Intrasarcomere [Ca2+] gradients in ventricular myocytes revealed by high speed digital imaging microscopy.
Cardiac muscle contraction is triggered by a small and brief Ca2+ entry across the t-tubular membranes, which is believed to be locally amplified by release of Ca2+ from the adjacent junctional sarcoplasmic reticulum (SR). As Ca2+ diffusion is thought to be markedly attenuated in cells, it has been predicted that significant intrasarcomeric [Ca2+] gradients should exist during activation. To directly test for this, we measured [Ca2+] distribution in single cardiac myocytes using fluorescent [Ca2+] indicators and high speed, three-dimensional digital imaging microscopy and image deconvolution techniques. Steep cytosolic [Ca2+] gradients from the t-tubule region to the center of the sarcomere developed during the first 15 ms of systole. The steepness of these [Ca2+] gradients varied with treatments that altered Ca2+ release from internal stores. Electron probe microanalysis revealed a loss of Ca2+ from the junctional SR and an accumulation, principally in the A-band during activation. We propose that the prolonged existence of [Ca2+] gradients within the sarcomere reflects the relatively long period of Ca2+ release from the SR, the localization of Ca2+ binding sites and Ca2+ sinks remote from sites of release, and diffusion limitations within the sarcomere. The large [Ca2+] transient near the t-tubular/ junctional SR membranes is postulated to explain numerous features of excitation-contraction coupling in cardiac muscle.
Isenberg
G
G
Martin-Luther-Universität, Halle-Wittenberg, Germany.
Etter
E F
EF
Wendt-Gallitelli
M F
MF
Schiefer
A
A
Carrington
W A
WA
Tuft
R A
RA
Fay
F S
FS
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D004577:Electron Probe Microanalysis / Q000295:instrumentation / Q000379:methods; D005456:Fluorescent Dyes; D006168:Guinea Pigs; D006352:Heart Ventricles; D066298:In Vitro Techniques; D007700:Kinetics; D008961:Models, Structural; D009206:Myocardium / Q000378:metabolism* / Q000648:ultrastructure; D012518:Sarcomeres / Q000378:metabolism* / Q000648:ultrastructure; D012519:Sarcoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure; D013997:Time Factors
D005456:Fluorescent Dyes; D002118:Calcium