Submitted on May 12, 2003
Accepted on September 22, 2003
Ca transients from Ca channel activity in rat cardiac myocytes reveal the dynamics of dyad cleft and troponin C Ca binding
Sivan Vadakkadath Meethal1, Katherine T Potter1, David Redon1, Dennis M Heisey1, and Robert A Haworth1*
1 Surgery, University of Wisconsin, Madison, WI, USA
* To whom correspondence should be addressed. E-mail: haworth{at}surgery.wisc.edu.
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 have simultaneously measured the time course of the rise in integrated ICa and the rise in [CaFura-2] with high time resolution in rat myocytes, for conditions where Ca entry is only via L-type Ca channels and SR Ca release is blocked, and have 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 [CaFura-2] follows the time course of integrated ICa plus a brief delay (1.36 +/- 0.43msec, 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 (at least 3msec) 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 if low affinity cleft Ca binding sites were present; 4) the data could only be fit to the model if myofibrillar troponin C Ca binding were low affinity (4.56µM), like that of soluble troponin C, instead of the high affinity value usually used (0.38µM). In a "good model", the rate constants for Ca binding and dissociation were 0.375 times the values for soluble TnC; 5) consequently, intracellular Ca buffering at the rise of the Ca transient is inferred to be low.
