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PubMed1998

Sexual dimorphism in broiler chick embryos and embryonic muscle development in late incubation.

Studies were conducted to compare the weights and the characteristics of the Pectoralis superficialis, Semimembranosus, and Gastrocnemius of male and female chicken embryos at 16 and 20 d of incubation. Male embryos were significantly heavier than females at 16 d of incubation, but not at 20 d. The cross-sectional area of myofibers in the P. superficialis of 20-d-old female embryos was greater than that of males and the area of Semimembranosus myofibers was greater at 16 and 20 d. The Semimembranosus apparent myofiber number of 16-d-old male embryos was greater than of females (P = 0.002). There was a significant increase in apparent myofiber number in this muscle between 16 and 20 d of incubation but there was no sex difference at 20 d. The number of Type I myofibers per 30,000 microm2 area of the Gastrocnemius of 20-d-old male embryos was greater than that in females (P = 0.06). There were no significant sex differences in the protein concentration, DNA concentration, or protein:DNA ratio in the muscles at either age when data were pooled across the whole population. There were significant differences in these characteristics between sire families and in some sire families significant sex differences were noted. The protein:DNA ratio of the muscles increased between 16 and 20 d of incubation and the mitotic index decreased. These data suggest that the muscles of male embryos have more but smaller, myofibers than females, which may be responsible for the sex difference in embryo weight and provide the framework for the greater posthatching muscle growth.

Animals↗
PubMed1985

Enzyme histochemical studies in an ontogeny study of muscle development in Ossabaw and decapitated fetuses: cellular reactions.

Fetuses were decapitated in one uterine horn in each of 14 sows at 45 d of gestation. Control (C) and decapitated (D) fetuses were removed by Caesarean section from three sows at 65 d of gestation (total of 10 D and 10 C fetuses), two sows at 85 d (six D and six C fetuses) and nine sows at 110 d (nine C and nine D fetuses) of gestation (Exp. 1). In Exp. 2, four to six fetuses were removed from each of two Ossabaw (O) gilts and three crossbred (C, Landrace X Yorkshire) gilts at 70 d of gestation, from three C and O gilts at 90 d of gestation and from three C and two O gilts at 110 d of gestation. In Exp. 1, one semitendinosis muscle was removed for histochemistry, whereas the contralateral muscle was removed and weighed. A medial portion of biceps femoris muscle was removed and used for histochemistry in Exp. 2. In both experiments, transverse sections (cryostat) of muscle were stained for lipid, glycogen (PAS) and the following enzymes: acid ATPase, NADH-TR, NADPH-TR, malate dehydrogenase (NAD- and NADP-dependent reactions; MDH), succinate dehydrogenase (SDH), alpha-glycerol phosphate dehydrogenase (with and without NAD; alpha-GPDH), isocitrate dehydrogenase (NAD dependent; ICDH), esterase, lipoprotein lipase and lipase. In Exp. 1, body and muscle weights of the two groups were not significantly different (P greater than .05) at 65 d of gestation, whereas D fetuses were smaller and had lighter weight muscles (P less than .05) at 85 d of gestation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

SV40 T antigen inhibits expression of MyoD and myogenin, up-regulates Myf-5, but does not affect early expression of desmin or alpha 7 integrin during muscle development.

The terminal stage of myogenic development is marked by the cessation of replication, fusion, and expression of genes which encode the myofibrillar proteins. Prior to terminal differentiation one or more stages of myogenic development take place. Expression of alpha 7 integrin and desmin have been used as markers for these earlier stages of myogenesis. Both proteins are expressed in replicating secondary myoblasts prior to terminal differentiation and when these cells differentiate further, the expression of the alpha 7 integrin and desmin genes is up-regulated. To determine whether the stages of myogenesis which precede terminal development and the factors which regulate them are distinct, the expression of alpha 7 integrin and desmin was assayed in a variety of myogenic cell lines in which terminal differentiation was inhibited. L8E63 and C2 myoblasts in which terminal differentiation was inhibited by SV40 large T antigen, adenovirus E1A protein, or ras and an L6 mutant whose terminal differentiation is sensitive to alpha-amanitin were studied. In all cases, when terminal myogenic differentiation is inhibited the basal levels of desmin and alpha 7 expression are not altered. Under these same conditions expression of the myogenic regulatory genes myogenin and MyoD also were inhibited whereas Myf-5 persisted. These results indicate that expression of the early myogenic phenotype and terminal differentiation are discrete and independent stages of myogenesis and that different transcription factors likely regulate the expression of each stage. In contrast with myoblasts in cultures of newborn rat hindlimb cells and the C2 cell line, myogenic cells derived from C3H10T1/2 cells by treatment with 5-azacytidine or by transfection with MyoD, Myf-5, or MRF4 do not express desmin as replicating myoblasts but do so upon terminal differentiation. This indicates that in vitro, terminal differentiation can proceed in the absence of the phenotypes that normally develop earlier and that the conversion of 10T1/2 cells to myogenic cells can bypass developmental stages which normally occur in vivo. These results are discussed in the context of a model of the myogenic lineage that is based on the expression of desmin.

