Activation of single cardiac and skeletal ryanodine receptor channels by flash photolysis of caged Ca2+

Biophys J. 1994 Jun;66(6):1879-86. doi: 10.1016/S0006-3495(94)80981-6.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acetates / pharmacology*
  • Animals
  • Calcium / metabolism*
  • Calcium Channels / physiology*
  • Chelating Agents / pharmacology*
  • Dogs
  • Electric Conductivity / drug effects
  • Electric Conductivity / physiology
  • Ethylenediamines / pharmacology*
  • Heart / physiology*
  • Kinetics
  • Lipid Bilayers
  • Membrane Potentials / drug effects
  • Membrane Potentials / physiology
  • Microsomes / physiology
  • Muscle Proteins / physiology*
  • Muscles / physiology*
  • Photolysis
  • Rabbits
  • Ryanodine Receptor Calcium Release Channel
  • Sarcoplasmic Reticulum / physiology*

Substances

  • Acetates
  • Calcium Channels
  • Chelating Agents
  • Ethylenediamines
  • Lipid Bilayers
  • Muscle Proteins
  • Ryanodine Receptor Calcium Release Channel
  • DM-nitrophen
  • Calcium