Abstract

At the vertebrate neuromuscular junction (nmj), ATP is known to be coreleased with acetylcholine from the synaptic vesicles. We have previously shown that the P2Y₁ receptor is localized at the nmj. Here, we extend the findings to show that another nucleotide receptor, P2Y₂, is also localized there and with P2Y₁ jointly mediates trophic responses to ATP. The P2Y₂ receptor mRNA in rat muscle increased during development and peaked in adulthood. The P2Y₂ receptor protein was shown to become restricted to the nmjs during embryonic development, in chick and in rat. In both rat and chick myotubes, P2Y₁ and P2Y₂ are expressed, increasing with differentiation, but P2Y₄ is absent. The P2Y₂ agonist UTP stimulated there inositol trisphosphate production and phosphorylation of extracellular signal-regulated kinases, in a dose-dependent manner. These UTP-induced responses were insensitive to the P2Y₁-specific antagonist MRS 2179 (2′-deoxy-N⁶-methyl adenosine 3′,5′-diphosphate diammonium salt). In differentiated myotubes, P2Y₂ activation induced expression of acetylcholinesterase (AChE) protein (but not control α-tubulin). This was shown to arise from AChE promoter activation, mediated by activation of the transcription factor Elk-1. Two Elk-1-responsive elements, located in intron-1 of the AChE promoter, were found by mutation to act in this gene activation initiated at the P2Y₂ receptor and also in that initiated at the P2Y₁ receptor. Furthermore, the promoters of different acetylcholine receptor subunits were also stimulated by application of UTP to myotubes. These results indicate that ATP regulates postsynaptic gene expressions via a common pathway triggered by the activation of P2Y₁ and P2Y₂ receptors at the nmjs.