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+],), with an initial transient phase followed by a sustained phase. Reverse transcription polymerase chain reaction (RT-PCR) showed the expression of P2Y(1), P2Y(2) and P2Y(12). The selective P2Y(1) receptor antagonist 2'-deoxy-N-6-methyl-adenosine- 3',5'diphosphate (MRS2179) and the Gi 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 11 (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, beta I, gamma and phospho-CaMKII.These results suggest that the activation of the P2Y(1) receptor triggers two different [Ca2+]i increase pathways, one IP3-dependent and the other kinase-dependent. (c) 2006 Elsevier B.V. All rights reserved.