Frog alpha- and beta-ryanodine receptors provide distinct intracellular Ca2+ signals in a myogenic cell line.
BACKGROUND
In frog skeletal muscle, two ryanodine receptor (RyR) isoforms, alpha-RyR and beta-RyR, are expressed in nearly equal amounts. However, the roles and significance of the two isoforms in excitation-contraction (E-C) coupling remains to be elucidated.


RESULTS
In this study, we expressed either or both alpha-RyR and beta-RyR in 1B5 RyR-deficient myotubes using the herpes simplex virus 1 helper-free amplicon system. Immunological characterizations revealed that alpha-RyR and beta-RyR are appropriately expressed and targeted at the junctions in 1B5 myotubes. In Ca(2+) imaging studies, each isoform exhibited caffeine-induced Ca(2+) transients, an indicative of Ca(2+)-induced Ca(2+) release (CICR). However, the fashion of Ca(2+) release events was fundamentally different: alpha-RyR mediated graded and sustained Ca(2+) release observed uniformly throughout the cytoplasm, whereas beta-RyR supported all-or-none type regenerative Ca(2+) oscillations and waves. alpha-RyR but not beta-RyR exhibited Ca(2+) transients triggered by membrane depolarization with high [K(+)](o) that were nifedipine-sensitive, indicating that only alpha-RyR mediates depolarization-induced Ca(2+) release. Myotubes co-expressing alpha-RyR and beta-RyR demonstrated high [K(+)](o)-induced Ca(2+) transients which were indistinguishable from those with myotubes expressing alpha-RyR alone. Furthermore, procaine did not affect the peak height of high [K(+)](o)-induced Ca(2+) transients, suggesting minor amplification of Ca(2+) release by beta-RyR via CICR in 1B5 myotubes.


CONCLUSIONS
These findings suggest that alpha-RyR and beta-RyR provide distinct intracellular Ca(2+) signals in a myogenic cell line. These distinct properties may also occur in frog skeletal muscle and will be important for E-C coupling.
Kashiyama
Taku
T
Department of Pharmacology, Juntendo University School of Medicine, Tokyo, Japan.
Murayama
Takashi
T
Suzuki
Erika
E
Allen
Paul D
PD
Ogawa
Yasuo
Y
D000818:Animals; D015153:Blotting, Western; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D007150:Immunohistochemistry; D051379:Mice; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D011892:Rana catesbeiana / Q000378:metabolism*; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics / Q000378:metabolism*
D020033:Protein Isoforms; D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D002118:Calcium