Development of platelet inhibition by cAMP during megakaryocytopoiesis.

Prostacyclin is a potent inhibitor of agonist-induced Ca2+ increases in platelets, but in the megakaryocytic cell line MEG-01 this inhibition is absent. Using human megakaryocytic cell lines representing different stages in megakaryocyte (Mk) maturation as well as stem cells and immature and mature megakaryocytes, we show that the inhibition by prostacyclin develops at a late maturation stage shortly before platelets are formed. This late appearance is not caused by insufficient cAMP formation or absent protein kinase A (PKA) activity in immature cells. Instead, the appearance of Ca2+ inhibition by prostacyclin is accompanied by a sharp increase in the expression of the catalytic subunit of PKA (PKA-C) but not by changes in the expression of the PKA-regulatory subunits Ialpha/beta, IIalpha, and IIbeta. Overexpression of PKA-C in the megakaryocytic cell line CHRF-288-11 potentiates the Ca2+ inhibition by prostacyclin. Thus, up-regulation of PKA-C appears to be a key step in the development of Ca2+ inhibition by prostacyclin in platelets.