Dimensions of calcium release domains in frog skeletal muscle fibers

The spatiotemporal properties of the Ca2+ release process in skeletal muscle fibers were determined using an improved confocal spot detection system. Muscle fibers were loaded with the low affinity fluorescent Ca2+ indicator OGB-5N and localized action potential-induced fluorescence signals were recorded from consecutive locations separated by 200 nm within a single sarcomere. Three-dimensional reconstructions on the Ca2+ transients illustrate the existence of domains of increased fluorescence around Ca2+ release sites in the neighborhood of the T-tubules. We estimated the dimensions of these domains by drawing isochronal curves ((delta) F/F vs. spot position) and fitting Gaussian profiles to them. It was found that the earliest detectable full-width-at-half- maximum of these profiles was 0.77 +/- 0.25 micrometers and increased rapidly with time to 1.4 +/- 0.2 micrometers at their peak (18 degree(s)C). A brief, but statistically significant delay of 0.8 +/- 0.42ms was observed between the onset of the fluorescent transients at the Z- and M-lines. Our results are compatible with the possibility that, in response to AP stimulation, Ca2+ is not released exclusively from the junctional region of the sarcoplasmic reticulum, but from a broader expanse of the triadic region.