The metabotropic P2Y4 receptor participates in the commitment to differentiation and cell death of human neuroblastoma SH-SY5Y cells

Extracellular nucleotides exert a variety of biological actions through different subtypes of P2 receptors. Here we characterized in the human neuroblastoma SH-SY5Y cells the simultaneous presence of various P2 receptors, belonging to the P2X ionotropic and P2Y metabotropic (2,4,6,7) and P2Y(1,2,4,6) families. Western blot analysis detected the P2X(1,) but not the P2X(3) and P2Y(12) receptors. We then investigated which biological effects were mediated by the P2Y(4) subtype and its physiological pyrimidine agonist UTP. We found that neuronal differentiation of the SH-SY5Y cells with dibutiryl-cAMP increased the expression of the P2Y(4) protein and that UTP itself was able to positively interfere with neuritogenesis. Moreover, transient transfection and activation of P2Y(4) also facilitated neuritogenesis in SH-SY5Y cells, as detected by morphological phase contrast analysis and confocal examination of neurofilament proteins NFL. This was concurrent with increased transcription of immediate-early genes linked to differentiation such as cdk-5 and NeuroD6, and activity of AP-1 transcription family members such as c-fos, fos-B, and jun-D. Nevertheless, a prolonged activation of the P2Y(4) receptor by UTP also induced cell death, both in naive, differentiated, and P2Y(4)-transfected SH-SY5Y cells, as measured by direct count of intact nuclei and cytofluorimetric analysis of damaged DNA. Taken together, our data indicate that the high expression and activation of the P2Y(4) receptor participates in the neuronal differentiation and commitment to death of SH-SY5Y cells. (C) 2004 Elsevier Inc. All rights reserved.