley
authors.forename: Sylviane
authors.initials: S
authors.lastname: Levy-Toledano
authors.forename: Aida
authors.initials: A
authors.lastname: Habib
authors.forename: Alan T
authors.initials: AT
authors.lastname: Nurden
mesh_terms: D000244:Adenosine Diphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000328:Adult; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000648:ultrastructure*; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D003594:Cytoplasmic Granules / Q000648:ultrastructure*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005340:Fibrinogen / Q000032:analysis; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007425:Intracellular Membranes / Q000737:chemistry*; D008533:Megakaryocytes / Q000648:ultrastructure; D008854:Microscopy, Electron; D015539:Platelet Activation / Q000187:drug effects; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000032:analysis; D018048:Receptors, Purinergic P2 / Q000097:blood*; D058472:Receptors, Purinergic P2Y1; D017482:Receptors, Thromboxane / Q000097:blood*; D013873:Thionucleotides / Q000494:pharmacology
chemical_list: D000911:Antibodies, Monoclonal; D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; C546382:P2RY1 protein, human; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D017482:Receptors, Thromboxane; D013873:Thionucleotides; C061750:7-(3-(3-hydroxy-4-(4'-iodophenoxy)-1-butenyl)-7-oxabicyclo(2.2.1)heptan-2-yl)-5-heptenoic acid; C030812:adenosine 5'-O-(2-thiodiphosphate); D000244:Adenosine Diphosphate; D005340:Fibrinogen