However, to our surprise, only ∼0.2% of BK channel clusters colocalize with RYR clusters (Fig. 5 B). This result alone suggests that a physical coupling between RYRs and BK channels is not required for Ca2+ sparks to generate STOCs. Interestingly, the distance distribution from RYR clusters to their closest BK channel clusters (Fig. 5 B) suggests that a Ca2+ spark could act on multiple BK channel clusters to give rise to a STOC. Several lines of evidence indeed indicate that this is the case. First, Ca2+ sparks generate a Ca2+ microdomain with an area of ∼500 nm in radius within which [Ca2+] is >3.5 µM (Bao et al., 2008). Second, within such a Ca2+ microdomain, the BK channel can be activated with a Po of 0.5 in smooth muscle and in our model (Fig. 6; Tanaka et al., 1997). Third, within the Ca2+ microdomain, there are, on average, two to three BK channel clusters. With these new results, we propose that a Ca2+ spark site is composed of a single cluster of RYRs and a variable number of clustered BK
