The growth of muscle nerves in relation to the formation of primary myotubes in the developing chick forelimb

Abstract

A study has been made of the development of muscle nerves to primary myotube clusters destined to become the flexor carpi ulnaris (fcu) and flexor digitorum profundus (fdp) muscles in the avian forelimb. Myotubes and nerves were identified by immunofluorescent techniques using antibodies to the heavy and light subunits of myosin and neurofilament, respectively. At stage 24 the main ventral nerve trunk (the brachialis longus inferior nerve; bli n) had entered the limb before the appearance of myotubes in the limb. At stage 25/26 the bli n within the ventral compartment of the forearm had given rise to the interosseus nerve (in n) and the medial-ulnar nerve (m-u n) at the junction of the stylopodium and zeugopodium. The first few myotubes of the fdp and fcu muscles were observed at this level within the ventral premuscle cell mass; however, no nerves projected toward these myotubes from either the in n or the m-u n at this time. At stage 26/27 the fcu and the fdp muscles each consisted of clusters of 20-40 myotubes; each cluster was clearly delineated within the ventral premuscle cell mass. By this time small groups of axons had left the in n and the m-u n to grow into the fdp and fcu myotube clusters, respectively; these axons formed the muscle nerves. At stage 28/29 the number of primary myotubes in the clusters composing the fdp and fcu muscles had greatly increased, as did the size of the muscle nerves; each muscle was still clearly identifiable within the ventral muscle mass. By stage 32 the fdp and fcu muscles had clearly separated and the muscle nerves had divided into several well-spaced branches within each muscle. The present observations show that the main nerve trunks grow into the limb before the formation of myotubes expressing myosin isozymes. When myotubes do form they appear in small clusters at specific sites within the premuscle mass, before muscle nerves appear; a distinct muscle is destined to form from each of these clusters. Muscle nerves first branch from the main limb nerves when the myotube cluster contains more than about ten myotubes.

Developmental relations between sixth nerve motor neurons and their targets in the chick embryo

“…In the case of limb motor axons these early contacts are not obligatorily linked to specific target recognition (Lance-Jones, 1986, 198813;Tosney, 19881, which occurs several days later. Our data for the IVth nerve in the non-somitic head mesoderm complements evidence that nerves contact epaxial muscle precursors while they are still in the myoepithelial phase (Oakley and Tosney, 19931, although primary myocyte formation and segregation of individual muscles occur independant of target matching by motor nerves (Noakes et al, 1986;Fredette and Landmesser, 1991;Ashby et al, 1993). However, both the orientation of elongating motor axons and patterns of muscle segregation are determined primarily by cues resident with limb somatic mesoderm (Lance-Jones and Dias, 1991).…”

Ultrastructural and morphometric analysis of the separation of two thigh muscles in the chick

“…Alternatively, cleavage has also been suggested to result directly from a variety of possible (but yet to be documented) changes in the myogenic cells. For instance, myogenic cells might not differentiate within cleavage zones (Noakes et al, 1986), or might stop or reverse their differentiation (Shellswell, 1977;Shellswell and Wolpert, 1977); or their adhesivity might decrease and result in the sorting out of cells and the separation of muscles (see Shellswell and Wolpert;. To address these possible mechanisms, we systematically sampled populations in the region of muscle separation before, during, and after separation and characterized them ultrastructurally.…”

A quantitative study of the segmental distribution of somitic cells in the developing chick limb bud using laser-scanning confocal microscopy.

“…Embryos were fixed at HH stages 26-27 as described for E4 embryos, Intact wings and trunks were immersed overnight in a 1:jOO dilution of ascites 2H2 monoclonal antibody (Noakes et al, 1986) to myosin light chains. After several washes in PBS, the wings were immersed in a 1:50 dilution of FITC-conjugated sheep anti-mouse IgG (Silenus; Melbourne, Australia) and washed again in PBS.…”

“…Myotubes were identified immunocytochemically in sections of limb with monoclonal antibody 2H2 to myosin light chains (Noakes et al, 1986). Antibody binding was detected with a fluorescein-conjugated anti-mouse antibody.…”

“…The cells aggregate into the dorsal and ventral premuscle masses from HH stage 23 (Hayashi and Ozawa, 1991;Schramm and Solursh, 1990;Newman et al, 1981) and express the myogenic regulatory factor MyoD a short time later (Williams and Ordahl, 1994). Fusion of myoblasts into myotubes and expression of muscle-specific proteins begins around HH stage 26 in the wing ( Noakes et al, 1986).…”