The role of luminal Ca²⁺ in the generation of Ca²⁺ waves in rat ventricular myocytes

1. We used confocal Ca²⁺ imaging and fluo-3 to investigate the transition of localized Ca²⁺ releases induced by focal caffeine stimulation into propagating Ca²⁺ waves in isolated rat ventricular myocytes. 2. Self-sustaining Ca²⁺ waves could be initiated when the cellular Ca²⁺ load was increased by elevating the extracellular [Ca²⁺] ([Ca²⁺]₀) and they could also be initiated at normal Ca²⁺ loads when the sensitivity of the release sites to cytosolic Ca²⁺ was enhanced by low doses of caffeine. When we prevented the accumulation of extra Ca²⁺ in the luminal compartment of the sarcoplasmic reticulum (SR) with thapsigargin, focal caffeine pulses failed to trigger self-sustaining Ca²⁺ waves on elevation of [Ca²⁺]₀. Inhibition of SR Ca²⁺ uptake by thapsigargin in cells already preloaded with Ca²⁺ above normal levels did not prevent local Ca²⁺ elevations from triggering propagating waves. Moreover, wave velocity increased by 20%. Tetracaine (0.75 nM) caused transient complete inhibition of both local and propagating Ca²⁺ signals, followed by full recovery of the responses due to increased SR Ca²⁺ accumulation. 3. Computer simulations using a numerical model with spatially distinct Ca²⁺ release sites suggested that increased amount of releasable Ca²⁺ might not be sufficient to generate self-sustaining Ca²⁺ waves under conditions of Ca²⁺ overload unless the threshold of release site Ca²⁺ activation was set at relatively low levels (< 1.5 μM). 4. We conclude that the potentiation of SR Ca²⁺ release channels by luminal Ca²⁺ is an important factor in Ca²⁺ wave generation. Wave propagation does not require the translocation of Ca²⁺ from the spreading wave front into the SR. Instead, it relies on luminal Ca²⁺ sensitizing Ca²⁺ release channels to cytosolic Ca²⁺.