Effects of cytochaslasin B and colcemide on myogenic cultures.

Abstract

Muscle cultures treated with cytochalasin B yield mono-and oligonucleated cells of two kinds: (i) arborized, replicating precursor myogenic cells and fibroblasts; and (ii) round, post-mitotic, terminally differentiating myoblasts and myotubes. The arb6rized cells do not bind fluorescein-labeled antibody against myosin, do not contract rhythmically, and do not display hexagonally stacked thick and thin filaments. The round, mononucleated myoblasts and round, oligonucleated myotubes bind the fluorescein-labeled antibody against myosin, contract rhythmically, and display clusters of hexagonallystacked thick and thin filaments. When cytochalasin B is removed and replaced by colcemide, the arborized cells, but not the post-mitotic muscle cells, acquire a radial symmetry and are induced to assemble massive, meandering cables that may occupy over 25% of the cell volume. These tortuous cables are positively birefringent and consist exclusively of enormous numbers of 100-A, intermediate-sized filaments.After 3 days in vitro primary cultures of embryonic chick muscle contain immature, post-mitotic, multinucleated myotubes, post-mitotic mononucleated myoblasts, replicating myogenic cells, replicating fibrogenic cells, and replicating mesenchyme cells (1,2,40). The addition of cytochalasin B to these cultures promptly blocks fusion of myoblasts into myotubes. Cytochalasin B also rapidly blocks locomotion of all replicating cells, induces them to "retract," and gradually transforms them into fully arborized cells (3-6). Cytochalasin B does not arborize immature myotubes, but it does cause them to retract from the clot, and subsequently the myotubes degenerate. Because cytocholasin B blocks, not the initiation, but the completion, of cytokinesis (6, 7), these cultures show an increase in oligonucleated cells with time. Cytocholasin B does not significantly impair either the initiation or continued synthesis of myosin, actin, or tropomyosin, for by day 6 the post-mitotic cells generated in these cultures contract spontaneously (3, 6).The addition of colcemide to 3-day-old muscle cultures results in the metaphase-arrest of many mononucleated myogenic and fibrogenic cells. Colcemide fragments the elongated, immature myotubes into multinucleated myosacs and dismantles their microtubules. The myosacs are rich in "lakes" of i00-A, intermediate-sized filaments, and it has been proposed that there is an inverse correlation between the microtubules in myotubes and the 100-A filaments in myosacs (6,8). Colcemide has no detectable effect on the synthesis of contractile proteins, their organization into hexagonally-stacked thick and thin filaments, or their interaction required for spontaneous contractions (9). We wish to report that muscle cultures exposed to cytochalasin B gen-513 erate large numbers of post-mitotic, mononucleated myoblasts rich in scattered stacks of thick and thin filaments. Although Z. H, and I bands are atypical, these scattered stacks of thick and thin filaments permit the cell to contract spontaneously. W...

Fusion of isolated myoblast plasma membranes. An approach to the mechanism

“…Ca 2 + -induced fusion of isolated plasma membranes was not affected by cytochalasin B at concentrations which have been shown to inhibit myotube formation in the monolayer [28,29]. In a control experiment the solvent Me 2 SO was found to be slightly inhibitory (Table IV).…”

“…Cytochalasin B, a drug interacting with microfilaments, is known to inhibit myotube formation [28,29]. Ca 2 +-induced fusion of isolated plasma membranes, however, was in sensitive to this drug (Table IV).…”

Effect of oncogenic virus on muscle differentiation.

“…The formation of myotubes during the first 3 or 4 days of the primary virus-treated cultures, but not thereafter, merits discussion, particularly regarding changes in the cell surface of myogenic cells concerned with fusion (19,20 (8,20 (14,23,26,29). The block by these agents is readily reversible, and it is unlikely that BrdUrd or a viral transformation product suppresses the individual structural genes for myosin, actin, creatine kinase, and the genes regulating the events required for fusion.…”

“…Normal presumptive myoblasts are obligated to synthesize DNA and pass through a quantal cell cycle to yield post-mitotic myoblasts capable of fusing (19,20,23 (23).…”

Hypertrophy-induced increase of intermediate filaments in vascular smooth muscle.

“…J.) (4,29,33,34,38,55,56) .In normal adult mammalian smooth muscle, intermediate filaments generally surround dense bodies on which actin filaments insert (2) . Intermediate filaments are very numerous in cultured vascular (45-47) and other smooth muscles (11)(12)(13)60).…”