The neuromuscular junction is the chemical synapse between motor neurons and skeletal muscle fibers. It is designed to reliably convert the action potential from the presynaptic motor neuron into the contraction of the postsynaptic muscle fiber. Diseases that affect the neuromuscular junction may cause failure of this conversion and result in loss of ambulation and respiration. The loss of motor input also causes muscle wasting as muscle mass is constantly adapted to contractile needs by the balancing of protein synthesis and protein degradation. Finally, neuromuscular activity and muscle mass have a major impact on metabolic properties of the organisms. This review discusses the mechanisms involved in the development and maintenance of the neuromuscular junction, the consequences of and the mechanisms involved in its dysfunction, and its role in maintaining muscle mass during aging. As life expectancy is increasing, loss of muscle mass during aging, called sarcopenia, has emerged as a field of high medical need. Interestingly, aging is also accompanied by structural changes at the neuromuscular junction, suggesting that the mechanisms involved in neuromuscular junction maintenance might be disturbed during aging. In addition, there is now evidence that behavioral paradigms and signaling pathways that are involved in longevity also affect neuromuscular junction stability and sarcopenia.

Myogenin and MyoD are proteins that bind to the regulatory regions of a battery of skeletal muscle genes and can activate their transcription during muscle differentiation. We have recently found that both proteins interact with the enhancer of the nicotinic acetylcholine receptor (nAChR) a subunit, a gene that is regulated by innervation. This observation prompted us to study if myogenin and MyoD transcript levels are also regulated by skeletal muscle innervation. Using Northern blot analysis, we found that MyoD and myogenin mRNA levels begin to decline at embryonic day 17 and attain adult levels in muscle of newborn and 3-week-old mice, respectively. In contrast, nAChR mRNAs are highest in newborn and 1-week-old mouse muscle and decline thereafter to reach adult levels in 3-week-old mice. To determine if the downregulation of myogenin and MyoD mRNA levels during development is due to innervation, we quantitated message levels in adult calf muscles after denervation. We found that in denervated muscle myogenin and MyoD mRNAs reach levels that are approximately 40-and 15-fold higher than those found in innervated muscle. Myogenin mRNAs begin to accumulate rapidly between 8 and 16 hr after denervation, and MyoD transcripts levels begin to increase sharply between 16 hr and 1 day after denervation. The increases in myogenin and MyoD mRNA levels precede the rapid accumulation of nAChR a-subunit transcripts; receptor mRNAs begin to accumulate significantiy after 1 day of denervation. The effects of denervation are specific because skeletal a-actin mRNA levels are not affected by denervation. In addition, we found that the repression of myogenin and MyoD expression by innervation is due, at least in part, to "electrical activity." Direct stimulation of soleus muscle with extracellular electrodes repressed the increase of myogenin and MyoD transcripts after denervation by 4-to 3-fold, respectively. In view of these results, it is interesting to speculate that myogenin and/or MyoD may regulate a repertoire ofskeletal muscle genes that are down-regulated by electrical activity.Development of skeletal muscle cells is characterized by a series ofevents that include commitment, differentiation, and maturation. Myoblasts arise from the commitment of pluripotential mesodermal cells to the myogenic lineage. The myoblasts proliferate and later differentiate and fuse to form multinucleated embryonic myotubes. Differentiation is characterized by the transcriptional activation of a battery of muscle-specific genes coding for metabolic enzymes, contractile proteins, ion channels, and neurotransmitter receptors (1-4

. To elucidate the nature of signals that control the level and spatial distribution of mRNAs encoding acetylcholine receptor (AChR), a-, ß-, y-, S-and e-subunits in muscle fibers chronic paralysis was induced in rat leg muscles either by surgical denervation or by different neurotoxins that cause disuse of the muscle or selectively block neuromuscular transmission pre-or postsynaptically and cause an increase of AChRs in muscle membrane. After paralysis, the levels and the spatial distributions of the different subunitspecific mRNAs change discoordinately and seem to follow one of three different patterns depending on the subunit mRNA examined . The level of e-subunit mRNA and its accumulation at the end-plate are largely independent on the presence of the nerve or electrical muscle activity. In contrast, the y-subunit mRNA level is tightly coupled to innervation . It is undetectable or low in innervated normally active muscle and in innervated but disused muscle, whereas it is abundant along T HE nicotinic acetylcholine receptor (AChR)' of mammalian skeletal muscle is a heterooligomeric membrane protein composed of a-, ß-, S-, and either -or y e-subunits Witzemann et al ., 1990) . Functionally, the two AChR subtypes (termed AChRy and AChRe, respectively) differ in their gating and ion conductance properties and their density in the sarcolemmal membrane changes during development . Before innervation, the fetal AChR subtype containing the y-subunit is distributed over the entire surface of the muscle fiber and accumulates at the end-plate as the nerve contacts the muscle fiber. The adult AChR subtype containing the e-subunit appears only postnatally and is predominantly localized at the end-plate. Concomitantly with the switch of the AChR subtypes in the end-plate, the fetal AChRs disappear from the extrajunctional membrane. When the muscle is surgically denervated the fetal AChRs are expressed again along the whole fiber length and disappear upon reinnervation . Thus the abun-1. Abbreviations used in this paper: AChR, acetylcholine receptor ; ci-BuTX, a-bungarotoxin ; BuTX, botulinum toxin; m.e.p.c ., miniature end-plate current; m.e .p.p., miniature end-plate potential; TTX, tetrodotoxin .® The Rockefeller University Press, 0021-9525/91/07/125/17 $2 .00 The Journal of Cell Biology, Volume 114, Number 1, July 1991125-141 the whole fiber length in denervated muscle or in muscle in which the neuromuscular contact is intact but the release of transmitter is blocked . The a-, ß-, and S-subunit mRNA levels show a different pattern . Highest amounts are always found at end-plate nuclei irrespective of whether the muscle is innervated, denervated, active, or inactive, whereas in extrasynaptic regions they are tightly controlled by innervation partially through electrical muscle activity. The changes in the levels and distribution of y-and e-subunit-specific mRNAs in toxin-paralyzed muscle correlate well with the spatial appearance of functional fetal and adult AChR channel subtypes along the muscle fiber. Th...

IN mammalian muscle, the subunit composition of the nicotinic acetylcholine receptor (AChR) and the distribution of AChRs along the fibre are developmentally regulated. In fetal muscle, AChRs are distributed over the entire fibre length whereas in adult fibres they are concentrated at the end-plate. We have used in situ hybridization techniques to measure the development of the synaptic localization of the messenger RNAs (mRNAs) encoding the alpha-subunit and the epsilon-subunit of the rat muscle AChR. The alpha-subunit is present in both fetal and adult muscle, whereas the epsilon-subunit appears postnatally and specifies the mature AChR subtype. The synaptic localization of alpha-subunit mRNA in adult fibres may arise from the selective down-regulation of constitutively expressed mRNA from extrasynaptic fibre segments. In contrast, epsilon-subunit mRNA appears locally at the site of neuromuscular contact and its accumulation at the end-plate is not dependent on the continued presence of the nerve terminal very early during synapse formation. This suggests that epsilon-subunit mRNA expression is induced locally via a signal which is restricted to the end-plate region and is dependent on the presence of the nerve only during a short period of early neuromuscular contact. Evidently, several mechanisms operate to confine AChR mRNAs to the adult end-plate region, and the levels of alpha-subunit and epsilon-subunit mRNAs depend on these mechanisms to differing degrees.