Release and synthesis of acetylcholine at ectopic neuromuscular junctions in the rat

1. The ability of axons in the superficial fibular nerve to synthesize and release acetylcholine (ACh) has been studied before and after the formation of ectopic neuromuscular junctions (NMJs) with denervated soleus muscles of adult rats. 2. The central end of the severed fibular nerve was transplanted to the surface of the soleus muscle. After 3.5-5 weeks the soleus muscle was denervated in one group of rats by cutting the tibial nerve, allowing the formation of functional ectopic NMJs within a few days. In other rats the tibial nerve remained intact, preventing the formation of functional ectopic NMJs. 3. A month later the content of ACh, the levels of activity of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE), and the amount of ACh released by depolarization by exposure to 50 mM KCl were measured in segments of isolated muscles that (i) contained normal or ectopic NMJs, (ii) were free of nerve or (iii) contained nerve that had not made NMJs. 4. Regions of muscles with ectopic nerve growth in which new NMJs had not formed contained substantial amounts of ACh and ChAT but no AChE. No detectable release of ACh could be evoked from these regions. 5. In muscles in which ectopic NMJs had formed after cutting the tibial nerve, the amounts of ACh and ChAT were about one-fifth of those in the regions of innervation of control muscles. ACh release could be evoked from the region of ectopic nerve growth in amounts nearly as great as those released from NMJs in normal and contralateral control muscles. 6. We conclude that the ability of the terminal parts of mature motor axons to synthesize and store ACh is largely independent of functional contact with muscle fibres. By contrast, the ability to release ACh in substantial amounts only develops when NMJs are formed. The possible significance of this situation for the development of synapses is discussed.