Voltage- and ligand-gated ryanodine receptors are functionally separated in developing C2C12 mouse myotubes
- 1In order to further understand the role of voltage- and ligand-gated ryanodine receptors in the control of intracellular Ca2+ signalling during myogenesis, changes in cytosolic free calcium concentration ([Ca2+]i) were investigated by fura-2 videoimaging in C2C12 mouse myotubes developing in vitro.
- 2A synchronous [Ca2+]i increase was observed after depolarisation with high [K+], while the Ca2+ response propagated as a wave following caffeine administration. Application of the two stimuli to the same myotube often revealed the existence of cellular zones that were responsive to depolarisation but not to caffeine.
- 3Focal application of high [K+] promoted a [Ca2+]i response detectable only in the cellular areas close to the pipette tip, while focal application of caffeine elicited a [Ca2+]i increase which spread as a Ca2+ wave. Buffering of [Ca2+]i by BAPTA did not affect the pattern of the depolarisation-induced [Ca2+]i transient but abolished the Ca2+ waves elicited by caffeine.
- 4When high [K+] and caffeine were applied in sequence, reciprocal inhibition of the [Ca2+]i responses was observed.
- 5Our results suggest that the different spatial patterns of [Ca2+]i responses are due to uneven distribution of voltage- and ligand-gated ryanodine receptors within the myotube. These two types of receptor control two functionally distinct Ca2+ pools which are part of a common intracellular compartment. Finally, the two differently operated ryanodine receptor channels appear to be independently activated, so that a mechanism of Ca2+-induced Ca2+ release is not required to sustain the global response in C2C12 myotubes.