Formation of sarcomeres in developing myotubes: role of mechanical stretch and contractile activation.
In a series of experiments, cultured myotubes were exposed to passive stretch or pharmacological agents that block contractile activation. Under these experimental conditions, the formation of Z lines and A bands (morphological structures, resulting from the specific structural alignment of sarcomeric proteins, necessary for contraction) was assessed by immunofluorescence. The addition of an antagonist of the voltage-gated Na(+) channels [tetrodotoxin (TTX)] for 2 days in developing rat myotube cultures led to a nearly total absence of Z lines and A bands. When contractile activation was allowed to resume for 2 days, the Z lines and A bands reappeared in a significant way. The appearance of Z lines or A bands could not be inhibited nor facilitated by the application of a uniaxial passive stretch. Electrical stimulation of the cultures increased sarcomere assembly significantly. Antagonists of L-type Ca(2+) channels (verapamil, nifedipine) combined with electrical stimulation led to the absence of Z lines and A bands to the same degree as the TTX treatment. Western blot analysis did not show a major change in the amount of sarcomeric alpha-actinin nor a shift in myosin heavy chain phenotype as a result of a 2-day passive stretch or TTX treatment. Results of experiments suggest that temporal Ca(2+) transients play an important factor in the assembly and maintenance of sarcomeric structures during muscle fiber development.
De Deyne
P G
PG
Department of Orthopedics, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA. pdedeyne@smail.umaryland.edu
D000185:Actinin / Q000032:analysis / Q000276:immunology; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D024510:Muscle Development; D018485:Muscle Fibers, Skeletal / Q000737:chemistry / Q000502:physiology / Q000648:ultrastructure*; D018482:Muscle, Skeletal / Q000166:cytology* / Q000254:growth & development / Q000502:physiology; D018995:Myosin Heavy Chains / Q000032:analysis / Q000276:immunology; D009543:Nifedipine / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012518:Sarcomeres / Q000737:chemistry / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D013779:Tetrodotoxin / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
D002121:Calcium Channel Blockers; D002614:Chelating Agents; D000185:Actinin; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D013779:Tetrodotoxin; D004533:Egtazic Acid; D012964:Sodium; D014700:Verapamil; D018995:Myosin Heavy Chains; D009543:Nifedipine; D002118:Calcium