1. Single latent pacemaker cells were isolated from cat right atrium, and studied in a whole-cell configuration using a nystatin-perforated patch recording method. The nystatin method avoids alterations in intracellular Ca2+, cellular constituents and run-down of ionic currents. 2. Depolarizing voltage clamp pulses from -40 mV elicited L-type Ca2+ current (I(Ca)) that exhibited an initial rapid phase of inactivation followed by a secondary slower inward current component that decayed over about 100 ms. The secondary inward component appeared as a slowly decaying inward tail current following short (10-40 ms) depolarizing clamp steps. 3. Slowly decaying inward currents were abolished by internally dialysing pacemaker cells with 2 mM EGTA using a ruptured patch recording method. Inward tail currents were also abolished by exposure to 1 μM ryanodine and significantly decreased by replacing 85% of external Na+ with lithium, without effect on peak I(Ca). These findings identify a Na+-Ca2+ exchange current (I(Na-Ca)) that is mediated by sarcoplasmic reticulum (SR) Ca2+ release. 4. Properties of I(Na-Ca) and I(Ca) differed significantly: (i) I(Ca) exhibited a bell-shaped voltage dependence that peaked at 0 mV and decreased at more positive voltages. I(Na-Ca) was maximal at -10 mV and remained relatively constant at more positive voltages; (ii) a paired pulse protocol showed that the time course of I(Na-Ca) recovery (5 s) was significantly longer than that of I(Ca) (2 s); (iii) cadmium (50 μM) induced an inhibition of I(Ca) that did not correlate in time with changes in I(Na-Ca). 5. The duration of depolarizing steps between 10 and 120 ms had no effect on the time course of I(Na-Ca) tail currents. 6. Isoprenaline ≥ 5 x 10-8 M significantly increased peak I(Ca) amplitude, peak I(Na-Ca) amplitude, accelerated I(Na-Ca) rate of decay and decreased the absolute time of I(Na-Ca) decay. 7. Free-running pacemaker action potentials were clamped during diastole at either -40 or -70 mV (maximum diastolic potential) for variable periods of time. At times between 0.2 and 1 s, I(Na-Ca) exhibited a voltage-dependent increase in amplitude over time, i.e. I(Na-Ca) recovered more rapidly from -70 mV than from -40 mV. At times > 2 s, I(Na-Ca) exhibited a voltage-dependent decline in amplitude over time, i.e. from -40 mV I(Na-Ca) decreased by 10% of maximum whereas from -70 mV I(Na-Ca) decreased by 60% of maximum. 8. Interruption of free-running pacemaker action potentials during the late phase of diastolic depolarization (-50 mV) elicited small, transient and background inward currents. Ryanodine (1 μM) decreased diastolic slope, increased pacemaker cycle length and abolished inward current. In other experiments, isoprenaline (5 x 10-8 M) induced an increase in diastolic slope and inward currents (at -50 mV) that were inhibited by 1 μM ryanodine. 9. These experiments provide an analysis of I(Na-Ca) in latent atrial pacemaker cells using a nystatin-perforated patch method. The time course and voltage dependence of I(Na-Ca) suggest that it contributes to the duration of the pacemaker action potential. In addition, I(Na-Ca) contributes significantly to the slope of diastolic depolarization, and may partially mediate the positive chronotropic response to isoprenaline of latent atrial pacemakers.
|Original language||English (US)|
|Number of pages||23|
|Journal||Journal of Physiology|
|Publication status||Published - 1993|
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