First Neuromuscular Contact Correlates with Onset of Primary Myogenesis in Rat and Mouse Limb Muscles

Abstract

Skeletal muscle development has been the focus of intensive study for many decades. Recent advances in genetic manipulation of the mouse have increased our understanding of the cell signalling involved in the development of muscle progenitors which give rise to adult skeletal muscles and their stem cell populations. However, the influence of a vital tissue type – the peripheral nerve—has largely been ignored since its earliest descriptions. Here we carefully describe the timing in which myogenic progenitors expressing Pax3 and Pax7 (the earliest markers of myogenic cells) enter the limb buds of rat and mouse embryos, as well as the spatiotemporal relationship between these progenitors and the ingrowing peripheral nerve. We show that progenitors expressing Pax3 enter the limb bud one full day ahead of the first neurites and that Pax7-expressing progenitors (associated with secondary myogenesis in the limb) are first seen in the limb bud at the time of nerve entry and in close proximity to the nerve. The initial entry of the nerve also coincides with the first expression of myosin heavy chain showing that the first contact between nerves and myogenic cells correlates with the onset of myogenic differentiation. Furthermore, as the nerve grows into the limb, Pax3 expression is progressively replaced by Pax7 expression in myogenic progenitors. These findings indicate that the ingrowing nerve enters the limb presumptive muscle masses earlier than what was generally described and raises the possibility that nerve may influence the differentiation of muscle progenitors in rodent limbs.

Making muscle: skeletal myogenesisin vivoandin vitro

“…Fiber type specificity is controlled in part by the transcription factors Six and Eya, which promote fast-type fiber diversity (Grifone et al, 2004;Richard et al, 2011). Fiber type also depends on neural input, which is established during fetal stages -as early as E14.5 in mouse limb (Hurren et al, 2015;Martin, 1990). Thus, the final fiber type of a given muscle is the product of its developmental history, its innervation and, during postnatal life, the physiological demands placed upon it (Schiaffino and Reggiani, 2011).…”

Evolution of the muscular system in tetrapod limbs

“…The timing of the first contact between nerve axons and developing muscles varies among tetrapods. In mammals and anurans, nerve axons enter the limb premuscle masses almost concurrent with premuscle mass formation [ 254 – 256 ], while in birds the axons remain at the plexus region prior to the onset of primary myotube formation [ 257 , 258 ]. Considering this interspecific difference, interaction between the nerve axon and developing muscle may not be required for the morphogenesis of limb muscles before the primary myotube formation.…”

Altered mitochondrial bioenergetics are responsible for the delay in Wallerian degeneration observed in neonatal mice

“…The morphological events occurring in the first postnatal month in the mouse have been extensively characterized, but the lack of available tissue makes it very difficult to know if and when these morphological changes occur into humans. In mice, axons of the ventral ramus appear to grow out from the spinal cord at around E12.5, reach the dorsal and ventral muscle masses at around E13.5, and initial nerve muscle contacts are formed at around E14.5 (Hurren et al, 2015). Acetylcholine receptor clusters are not observed until around E14, becoming numerous at around E15 (Hurren et al, 2015).…”

“…In mice, axons of the ventral ramus appear to grow out from the spinal cord at around E12.5, reach the dorsal and ventral muscle masses at around E13.5, and initial nerve muscle contacts are formed at around E14.5 (Hurren et al, 2015). Acetylcholine receptor clusters are not observed until around E14, becoming numerous at around E15 (Hurren et al, 2015). This suggests formation of neuromuscular contacts occurs at around 60 to 70% of the way through gestation.…”