Activation of P2Y1 receptor triggers two calcium signaling pathways in bone marrow erythroblasts.
In this study, we describe the presence of P2 receptor subtypes and Ca2+ signaling in erythroblasts. ATP and ADP produced a biphasic increase of intracellular Ca2+ concentration ([Ca2+]i), with an initial transient phase followed by a sustained phase. Reverse transcription polymerase chain reaction (RT-PCR) showed the expression of P2Y1, P2Y2 and P2Y12. The selective P2Y1 receptor antagonist 2'-deoxy-N6-methyl-adenosine-3',5'-diphosphate (MRS2179) and the G(i) protein inhibitor pertussis toxin blocked Ca2+ increase. The initial transient [Ca2+]i increase phase was sensitive to the 1,4,5-inositol trisphosphate (IP3) receptor blocker 2-aminoethoxy-diphenylborate (2-APB), while the sustained phase was sensitive to the protein kinase C (PKC) inhibitor 2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimide (GF109203X) and calcium calmodulin kinase II (CaMKII) inhibitor 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN-62). In addition, the PKC activator phorbol-12,13-dibutyrate (PDBu) produced increase of [Ca2+]i. Flow cytometry analysis showed the expression of Ca2+-dependent PKC alpha, betaI, gamma and phospho-CaMKII. These results suggest that the activation of the P2Y1 receptor triggers two different [Ca2+]i increase pathways, one IP3-dependent and the other kinase-dependent.
Department of Biophysics, Universidade Federal de São Paulo, Escola Paulista de Medicina, Rua Botucatu 862, 04023-062, São Paulo, SP, Brazil. jean@biofis.epm.br
Edgar Julian
EJ
Paredes-Gamero
Rogério Bastos
RB
Craveiro
João Bosco
JB
Pesquero
Jerônimo Pereira
JP
França
Maria Etsuko Miamoto
ME
Oshiro
Alice Teixeira
AT
Ferreira
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D001896:Boron Compounds / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D020013:Calcium Signaling; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004900:Erythroblasts / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D007211:Indoles / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000187:drug effects / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D058472:Receptors, Purinergic P2Y1
D001896:Boron Compounds; D015220:Calcium Channels; D007211:Indoles; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008301:Maleimides; C111914:N(6)-methyl-2'-deoxyadenosine 3',5'-diphosphate; C546383:P2ry1 protein, mouse; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D000244:Adenosine Diphosphate; C063302:KN 62; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; C109986:2-aminoethoxydiphenyl borate; D011493:Protein Kinase C; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C070515:bisindolylmaleimide I