Axonal Regeneration and Neuronal Function Are Preserved in Motor Neurons Lacking ß-Actin In Vivo

The proper localization of ß-actin mRNA and protein is essential for growth cone guidance and axon elongation in cultured neurons. In addition, decreased levels of ß-actin mRNA and protein have been identified in the growth cones of motor neurons cultured from a mouse model of Spinal Muscular Atrophy (SMA), suggesting that ß-actin loss-of-function at growth cones or pre-synaptic nerve terminals could contribute to the pathogenesis of this disease. However, the role of ß-actin in motor neurons in vivo and its potential relevance to disease has yet to be examined. We therefore generated motor neuron specific ß-actin knock-out mice (Actb-MNsKO) to investigate the function of ß-actin in motor neurons in vivo. Surprisingly, ß-actin was not required for motor neuron viability or neuromuscular junction maintenance. Skeletal muscle from Actb-MNsKO mice showed no histological indication of denervation and did not significantly differ from controls in several measurements of physiologic function. Finally, motor axon regeneration was unimpaired in Actb-MNsKO mice, suggesting that ß-actin is not required for motor neuron function or regeneration in vivo

Neural agrin controls maturation of the excitation-contraction coupling mechanism in human myotubes developing in vitro

The aim of this study was to elucidate the mechanisms responsible for the effects of innervation on the maturation of excitation-contraction coupling apparatus in human skeletal muscle. For this purpose, we compared the establishment of the excitation-contraction coupling mechanism in myotubes differentiated in four different experimental paradigms: 1) aneurally cultured, 2) cocultured with fetal rat spinal cord explants, 3) aneurally cultured in medium conditioned by cocultures, and 4) aneurally cultured in medium supplemented with purified recombinant chick neural agrin. Ca2+ imaging indicated that coculturing human muscle cells with rat spinal cord explants increased the fraction of cells showing a functional excitation-contraction coupling mechanism. The effect of spinal cord explants was mimicked by treatment with medium conditioned by cocultures or by addition of 1 nM of recombinant neural agrin to the medium. The treatment with neural agrin increased the number of human muscle cells in which functional ryanodine receptors (RyRs) and dihydropyridine-sensitive L-type Ca2+ channels were detectable. Our data are consistent with the hypothesis that agrin, released from neurons, controls the maturation of the excitation-contraction coupling mechanism and that this effect is due to modulation of both RyRs and L-type Ca2+ channels. Thus, a novel role for neural agrin in skeletal muscle maturation is proposed

CD24 is expressed by myofiber synaptic nuclei and regulates synaptic transmission

The genes encoding several synaptic proteins, including acetylcholine receptors, acetylcholinesterase, and the muscle-specific kinase, MuSK, are expressed selectively by a small number of myofiber nuclei positioned near the synaptic site. Genetic analysis of mutant mice suggests that additional genes, expressed selectively by synaptic nuclei, might encode muscle-derived retrograde signals that regulate the differentiation of motor axon terminals. To identify candidate retrograde signals, we used a microarray screen to identify genes that are preferentially expressed in the synaptic region of muscle, and we analyzed one such gene, CD24, further. We show that CD24, which encodes a small, variably and highly glycosylated, glycosylphosphatidylinositol (GPI)-linked protein, is expressed preferentially by myofiber synaptic nuclei in embryonic and adult muscle, and that CD24 expression is restricted to the central region of muscle independent of innervation. Moreover, we show that CD24 has a role in presynaptic differentiation, because synaptic transmission is depressed and fails entirely, in a cyclical manner, after repetitive stimulation of motor axons in CD24 mutant mice. These deficits in synaptic transmission, which are accompanied by aberrant stimulus-dependent uptake of AM1–43 from axons, indicate that CD24 is required for normal presynaptic maturation and function. Because CD24 is also expressed in some neurons, additional experiments will be required to determine whether pre- or postsynaptic CD24 mediates these effects on presynaptic development and function