Local, stochastic release of Ca2+ in voltage-clamped rat heart cells: visualization with confocal microscopy.
1. Confocal microscopy and the fluorescent Ca2+ indicator fluo-3 (K+ salt) were used to measure cytosolic free calcium ion concentration ([Ca2+]) during excitation-contraction (E-C) coupling in single, voltage-clamped, rat cardiac ventricular cells. 2. Local [Ca2+]i transients were measured nearly simultaneously in different, separate, subcellular volumes of approximately 2.0 microns 3. During depolarization, local [Ca2+]i transients were distinctly different from each other and from whole-cell [Ca2+]i transients. These differences were particularly apparent during small depolarizations, and were substantially reduced by ryanodine. 3. Components of the local [Ca2+]i transients, particularly those evoked by small depolarizations, were closely similar, in time course and amplitude, to spontaneous local [Ca2+]i transients, or 'sparks' (which have been shown previously to be Ca2+ released from sarcoplasmic reticulum). 4. Analysis of local [Ca2+]i transients in the spatial frequency domain (power spectrum) revealed that high power at spatial frequencies of 0.05-0.2 microns-1 was always associated with spontaneous calcium 'sparks' and with local [Ca2+]i transients evoked by small depolarizing pulses (e.g. to -31 mV). Evoked local [Ca2+]o transients in the presence of ryanodine, and those evoked by depolarization to very positive clamp-pulse potentials (+45 mV), were associated with considerably lower power at this frequency. 5. The results suggest that whole-cell [Ca2+]i transients evoked by voltage-clamp depolarization, and thus by L-type Ca2+ current, are comprised of local [Ca2+]i transients that are similar to the spontaneous calcium 'sparks'. At very positive clamp-pulse potentials, however, the electrically evoked local [Ca2+]i transients may be smaller, perhaps as a result of smaller unitary L-type Ca2+ current.
Department of Physiology, University of Maryland, School of Medicine, Baltimore 21201.
J R
JR
López-López
P S
PS
Shacklock
C W
CW
Balke
W G
WG
Wier
D000814:Aniline Compounds; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D005456:Fluorescent Dyes; D006321:Heart / Q000187:drug effects; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000502:physiology; D018613:Microscopy, Confocal; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D051381:Rats; D017208:Rats, Wistar; D012433:Ryanodine / Q000494:pharmacology; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D014966:Xanthenes
D000814:Aniline Compounds; D015220:Calcium Channels; D005456:Fluorescent Dyes; D014966:Xanthenes; D012433:Ryanodine; C059715:Fluo-3; D002118:Calcium