Activation of single cardiac and skeletal ryanodine receptor channels by flash photolysis of caged Ca2+.
Single ryanodine-sensitive sarcoplasmic reticulum (SR) Ca2+ release channels isolated from rabbit skeletal and canine cardiac muscle were reconstituted in planar lipid bilayers. Single channel activity was measured in simple solutions (no ATP or Mg2+) with 250 mM symmetrical Cs+ as charge carrier. A laser flash was used to photolyze caged-Ca2+ (DM-nitrophen) in a small volume directly in front of the bilayer. The free [Ca2+] in this small volume and in the bulk solution was monitored with Ca2+ electrodes. This setup allowed fast, calibrated free [Ca2+] stimuli to be applied repetitively to single SR Ca2+ release channels. A standard photolytically induced free [Ca2+] step (pCa 7-->6) was applied to both the cardiac and skeletal release channels. The rate of channel activation was determined by fitting a single exponential to ensemble currents generated from at least 50 single channel sweeps. The time constants of activation were 1.43 +/- 0.65 ms (mean +/- SD; n = 5) and 1.28 +/- 0.61 ms (n = 5) for cardiac and skeletal channels, respectively. This study presents a method for defining the fast Ca2+ regulation kinetics of single SR Ca2+ release channels and shows that the activation rate of skeletal SR Ca2+ release channels is consistent with a role for CICR in skeletal muscle excitation-contraction coupling.
Györke
S
S
Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston 77555-0641.
Vélez
P
P
Suárez-Isla
B
B
Fill
M
M
D000085:Acetates / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000502:physiology*; D002614:Chelating Agents / Q000494:pharmacology*; D004285:Dogs; D004553:Electric Conductivity / Q000187:drug effects / Q000502:physiology; D005029:Ethylenediamines / Q000494:pharmacology*; D006321:Heart / Q000502:physiology*; D007700:Kinetics; D008051:Lipid Bilayers; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008861:Microsomes / Q000502:physiology; D009124:Muscle Proteins / Q000502:physiology*; D009132:Muscles / Q000502:physiology*; D010782:Photolysis; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel; D012519:Sarcoplasmic Reticulum / Q000502:physiology*
D000085:Acetates; D015220:Calcium Channels; D002614:Chelating Agents; D005029:Ethylenediamines; D008051:Lipid Bilayers; D009124:Muscle Proteins; D019837:Ryanodine Receptor Calcium Release Channel; C057014:DM-nitrophen; D002118:Calcium