Effects of purified recombinant neural and muscle agrin on skeletal musclefibers in vivo

Aggregation of acetylcholine receptors (AChRs) in muscle fibers by nerve-de rived agrin plays a key role in the formation of neuromuscular junctions. S o far, the effects of agrin on muscle fibers have been studied in culture s ystems, transgenic animals, and in animals injected with agrin-cDNA constru cts. We have applied purified recombinant chick neural and muscle agrin to rat soleus muscle in vivo and obtained the following results. Both neural a nd muscle agrin bind uniformly to the surface of innervated and denervated muscle fibers along their entire length. Neural agrin causes a dose-depende nt appearance of AChR aggregates, which persist greater than or equal to7 w k after a single application. Muscle agrin does not cluster AChRs and at 10 times the concentration of neural agrin does not reduce binding or AChR-ag gregating activity of neural agrin. Electrical muscle activity affects the stability of agrin binding and the number, size, and spatial distribution o f the neural agrin-induced AChR aggregates. Injected agrin is recovered fro m the muscles together with laminin and both proteins coimmunoprecipitate, indicating that agrin, binds to laminin in vivo. Thus, the present approach provides a novel, simple, and efficient method for studying the effects of agrin on muscle under controlled conditions in vivo.