Mitochondrial and sarcolemmal Ca2+ transport reduce [Ca2+]i during caffeine contractures in rabbit cardiac myocytes.

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Abstract
1. Contraction and intracellular Ca2+ (Ca2+i) transients were measured in isolated rabbit ventricular myocytes during twitches and contractures induced by rapid application of 10 mM‐caffeine. 2. The amplitude of caffeine‐induced contractures and the accompanying Ca2+i transients were larger than during normal twitches and also declined more slowly. This may be because only a fraction of sarcoplasmic reticulum (SR) Ca2+ is released during a normal twitch, or because of a temporal overlap of SR Ca2+ release and uptake during the twitch. 3. When a caffeine contracture was initiated in Na(+)‐free, Ca(2+)‐free medium (to prevent sarcolemmal Na(+)‐Ca2+ exchange) the contracture and Ca2+i transient were larger and decreased much more slowly. Thus, Ca2+ extrusion via Na(+)‐Ca2+ exchange may limit the amplitude of caffeine‐induced contractures. 4. Relaxation half‐time (t1/2) for the twitch (0.17 +/‐ 0.03 s) was increased to 0.54 +/‐ 0.07 s for caffeine contractures in control solution and 8.8 +/‐ 1 s for caffeine‐induced contractures in Na(+)‐free, Ca(2+)‐free solution. These results confirm that the SR Ca2+ pump and Na(+)‐Ca2+ exchange are the predominant mechanisms for cytoplasmic Ca2+ removal during relaxation. However slower mechanisms can still reduce intracellular [Ca2+]. 5. Relaxation of caffeine contractures in Na(+)‐free solution was further slowed when (a) mitochondrial Ca2+ uptake was inhibited with the oxidative phosphorylation uncoupler, FCCP (t1/2 = 19.7 +/‐ 3.2 s), or (b) the sarcolemmal Ca(2+)‐ATPase pumping ability was depressed by a large transmembrane [Ca2+] gradient (t1/2 = 27.5 +/‐ 6.9 s). 6. When the four Ca2+ transport systems were simultaneously inhibited (i.e. SR Ca2+ pump, Na(+)‐Ca2+ exchange, mitochondrial Ca2+ uptake and sarcolemmal Ca2+ pump), relaxation was practically abolished, but the cell could recover quickly when Na+ was reintroduced and caffeine removed. 7. We conclude that, under our experimental conditions, the sarcolemmal Ca2+ pump and mitochondria are approximately 37‐ and 50‐fold slower than the Na(+)‐Ca2+ exchange at removing Ca2+ from the cytoplasm. Additionally, the SR Ca2+ pump is about 3‐4 times faster than Na(+)‐Ca2+ exchange.