Cyclic AMP raises intracellular Ca2+ in human megakaryocytes independent of protein kinase A

The immature megakaryoblastic cell line MEG-01 responds to iloprost with an increase in cytosolic Ca2+ and CAMP. The Ca2+ response is almost absent in CHRF-288-11 cells, but CAMP formation is preserved in this more mature megakaryoblastic cell line. Also, in human hematopoietic stem cells, iloprost induces a Ca2+ response and CAMP formation. The Ca2+ response is downregulated during megakaryocytopoiesis, but CAMP formation remains unchanged. The Ca2+ increase may be caused by CAMP-mediated inhibition of Ca2+ sequestration, because it is (1) independent of Ca2+ entry; (2) mimicked by forskolin. an activator of adenylyl cyclase, and isobutylmethylxanthine, an inhibitor of phosphodiesterases; and (3) preserved in the presence of inhibitors of protein kinase A and inositol-1,4,5-triphosphate receptors. The small GTPase Rapt has been implicated in the control of Ca2+ sequestration. Indeed, Rapt activation parallels the iloprost- and forskolin-induced Ca2+ increase and is unaffected by the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N',-tetraacetic acid-AM. These findings reveal a novel mechanism for elevating cytosolic Ca2+ by CAMP, possibly via GTP-Rap1.