The spark and its ember - Separately gated local components of Ca2+ release in skeletal muscle

Amplitude, spatial width, and rise time of Ca2+ sparks were compared in hog fast-twitch muscle, in three conditions that alter activation of release c hannels by [Ca2+]. A total of similar to 17,000 sparks from 30 cells were e valuated. In cells under voltage clamp, caffeine (0.5 or 1 mM) increased av erage spark width by 28%, rise time by 18%, and amplitude by 7%. Increases in width were significant even among events of the same rise time. Spontane ous events recorded in permeabilized fibers with low internal [Mg2+] (0.4 m M), had width and rise times greater than in reference, and not significant ly different than those in caffeine. The spark average in reference rides o n a continuous fluorescence "ridge" and is continued by an "ember," a prolo ngation of width similar to 1 mu m and amplitude <0.2, vanishing in similar to 100 ms. Ridge and ember were absent in caffeine and in permeabilized ce lls. Ex posure of voltage-clamped cells to high internal [Mg2+] (7 mM) had effects opposite to caffeine, reducing spark width by 26% and amplitude by 27%. In high [Mg2+], the ember was visible in individual sparks as a prolon gation of variable duration and amplitude up to 1.2. Based on simulations a nd calculation of Ca2+ release flux from averaged sparks, the increase in s park width caused by caffeine was interpreted as evidence of an increase in radius of the release source-presumably by recruitment of additional chann els. Conversely, spark narrowing suggests loss of contributing channels in high Mg2+. Therefore, these changes in spark width at constant rise times a re evidence of a multichannel origin of sparks. Because ridge and ember wer e reduced by promoters of Ca2+-dependent activation (caffeine, low [Mg2+]) and became more visible in the presence of its inhibitors, they are probabl y manifestations of Ca2+ release directly operated by voltage sensors.