title: Calcium influx through a possible coupling of cation channels impacts skeletal muscle satellite cell activation in response to mechanical stretch.
abstract: When skeletal muscle is stretched or injured, satellite cells, resident myogenic stem cells positioned beneath the basal lamina of mature muscle fibers, are activated to enter the cell cycle. This signaling pathway is a cascade of events including calcium-calmodulin formation, nitric oxide (NO) radical production by NO synthase, matrix metalloproteinase activation, release of hepatocyte growth factor (HGF) from the extracellular matrix, and presentation of HGF to the receptor c-met, as demonstrated by assays of primary cultures and in vivo experiments. Here, we add evidence that two ion channels, the mechanosensitive cation channel (MS channel) and the long-lasting-type voltage-gated calcium-ion channel (L-VGC channel), mediate the influx of extracellular calcium ions in response to cyclic stretch in satellite cell cultures. When applied to 1-h stretch cultures with individual inhibitors for MS and L-VGC channels (GsMTx-4 and nifedipine, respectively) or with a less specific inhibitor (gadolinium chloride, Gd), satellite cell activation and upstream HGF release were abolished, as revealed by bromodeoxyuridine-incorporation assays and Western blotting of conditioned media, respectively. The inhibition was dose dependent with a maximum at 0.1 μM (GsMTx-4), 10 μM (nifedipine), or 100 μM (Gd) and canceled by addition of HGF to the culture media; a potent inhibitor for transient-type VGC channels (NNC55-0396, 100 μM) did not show any significant inhibitory effect. The stretch response was also abolished when calcium-chelator EGTA (1.8 mM) was added to the medium, indicating the significance of extracellular free calcium ions in our present activation model. Finally, cation/calcium channel dependencies were further documented by calcium-imaging analyses on stretched cells; results clearly demonstrated that calcium ion influx was abolished by GsMTx-4, nifedipine, and EGTA. Therefore, these results provide an additional insight that calcium ions may flow in through L-VGC channels by possible coupling with adjacent MS channel gating that promotes the local depolarization of cell membranes to initiate the satellite cell activation cascade.
authors.affiliation: Department of Animal and Marine Bioresource Sciences, Graduate School of Agriculture, Kyushu University, Hakozaki, Fukuoka, Japan.
authors.forename: Minako
authors.initials: M
authors.lastname: Hara
authors.forename: Kuniko
authors.initials: K
authors.lastname: Tabata
authors.forename: Takahiro
authors.initials: T
authors.lastname: Suzuki
authors.forename: Mai-Khoi Q
authors.initials: MK
authors.lastname: Do
authors.forename: Wataru
authors.initials: W
authors.lastname: Mizunoya
authors.forename: Mako
authors.initials: M
authors.lastname: Nakamura
authors.forename: Shotaro
authors.initials: S
authors.lastname: Nishimura
authors.forename: Shoji
authors.initials: S
authors.lastname: Tabata
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Ikeuchi
authors.forename: Kenji
authors.initials: K
authors.lastname: Sunagawa
authors.forename: Judy E
authors.initials: JE
authors.lastname: Anderson
authors.forename: Ronald E
authors.initials: RE
authors.lastname: Allen
authors.forename: Ryuichi
authors.initials: R
authors.lastname: Tatsumi
mesh_terms: D000818:Animals; D001562:Benzimidazoles / Q000494:pharmacology; D015153:Blotting, Western; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D049109:Cell Proliferation; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D017077:Culture Media, Conditioned / Q000378:metabolism; D003521:Cyclopropanes / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D005682:Gadolinium / Q000494:pharmacology; D017228:Hepatocyte Growth Factor / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D015640:Ion Channel Gating; D008297:Male; D040542:Mechanotransduction, Cellular / Q000187:drug effects; D008564:Membrane Potentials; D008856:Microscopy, Fluorescence; D009281:Naphthalenes / Q000494:pharmacology; D009543:Nifedipine / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D051381:Rats; D032496:Satellite Cells, Skeletal Muscle / Q000187:drug effects / Q000378:metabolism*; D013111:Spider Venoms / Q000494:pharmacology; D013314:Stress, Mechanical; D013997:Time Factors
chemical_list: D001562:Benzimidazoles; D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D002614:Chelating Agents; D017077:Culture Media, Conditioned; D003521:Cyclopropanes; D036341:Intercellular Signaling Peptides and Proteins; C411249:MTx4 protein, Grammostola spatulata; D009281:Naphthalenes; D010455:Peptides; D013111:Spider Venoms; C484287:NNC 55-0396; D004533:Egtazic Acid; D017228:Hepatocyte Growth Factor; D005682:Gadolinium; D009543:Nifedipine; C038958:gadolinium chloride