Single channel and 45Ca2+ flux measurements of the cardiac sarcoplasmic reticulum calcium channel.
Purified canine cardiac sarcoplasmic reticulum vesicles were passively loaded with 45CaCl2 and assayed for Ca2+ releasing activity according to a rapid quench protocol. Ca2+ release from a subpopulation of vesicles was found to be activated by micromolar Ca2+ and millimolar adenine nucleotides, and inhibited by millimolar Mg2+ and micromolar ruthenium red. 45Ca2+ release in the presence of 10 microM free Ca2+ gave a half-time for efflux of 20 ms. Addition of 5 mM ATP to 10 microM free Ca2+ increased efflux twofold (t1/2 = 10 ms). A high-conductance calcium-conducting channel was incorporated into planar lipid bilayers from the purified cardiac sarcoplasmic reticulum fractions. The channel displayed a unitary conductance of 75 +/- 3 pS in 53 mM trans Ca2+ and was selective for Ca2+ vs. Tris+ by a ratio of 8.74. The channel was dependent on cis Ca2+ for activity and was also stimulated by millimolar ATP. Micromolar ruthenium red and millimolar Mg2+ were inhibitory, and reduced open probability in single-channel recordings. These studies suggest that cardiac sarcoplasmic reticulum contains a high-conductance Ca2+ channel that releases Ca2+ with rates significant to excitation-contraction coupling.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002132:Calcium Radioisotopes; D004285:Dogs; D007473:Ion Channels / Q000502:physiology*; D007700:Kinetics; D008051:Lipid Bilayers; D008564:Membrane Potentials; D009132:Muscles / Q000502:physiology; D012519:Sarcoplasmic Reticulum / Q000502:physiology*
D002132:Calcium Radioisotopes; D007473:Ion Channels; D008051:Lipid Bilayers; D002118:Calcium