Ca transients from Ca channel activity in rat cardiac myocytes reveal dynamics of dyad cleft and troponin C Ca binding.
The properties of the dyad cleft can in principle significantly impact excitation-contraction coupling, but these properties are not easily amenable to experimental investigation. We simultaneously measured the time course of the rise in integrated Ca current (ICa) and the rise in concentration of fura 2 with Ca bound ([Ca-fura 2]) with high time resolution in rat myocytes for conditions under which Ca entry is only via L-type Ca channels and sarcoplasmic reticulum (SR) Ca release is blocked, and compared these measurements with predictions from a finite-element model of cellular Ca diffusion. We found that 1) the time course of the rise of [Ca-fura 2] follows the time course of integrated ICa plus a brief delay (1.36 +/- 0.43 ms, n = 6 cells); 2) from the model, high-affinity Ca binding sites in the dyad cleft at the level previously envisioned would result in a much greater delay (>/=3 ms) and are therefore unlikely to be present at that level; 3) including ATP in the model promoted Ca efflux from the dyad cleft by a factor of 1.57 when low-affinity cleft Ca binding sites were present; 4) the data could only be fit to the model if myofibrillar troponin C (TnC) Ca binding were low affinity (4.56 microM), like that of soluble troponin C, instead of the high-affinity value usually used (0.38 microM). In a "good model," the rate constants for Ca binding and dissociation were 0.375 times the values for soluble TnC; and 5) consequently, intracellular Ca buffering at the rise of the Ca transient is inferred to be low.
Vadakkadath Meethal
Sivan
S
Department of Surgery, University of Wisconsin, Madison WI 53792, USA.
Potter
Katherine T
KT
Redon
David
D
Heisey
Dennis M
DM
Haworth
Robert A
RA
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism* / Q000502:physiology; D004553:Electric Conductivity; D005260:Female; D005456:Fluorescent Dyes / Q000378:metabolism; D016257:Fura-2 / Q000378:metabolism; D008955:Models, Cardiovascular; D032383:Myocytes, Cardiac / Q000378:metabolism*; D051381:Rats; D013997:Time Factors; D019209:Troponin C / Q000378:metabolism*
D015220:Calcium Channels; D005456:Fluorescent Dyes; D019209:Troponin C; D000255:Adenosine Triphosphate; D002118:Calcium; D016257:Fura-2