Elsevier

Neuroscience

Volume 148, Issue 3, 7 September 2007, Pages 700-711
Neuroscience

Developmental neuroscience
Abnormalities in neuromuscular junction structure and skeletal muscle function in mice lacking the P2X2 nucleotide receptor

https://doi.org/10.1016/j.neuroscience.2007.06.050Get rights and content

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 P2X2 ion channel receptor subunit is expressed during development of the skeletal NMJ, but not in adult muscle fibers, although it is re-expressed during muscle fiber regeneration. Using mice deficient for the P2X2 receptor subunit for ATP (P2X2−/−), 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 post-synaptic junctional folds rich in acetylcholine receptors (AChRs), NMJs in P2X2−/− 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 P2X2 receptor-mediated signaling in NMJ formation and suggest that purinergic signaling may play an as yet largely unrecognized part in synapse formation.

Section snippets

Animals

P2X2−/− mice were generated by introducing a deletion encompassing exons 2–11 into the mouse P2X2 gene (see Cockayne et al., 2005 for details).

Immunohistochemistry and histology

Four female wild-type and four P2X2−/− mice were killed by CO2 asphyxiation and death was confirmed by cervical dislocation according to Home Office (UK) regulations covering Schedule 1 procedures. All experiments conformed to the Royal Free and University College Medical School guidelines on the ethical use of animals; experiments were designed to

Results

P2X2−/− mice were fertile with no signs of gross pathology. We did not detect any significant differences in body mass (27.8 g±1.7 g, wild-type; 27.6 g±0.5 g, P2X2−/− mice). While differences in the muscle masses of wild-type and P2X2−/− extensor digitorum longus (10.9±0.7 mg, wild-type; 9.68±0.5 mg, P2X2−/−) and soleus muscles (9.1±0.44 mg, wild-type; 8.2±0.32 mg, P2X2−/−) were suggestive of muscle atrophy, values did not reach significance (P=0.15 for soleus muscle).

Discussion

Although ATP is well recognized as an important neurotransmitter in the CNS and peripheral nervous system mediating fast synaptic signaling (see Burnstock, 2007), the role of purinergic signaling in the formation and maintenance of synapses has not been fully investigated. In this study P2X2 receptor-deficient mice were used to demonstrate a role for ATP signaling in the formation of mature NMJs. Our results show that the normal expression of the P2X2 receptor protein on developing skeletal

Conclusion

In summary, we demonstrate that absence of the P2X2 receptor on skeletal muscle results in significant abnormalities in muscle structure and function. These effects can be explained most simply by proposing a role for the P2X2 receptor in the normal development of the NMJ. It is becoming increasingly clear that the development and maintenance of synapses is a complex process requiring a balance between stabilizing and de-stabilizing factors. A number of other factors have been shown to

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