Calcium transients in 1B5 myotubes lacking ryanodine receptors are relatedto inositol trisphosphate receptors

Potassium depolarization of skeletal myotubes evokes slow calcium waves tha t are unrelated to contraction and involve the cell nucleus (Jaimovich, E., Reyes, R., Liberona, J. L., and Powell, J. A. (2000) Am. J. Physiol. 278, C998-C1010). Studies were done in both the 1B5 (Ry53-/-) murine "dyspedic" myoblast cell line, which does not express any ryanodine receptor isoforms (Moore, R. A, Nguyen, H., Galceran, J., Pessah, I. N., and Alien, P. D. (19 98) J. Cell Biol. 140, 843-851), and C2C12 cells, a myoblast cell line that expresses all three isoforms. Although 1B5 cells lack ryanodine binding, t hey bind tritiated inositol (1,4,5)-trisphosphate. Both type 1 and type 3 i nositol trisphosphate receptors were immune-located in the nuclei of both c ell types and were visualized by Western blot analysis. After stimulation w ith 47 mM K+, inositol trisphosphate mass raised transiently in both cell t ypes. Both fast calcium increase and slow propagated calcium signals were s een in C2C12 myotubes. However, 1B5 myotubes (as well as ryanodine-treated C2C12 myotubes) displayed only a long-lasting, non-propagating calcium incr ease, particularly evident in the nuclei. Calcium signals in 1B5 myotubes w ere almost completely blocked by inhibitors of the inositol trisphosphate p athway: U73122, 2-aminoethoxydiphenyl berate, or xestospongin C. Results su pport the hypothesis that inositol trisphosphate mediates slow calcium sign als in muscle cell ryanodine receptors, having a role in their time course and propagation.