REGENERATING MUSCLE-FIBERS INDUCE DIRECTIONAL SPROUTING FROM NEARBY NERVE-TERMINALS - STUDIES IN LIVING MICE

The principal aim of this work was to better understand how regenerati ng muscle fibers become innervated in adult animals. To induce muscle regeneration, individual identified muscle fibers in a mouse were dama ged with a laser focused through a microscope. The muscle fiber that d egenerated and the muscle fiber that was formed in its place were foll owed by viewing the same site repeatedly over a period of 2 d to 40 we eks. Commonly, the nerve terminal innervating the irradiated muscle fi ber partially retracted during muscle fiber degeneration, and then spr outed to innervate the regenerating muscle fiber at the same site it h ad previously innervated the muscle fiber that was damaged. During the early phase of muscle regeneration we also observed sprouts originati ng from nerve terminals on adjacent muscle fibers. The new nerve growt h was a response to the regenerating muscle fiber rather than to the d egenerated fiber it replaced because repeated damage of the same site every 2-3 d over a 10 d period (to prevent regeneration) did not cause any sprouting. The direction of the sprouts on adjacent muscle fibers showed a bias toward the regenerating muscle fiber, although they avo ided the region occupied by the original nerve terminal. Forty percent of the sprouts managed to reach the regenerated fiber. Nonetheless, b y 11 d after muscle fiber damage, all sprouts had regressed, leaving t he new fiber innervated by the same motor axon that innervated the fib er that was damaged. On the other hand, when the overlying nerve termi nal as well as the muscle fiber was damaged, the sprouts from nearby m uscle fibers were both more numerous and more stable, and in five case s we observed two or more new synaptic junctions on the regenerating f iber originating from different axons. In one case we witnessed a prot racted competition between the original motor axon as it sprouted back and the sprouts from nearby junctions for sole innervation of the reg enerate. Ultimately, the surviving sprouts myelinated and became the p ermanent and exclusive input to the new fiber. These results indicate that regenerating muscle fibers emit a signal that induces directional sprouting from nearby undamaged nerve terminals. Reinnervation of the regenerating muscle fiber by one axon apparently prevents the mainten ance of such neurites. Because the process of muscle regeneration shar es many features in common with myogenesis during embryonic developmen t, it is likely that developing muscle fibers present an analogous sti mulus to ingrowing motor axons. Furthermore, the well-documented sprou ting observed following partial denervation seems similar to the sprou ting observed in this study, arguing that denervated muscle fibers use a mechanism to attract axons similar to that used by newly forming mu scle fibers.