Animals↗
PubMed1989

Effects of strenuous maternal exercise on fetal organ weights and skeletal muscle development in rats.

The purpose of the present study was to observe the effects of strenuous maternal aerobic exercise throughout gestation on fetal outcome in the rat. The strenuous exercise intensity consisted of a treadmill speed of 30 m.min-1 on a 10 degrees incline, for 120 min.day-1, 5 days.week-1. The rats were conditioned to run on a motor-driven treadmill by following a progressive two-week exercise program, so that by the end of the two weeks the rats were capable of running comfortably at this strenuous intensity in the non-pregnant state. Following the two-week running programme, the rats were paired by weight and randomly assigned to either a pregnant group that continued the running program throughout gestation (pregnant runner), or a pregnant group that did not continue the running program throughout pregnancy (pregnant control). At birth the neonates born to the pregnant running group did not differ in average neonatal body weight values, number per litter or total litter weight values when compared to controls, nor were superficial gross abnormalities observed in neonates born to the pregnant control or pregnant running groups. The strenuous maternal exercise intensity did not alter neonatal organ weight values (brain, heart, liver, lung, kidney), nor neonatal skeletal muscle (gastrocnemius, sternomastoid, diaphragm) when compared to control values. It is suggested that maternal exercise of this intensity throughout gestation does not affect fetal outcome in the rat, and may be due to the animals accustomization to the strenuous exercise protocol prior to pregnancy.

Animals↗
PubMed1996

Muscle development and attachment to the epidermis is accompanied by expression of beta 3 and beta 1 tubulin isotypes, respectively.

In Drosophila beta tubulins are encoded by a small gene family whose members are differentially expressed in a highly cell and tissue specific manner. Here we focus on the expression of the beta 3 tubulin isotype during mesoderm differentiation and beta 1 tubulin expression in the apodemes during embryonic development. The beta 3 tubulin isotype is first detectable at the extended germband stage shortly before the separation of somatic and visceral derivatives. Comparing the distribution of the beta 3 mRNA and the beta 3 isotype shows that the transcription of the beta 3 tubulin gene is cell type specifically repressed during differentiation of individual mesodermal derivatives, from which the dorsal vessel remains transcriptionally active until shortly before hatching. In contrast the beta 3 tubulin protein is detectable in all mesodermal derivatives. The beta 3 tubulin is an excellent marker to study mesoderm differentiation on a regulatory and cellular level using both genetics and molecular biology. In the visceral mesoderm, the expression of the beta 3 tubulin gene is regulated by homeotic gene products, while other transactivators regulate expression in the dorsal vessel and the body wall musculature. In the somatic mesoderm, the beta 3 tubulin allows to visualize myotube formation and insertion into the epidermis. This contact to the epidermal attachment sites (apodemes) induces beta 1 tubulin expression, as can be seen in double staining experiments. We determined a 14bp cis-regulatory enhancer element guiding expression of the beta 1 tubulin gene in these attachment sites. Using the beta 1 and beta 3 tubulin isotypes as markers we started to isolate mutants which are disturbed in muscle formation.

Animals↗
PubMed1995

Spontaneous opening of the acetylcholine receptor channel in developing muscle cells from normal and dystrophic mice.

Single-channel activity was recorded from cell-attached patches on skeletal muscle cells isolated from wild-type mice and from mice carrying the dy or mdx mutations. Spontaneous openings of the nicotinic acetylcholine receptor channel (nAChR) were detected in virtually all recordings from either dy/dy or dy/+ myotubes, but only infrequently from wild-type or mdx myotubes. Spontaneous openings were also present in most recordings from undifferentiated myoblasts from all of the mouse strains studied. The biophysical properties of the spontaneous activity were similar to those of the embryonic form of the nAChR in the presence of acetylcholine (ACh). Examination of the single-channel currents evoked by low concentrations of ACh showed a reduced sensitivity to the agonist in the dystrophic dy and mdx myotubes, but not in wild-type myotubes. The results suggest that alterations in nAChR function are associated with the pathogenesis of muscular dystrophy in the dy mouse.

Acetylcholine↗
PubMed1987

Calmodulin levels in developing muscle tissues and primary cultures of normal and dystrophic (UM-X7.1) hamsters.

