Pharmacological properties of purinergic receptors and their effects on proliferation and induction of neuronal differentiation of P19 embryonal carcinoma cells

We have used P19 embryonal carcinoma cells as in vitro model for early neurogenesis to study ionotropic P2X and metabotropic P2Y receptor-induced Ca2+ transients and their participation in induction of proliferation and differentiation. In embryonic P19 cells, P2Y1, P2Y2 and P2X4 receptors or P2X-heteromultimers with similar P2X4 pharmacology were responsible for ATP and ATP analogue-induced Ca2+ transients. In neuronal-differentiated cells, P2Y2, P2Y6, P2X2 and possibly P2X2/P2X6 heteromeric receptors were the major mediators of the elevations in intracellular free calcium concentration [Ca2+]i. We have collected evidence for the involvement of metabotropic purinergic receptors in proliferation induction of undifferentiated and neural progenitor cells by using a BrdU-incorporation assay. ATP-, UTP-, ADP-, 2-MeS-ATP- and ADP-βS-induced proliferation in P19 cells was mediated by P2Y1 and P2Y2 receptors as judged from pharmacological profiles of receptor responses. ATP-provoked acceleration of neuronal differentiation, determined by analysis of nestin and neuron-specific enolase gene and protein expression, also resulted from P2Y1 and P2Y2 receptor activation. Proliferation- and differentiation-induction involved the activation of inositol-trisphosphate sensitive intracellular Ca2+ stores.