title: Characterization of an acute muscle contraction model using cultured C2C12 myotubes.
abstract: A cultured C2C12 myotube contraction system was examined for application as a model for acute contraction-induced phenotypes of skeletal muscle. C2C12 myotubes seeded into 4-well rectangular plates were placed in a contraction system equipped with a carbon electrode at each end. The myotubes were stimulated with electric pulses of 50 V at 1 Hz for 3 ms at 997-ms intervals. Approximately 80% of the myotubes were observed to contract microscopically, and the contractions lasted for at least 3 h with electrical stimulation. Calcium ion (Ca²⁺) transient evoked by the electric pulses was detected fluorescently with Fluo-8. Phosphorylation of protein kinase B/Akt (Akt), 5' AMP-activated protein kinase (AMPK), p38 mitogen-activated protein kinase (p38), and c-Jun NH2-terminal kinase (JNK)1/2, which are intracellular signaling proteins typically activated in exercised/contracted skeletal muscle, was observed in the electrically stimulated C2C12 myotubes. The contractions induced by the electric pulses increased glucose uptake and depleted glycogen in the C2C12 myotubes. C2C12 myotubes that differentiated after exogenous gene transfection by a lipofection or an electroporation method retained their normal contractile ability by electrical stimulation. These findings show that our C2C12 cell contraction system reproduces the muscle phenotypes that arise invivo (exercise), in situ (hindlimb muscles in an anesthetized animal), and invitro (dissected muscle tissues in incubation buffer) by acute muscle contraction, demonstrating that the system is applicable for the analysis of intracellular events evoked by acute muscle contraction.
authors.affiliation: Department of Health Promotion Sciences, Graduate School of Human Health Sciences, Tokyo Metropolitan University, Tokyo, Japan.
authors.forename: Yasuko
authors.initials: Y
authors.lastname: Manabe
authors.forename: Shouta
authors.initials: S
authors.lastname: Miyatake
authors.forename: Mayumi
authors.initials: M
authors.lastname: Takagi
authors.forename: Mio
authors.initials: M
authors.lastname: Nakamura
authors.forename: Ai
authors.initials: A
authors.lastname: Okeda
authors.forename: Taemi
authors.initials: T
authors.lastname: Nakano
authors.forename: Michael F
authors.initials: MF
authors.lastname: Hirshman
authors.forename: Laurie J
authors.initials: LJ
authors.lastname: Goodyear
authors.forename: Nobuharu L
authors.initials: NL
authors.lastname: Fujii
mesh_terms: D000818:Animals; D020013:Calcium Signaling; D002454:Cell Differentiation; D002460:Cell Line; D004558:Electric Stimulation; D005947:Glucose / Q000378:metabolism; D006003:Glycogen / Q000378:metabolism; D007770:L-Lactate Dehydrogenase; D051379:Mice; D048055:Mitogen-Activated Protein Kinase 8 / Q000096:biosynthesis / Q000235:genetics; D008954:Models, Biological; D009119:Muscle Contraction; D018485:Muscle Fibers, Skeletal / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D010750:Phosphoproteins; D011994:Recombinant Proteins; D006003:Glycogen; D007770:L-Lactate Dehydrogenase; D048055:Mitogen-Activated Protein Kinase 8; D005947:Glucose