Abstract-ATP is co-released in significant quantities with acetylcholine from motor neurons at skeletal neuromuscular junctions (NMJ). However, the role of this neurotransmitter in muscle function remains unclear. The P2X 2 ion channel receptor subunit is expressed during development of the skeletal NMJ, but not in adult muscle fibers, although it is reexpressed during muscle fiber regeneration. Using mice deficient for the P2X 2 receptor subunit for ATP (P2X 2 ؊/؊ ), we demonstrate a role for purinergic signaling in NMJ development. Whereas control NMJs were characterized by precise apposition of pre-synaptic motor nerve terminals and postsynaptic junctional folds rich in acetylcholine receptors (AChRs), NMJs in P2X 2 ؊/؊ mice were disorganized: misapposition of nerve terminals and post-synaptic AChR expression localization was common; the density of post-synaptic junctional folds was reduced; and there was increased end-plate fragmentation. These changes in NMJ structure were associated with muscle fiber atrophy. In addition there was an increase in the proportion of fast type muscle fibers. These findings demonstrate a role for P2X 2 receptor-mediated signaling in NMJ formation and suggest that purinergic signaling may play an as yet largely unrecognized part in synapse formation. © 2007 IBRO. Published by Elsevier Ltd. All rights reserved.Key words: ATP, acetylcholine, knockout, mouse, nerve terminal, synapse.It is well established that ATP is co-released with acetylcholine (ACh) from motor nerve terminals (Redman and Silinsky, 1994;Redman and Silinsky, 1996). While ACh is the transmitter that mediates via nicotinic receptors muscle contraction in mature animals, during early development both ATP, acting via P2X ion channel receptors (Ralevic and Burnstock, 1998) and ACh mediate muscle responses (Kolb and Wakelam, 1983;Henning, 1997;Heilbronn and Eriksson, 1998). P2X 2 receptor subunits are expressed during early postnatal development, when neuromuscular junction (NMJ) maturation and patterning occur, but disappear in the adult (Ryten et al., 2001). Furthermore, this receptor is re-expressed in the later stages of muscle regeneration in the mdx mouse model of muscular dystrophy (Ryten et al., 2004;Jiang et al., 2005). Also direct postjunctional responses to ATP reappear after denervation of chick skeletal muscle (Wells et al., 1995).Electrophysiology, immunohistochemistry and reverse transcriptase polymerase chain reaction have demonstrated the expression of a range of purinoceptors in developing skeletal muscle, including P2X 2 , P2X 5 , P2X 6 , P2Y 1 , P2Y 2 and P2Y 4 (Kolb and Wakelam, 1983;Hume and Thomas, 1988;Thomas et al., 1991;Henning et al., 1992; Meyer et al., 1999a,b;Bo et al., 2000;Ruppelt et al., 2001;Ryten et al., 2001;Cheung et al., 2003). Specific roles in skeletal muscle fiber formation and function have been identified for two of these receptors. While activation of the P2Y 1 receptor has been shown to regulate acetylcholine receptor (AChR) and acetylcholinesterase (AChE) expression at NMJs...
