Ca(2+) spark sites in smooth muscle cells are numerous and differ in number of ryanodine receptors, large-conductance K(+) channels, and coupling ratio between them.
Ca(2+) sparks are highly localized Ca(2+) transients caused by Ca(2+) release from sarcoplasmic reticulum through ryanodine receptors (RyR). In smooth muscle, Ca(2+) sparks activate nearby large-conductance, Ca(2+)-sensitive K(+) (BK) channels to generate spontaneous transient outward currents (STOC). The properties of individual sites that give rise to Ca(2+) sparks have not been examined systematically. We have characterized individual sites in amphibian gastric smooth muscle cells with simultaneous high-speed imaging of Ca(2+) sparks using wide-field digital microscopy and patch-clamp recording of STOC in whole cell mode. We used a signal mass approach to measure the total Ca(2+) released at a site and to estimate the Ca(2+) current flowing through RyR [I(Ca(spark))]. The variance between spark sites was significantly greater than the intrasite variance for the following parameters: Ca(2+) signal mass, I(Ca(spark)), STOC amplitude, and 5-ms isochronic STOC amplitude. Sites that failed to generate STOC did so consistently, while those at the remaining sites generated STOC without failure, allowing the sites to be divided into STOC-generating and STOC-less sites. We also determined the average number of spark sites, which was 42/cell at a minimum and more likely on the order of at least 400/cell. We conclude that 1) spark sites differ in the number of RyR, BK channels, and coupling ratio of RyR-BK channels, and 2) there are numerous Ca(2+) spark-generating sites in smooth muscle cells. The implications of these findings for the organization of the spark microdomain are explored.
Zhuge
Ronghua
R
Department of Physiology, University of Massachusetts Medical Center, 55 Lake Ave. North, Worcester, MA 01655, USA.
Fogarty
Kevin E
KE
Baker
Stephen P
SP
McCarron
John G
JG
Tuft
Richard A
RA
Lifshitz
Lawrence M
LM
Walsh
John V
JV
D000818:Animals; D002024:Bufo marinus; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D003593:Cytoplasm / Q000378:metabolism; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008856:Microscopy, Fluorescence / Q000379:methods*; D032389:Myocytes, Smooth Muscle / Q000502:physiology*; D018408:Patch-Clamp Techniques; D024681:Potassium Channels, Calcium-Activated / Q000502:physiology*; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000502:physiology
D051036:Large-Conductance Calcium-Activated Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium