Adenosine 5’-triphosphate (ATP) is the fundamental energy source in all living organism. In addition to its vital role in energy production, ATP is also an essential extracellular signaling molecule. ATP is co-stored and co-released with neurotransmitters at central and peripheral neuronal synapses. Previous results indicated that muscular and neuronal P2Y
1 receptors, as well as muscular P2Y
2 receptor, could be activated by synaptic ATP. This activation leads to the up-regulation of post-synaptic gene expression, e.g. acetylcholine receptor (AChR) and acetylcholinesterase (AChE). However, the functional role of P2Y
2 receptor in the synapses is not fully revealed. In this project, primary culture of cortical neurons was employed as a model for studying the functional role of ATP/P2Y
2-sig...[
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Adenosine 5’-triphosphate (ATP) is the fundamental energy source in all living organism. In addition to its vital role in energy production, ATP is also an essential extracellular signaling molecule. ATP is co-stored and co-released with neurotransmitters at central and peripheral neuronal synapses. Previous results indicated that muscular and neuronal P2Y
1 receptors, as well as muscular P2Y
2 receptor, could be activated by synaptic ATP. This activation leads to the up-regulation of post-synaptic gene expression, e.g. acetylcholine receptor (AChR) and acetylcholinesterase (AChE). However, the functional role of P2Y
2 receptor in the synapses is not fully revealed. In this project, primary culture of cortical neurons was employed as a model for studying the functional role of ATP/P2Y
2-signaling. The mRNA and protein of P2Y
2 receptor was shown to be present in both rat cerebrum and cultured cortical neurons. During the differentiation of cultured cortical neurons, the expression of P2Y
2 receptor was significantly increased. From the results of immunocytofluorescence staining, P2Y
2 receptor was partially co-localized with post-synaptic marker, PSD-95, in cultured cortical neurons. More importantly, P2Y
2 receptor could be detected in lipid raft and shown to be co-existed with P2Y
1 receptor and other post-synaptic proteins in there. To demonstrate the molecular interaction within the complex of P2Y
1-P2Y
2-PSD-95, co-immunoprecipitation was applied, and which subsequently revealed that P2Y
2 receptor could interact directly with P2Y
1 receptor but not with PSD-95 in the complex.
In cultured cortical neurons, the activation of P2Y
2 receptor triggered a mitogen-activated protein (MAP) kinase signaling cascade and induced the phosphorylation of extracellular signal-regulated kinase (ERK), which was similar as that in muscles. Moreover, the activation of P2Y
2 receptor was shown to stimulate several cholinergic genes: AChE, choline acetyltransferase (ChAT) and proline-rich membrane anchor (PRiMA) of AChE, as well as the glutamatergic genes: NMDA receptor subunit NR1 and NR2A.
In summary, these results indicated that P2Y
2 receptor was existed in the brain and in neurons, where it was localized at the neuronal synapse and lipid raft: this synaptic localization might be mediated by the interaction of P2Y
1 receptor. Finally, the functional role of P2Y
2 receptor was proven to regulate the synaptic gene expression. This gene regulatory effect might participate in the modulation of synaptic plasticity, which strengthens the trophic role(s) of ATP during the formation and maintenance of neuron to neuron synapses.
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