Spontaneous and repetitive calcium transients in C2C12 mouse myotubes during in vitro myogenesis.
Fluorescence videomicroscopy was used to monitor changes in the cytosolic free Ca2+ concentration ([Ca2+]i) in the mouse muscle cell line C2Cl2 during in vitro myogenesis. Three different patterns of changes in [Ca2+]i were observed: (i) [Ca2+]i oscillations; (ii) faster Ca2+ events confined to subcellular regions (localized [Ca2+]i spikes) and (iii) [Ca2+]i spikes detectable in the entire myotube (global [Ca2+]i spikes). [Ca2+]i oscillations and localized [Ca2+]i spikes were detectable following the appearance of caffeine-sensitivity in differentiating C2Cl2 cells. Global [Ca2+]i spikes appeared later in the process of myogenesis in cells exhibiting coupling between voltage-operated Ca2+ channels and ryanodine receptors. In contrast to [Ca2+]i oscillations and localized [Ca2+]i spikes, the global events immediately stopped when cells were perfused either with a Ca2+-free solution, or a solution with TTX, TEA and verapamil. To explore further the mechanism of the global [Ca2+]i spikes, membrane currents and fluorescence signals were measured simultaneously. These experiments revealed that global [Ca2+]i spikes were correlated with an inward current. Moreover, while the depletion of the Ca2+ stores blocked [Ca2+]i oscillations and localized [Ca2+]i spikes, it only reduced the amplitude of global [Ca2+]i spikes. It is suggested that, during the earlier stages of the myogenesis, spontaneous and repetitive [Ca2+]i changes may be based on cytosolic oscillatory mechanisms. The coupling between voltage-operated Ca2+ channels and ryanodine receptors seems to be the prerequisite for the appearance of global [Ca2+]i spikes triggered by a membrane oscillatory mechanism, which characterizes the later phases of the myogenic process.
Lorenzon
P
P
Istituto di Fisiologia, Università di Trieste, Italy.
Giovannelli
A
A
Ragozzino
D
D
Eusebi
F
F
Ruzzier
F
F
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001030:Apamin / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D018613:Microscopy, Confocal; D018715:Microscopy, Video; D018482:Muscle, Skeletal / Q000166:cytology / Q000502:physiology*; D009991:Oscillometry; D011189:Potassium Chloride / Q000494:pharmacology; D019789:Tetraethylammonium; D013757:Tetraethylammonium Compounds / Q000494:pharmacology; D013779:Tetrodotoxin / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
D013757:Tetraethylammonium Compounds; D001030:Apamin; D002110:Caffeine; D013779:Tetrodotoxin; D004533:Egtazic Acid; D019789:Tetraethylammonium; D011189:Potassium Chloride; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D000255:Adenosine Triphosphate; D014700:Verapamil; D002118:Calcium