VOLTAGE-DEPENDENT AND TIME-DEPENDENT BLOCK OF I-F BY SR2+ IN RABBIT SINOATRIAL NODE CELLS

The hyperpolarization-activated cation current (I-f) was recorded in single myocytes dissociated from the rabbit sino-atrial node and the Sr2+-mediated block of I-f examined. In the presence of 0.1-10 mM Sr2+, the activation phase of I-f was followed by a slower decay during hyperpolarization. In the steady state I/V diagram, the Sr2+ block progressed with increasing hyperpolarization. Ba2+ also blocked I-f, but no time dependency could be observed. The blocking effect of Ca2+ was weak, and Mg2+ had little effect on I-f. The relationship between extracellular Sr2+ concentration [Sr2+](o) and the block was described by a Hill coefficient of 1. The half-saturating [Sr2+](o) were 2.0, 1.6, 1.1 and 0.65 mM at -90, -100, -110 and -120 mV, respectively. The rapid application of Sr2+ during the full activation of I-f using the jet-stream method induced an exponential decline of I-f. The reciprocal time constants were linearly correlated to [Sr2+](o), suggesting 1:1 binding stoichiometry. The fractional electrical distance for the binding site was approximately 0.5 from the external side of the channel. Based on the multiple closed and open states for the I-f channel, a mathematical model for the Sr2+ block was constructed in which the time course of I-f in the presence of Sr2+ was described by the sum of three exponential functions. Fitting the model to the original traces revealed blocking and unblocking rates similar to those obtained from the jet-stream method. At -110 mV, the blocking rate was 410 M(-1)s(-1) while the unblocking rate was 0.16 s(-1). These values are the smallest so far reported for time- and voltage-dependent blockade of ionic channels.