Calmodulin levels have been assessed in whole muscle and primary culture extracts in order to examine the relationship between calmodulin and the accumulation of calcium in dystrophic hamster muscle tissues. Significant decreases in both normal and dystrophic skeletal muscle, tongue, and heart calmodulin levels were observed between 2 and 12 weeks of age. Dystrophic values, however, tended to be somewhat higher than normal, especially in 12-week-old skeletal muscle total and soluble extracts (normal 29.7 and 0.6 and dystrophic 117.0 and 3.1 micrograms/g wet weight, respectively). No significant differences were observed in dystrophic myoblast (total 2.22-2.78, soluble 2.85-3.26 micrograms/mg protein) or fibroblast (total 2.64-2.94, soluble 2.54-3.60 micrograms/mg protein) calmodulin levels, except for a significant decrease in dystrophic fibroblast levels (total 1.97, soluble 2.18 micrograms/mg protein) at 7 days in culture. Elevated calmodulin levels in dystrophic muscle are discussed in terms of increases in intracellular Ca2+ concentrations and immature regenerating fibers.

Animals↗
PubMed1979

Alterations in lipid metabolism of developing muscle cells in culture.

Embryonic chick muscle cells in the infused (myoblast) and fused (myotube) states differentially incorporate [3H]palmitic acid into their cellular lipids. Myoblasts incorporated [3H]palmitic acid 6--7-fold greater than myotubes in the triglyceride fraction. The endogenous triglyceride levels, however, were similar for the two developmental stages indicating increased triglyceride turnover in the myoblast compared to that of the myotube. Myotubes showed an increased incorporation of [3H]palmitic acid into sphingomyelin compared to myoblasts. Myoblasts and myotubes had a similar phospholipid composition. The data indicate that fusion is characterized by a decrease in triglyceride turnover possibly reflecting a shift from oxidative to glycolytic metabolism to coincide with the fusion process.

Animals↗
PubMed1979

Effects of innervation by ciliary ganglia on developing muscle in vitro.

Electrophysiological and pharmacological properties of neuromuscular junctions formed in tissue culture between chick ciliary ganglia and chick skeletal muscle cells have been studied. Functional neuromuscular junctions are formed already within 24 h. No functional acetylcholine (ACh) esterase is present at these end-plates. The neurites conduct action potentials to the neuromuscular junctions, where EPPs are generated. Tetrodotoxin (TTX) blocks this nerve conduction but in the presence of TTX MEPP-like potentials remain whose amplitudes are lowered when the Mg2+/Ca2+ ratio in the medium is raised. It is speculated that these large TTX-resistant potentials are multiquantal. The relatively high resting membrane potential in non-innervated muscle fibers was not changed by innervation. ACh-sensitivity was determined by iontophoretical application of ACh to the myotubes. Non-innervated myotubes exhibited an evenly distributed ACh-sensitivity. Local differences in ACh-sensitivity were always gradual and never exceeded a factor of 3. Innervation did not alter the overall ACh-sensitivity, but on functionally innervated muscle cells loci hypersensitive to ACh were found. Hypersensitivity was located within sharply defined areas. Apparently the parasympathetic neurons of the ciliary ganglion are able to form functional neuromuscular junctions with skeletal muscle cells in tissue culture and to induce the formation of regions of high ACh-sensitivity.

Animals↗
PubMed1995

Subnanosecond polarized fluorescence photobleaching: rotational diffusion of acetylcholine receptors on developing muscle cells.

Polarized fluorescence recovery after photobleaching (PFRAP) is a technique for measuring the rate of rotational motion of biomolecules on living, nondeoxygenated cells with characteristic times previously ranging from milliseconds to many seconds. Although very broad, that time range excludes the possibility of quantitatively observing freely rotating membrane protein monomers that typically should have a characteristic decay time of only several microseconds. This report describes an extension of the PFRAP technique to a much shorter time scale. With this new system, PFRAP experiments can be conducted with sample time as short as 0.4 microseconds and detection of possible characteristic times of less than 2 microseconds. The system is tested on rhodamine-alpha-bungarotoxin-labeled acetylcholine receptors (AChRs) on myotubes grown in primary cultures of embryonic rat muscle, in both endogenously clustered and nonclustered regions of AChR distribution. It is found that approximately 40% of the AChRs in nonclustered regions undergoes rotational diffusion fast enough to possibly arise from unrestricted monomer Brownian motion. The AChRs in clusters, on the other hand, are almost immobile. The effects of rat embryonic brain extract (which contains AChR aggregating factors) on the myotube AChR were also examined by the fast PFRAP system. Brain extract is known to abolish the presence of endogenous clusters and to induce the formation of new clusters. It is found here that rotational diffusion of AChR in the extract-induced clusters is as slow as that in endogenous clusters on untreated cells but that rotational diffusion in the nonclustered regions of extract-treated myotubes remains rapid.

Animals↗