Abstract
1. Changes in cytosolic Ca2+ concentration (delta[Ca2+]c) were measured by global indo-1 fluorescence and compared with changes in subsarcolemmal Ca2+ concentration (delta[Ca2+]sl) indicated by Ca(2+)-activated K+ currents (IK(Ca)). 2. At -50 mV holding potential, 10mM caffeine increased both IK(Ca) and [Ca2+]c without measurable delay. While IK(Ca) peaked within 0.3 +/- 0.16 s (mean +/- S.D.) and decayed to 50% within 0.4 +/- 0.2 s, delta[Ca2+]c peaked within 1.5 +/- 0.5 s and decayed to 50% within 5.2 +/- 1.0 s. The different time courses support the idea that [Ca2+]sl and [Ca2+]c deviate. 3. When 10 mM caffeine was applied 20 s after an initial 2 s caffeine application, IK(Ca) was suppressed to 22 +/- 5% and delta [Ca2+]c to 40 +/- 4%. During the following 1 min caffeine-free period, IK(Ca) recovered to 61 +/- 7% while delta [Ca2+]c remained at 40 +/- 3%. The differences between IK(Ca) and delta[Ca2+]c suggest that Ca2+ deprivation and Ca2+ refilling is faster in peripheral than in central sarcoplasmic reticulum (SR). 4. During the loading period of indo-1, a spontaneous delta[Ca2+]c of 30-80 nM appeared both at -50 mV and at more positive potentials. The amplitude of spontaneous delta[Ca2+]c increased with the amplitude, the frequency or the fusion of spontaneous transient outward currents (STOCs). 5. Block of sarcolemmal Ca2+ fluxes by 1 mM La3+ increased [Ca2+]c by 250 +/- 100 nM and suppressed the spontaneous delta[Ca2+]c. However, La3+ did not significantly retard the rate of decay of STOCs which may therefore be limited by Ca2+ diffusion into the cytosol and not by Ca2+ extrusion. 6. The dissociation of IK(Ca) (or STOCs) and delta[Ca2+]c may indicate a Ca2+ concentration gradient during Ca2+ release directed from the sarcolemma towards the centre of the cell, which later reverses direction.