FIGURE 5.

Distal SR depletion with proximal caffeine is unaffected by Fluo-3. (A) Myocyte AM-loaded with carboxy-SNARF-1, a non-Ca²⁺ binding fluorophore. Upper panel: fluorescence line scan (y axis shows cell length) used to monitor cell shortening (carboxy-SNARF-1 excitation at 514 nm, emission at 580 nm). (Lower panel) time course of cell shortening (derived from upper panel) in response to caffeine. Protocol of Fig. 4 was applied. Cell initially exposed to SS + 0.3 mM Tet. At circle 1, the proximal end of the cell exposed to SS + 10 mM caffeine; at circle 2 the whole cell exposed to SS + 10 mM caffeine. (B) Ratio of second/first shortening-event is plotted versus time delay between circles 1 and 2 in the presence of intracellular carboxy-SNARF-1 (black; n = 8, 7) or Fluo-3 (gray; n = 6–13). Time constants (τ) obtained from monoexponential fits. The presence of the Ca²⁺-binding fluorophore, Fluo-3, does not affect the rate of decline of distal contraction (and hence distal SR-depletion) during proximal caffeine exposure.