Collagen mediates changes in intracellular calcium in primary mouse megakaryocytes through syk-dependent and -independent pathways

We have characterized changes in [Ca2+](i) in primary mouse megakaryocytes in response to fibrillar collagen and in response to cross-linking of the collagen receptor, the integrin alpha(2)beta(1). The response to collagen was markedly different from that seen to a triple helical collagen-related peptide (CRP), which signals via the tyrosine kinases p59(fyn) and p72(syk). This peptide binds to the collagen receptor glycoprotein VI (GPVI), but not to the integrin alpha(2)beta(1). Collagen elicited a sustained increase in [Ca2+](i) composed primarily of influx of extracellular Ca2+ with some Ca2+ release from internal stores. In contrast to CRP, this response was only partially (similar to 30%) inhibited by the src-family kinase inhibitor PP1 (10 mu mol/L) or by microinjection of the tandem SH2 domains of p72(syk), Collagen also caused an increase in [Ca2+](i) in megakaryocytes deficient in either p59(fyn) or p72(syk), although the response was reduced by approximately 40% in both cases: Cross-linking of the alpha(2) integrin increased [Ca2+](i) in these cells exclusively via Ca2+ influx. This response was reduced by approximately 50% after PP1 pretreatment, but was significantly increased in fyn-deficient megakaryocytes. Collagen therefore increases [Ca2+](i) in mouse megakaryocytes via multiple receptors, including GPVI, which causes Ca2+ mobilization, and alpha(2)beta(1) which stimulates a substantial influx of extracellular Ca2+. (C) 1999 by The American Society of Hematology.