Abstract
Cellular injury induces a complex series of events that involves Ca^2+^ signaling, cell communication, and migration. One of the first responses following mechanical injury is the propagation of a Ca^2+^ wave (Klepeis et al. [2001] J Cell Sci 114(Pt 23):4185β4195). The wave is generated by the extracellular release of ATP, which also induces phosphorylation of ERK (Yang et al. [2004] J Cell Biochem 91(5):938β950). ATP and other nucleotides, which bind to and activate specific purinergic receptors were used to mimic injury. Our goal was to determine which of the P2Y purinergic receptors are expressed and stimulated in corneal epithelial cells and which signaling pathways are activated leading to changes in cell migration, an event critical for wound closure. In this study, we demonstrated that the P2Y~1~, P2Y~2~, P2Y~4~, P2Y~6~, and P2Y~11~ receptors were present in corneal epithelial cells. A potency profile was determined by Ca^2+^ imaging for nucleotide agonists as follows: ATPββ₯βUTPβ>βADPββ₯βUDP. In contrast, negligible responses were seen for Ξ²,Ξ³βmeATP, a general P2X receptor agonist and adenosine, a P1 receptor agonist. Homologous desensitization of the Ca^2+^ response was observed for the four nucleotides. However, P2Y receptor internalization and degradation was not detected following stimulation with ATP, which is in contrast to EGFR internalization observed in response to EGF. ATP induced cell migration was comparable to that of EGF and was maximal at 1 ΞΌM. Cells exposed to ATP, UTP, ADP, and UDP demonstrated a rapid twofold increase in phosphorylation of paxillin at Y^31^ and Y^118^, however, there was no activation elicited by Ξ²,Ξ³βmeATP or adenosine. Additional studies demonstrated that wound closure was inhibited by reactive blue 2. These results indicate that P2Y receptors play a critical role in the injury repair process. Β© 2004 WileyβLiss, Inc.