Myocytes were dissociated from the sinoatrial (SA) node of rat heart using a new enzymatic dissociation technique. Only a small number of isolated SA node myocytes showed regular rhythmic contractions and spontaneous action potentials, and these were used in the present study. The spontaneous action potential was resistant to TTX, and the action potential parameters were similar to those of rabbit and guinea‐pig pacemaker cells. Major time‐ and voltage‐dependent currents were the delayed rectifier K+ current IKr, the L‐type Ca2+ current ICa,L and the sodium current INa. The hyperpolarization‐activated cation current (If) was recorded from ≈50 % of the cells with hyperpolarization beyond ‐90 mV. The instantaneous current jump at the onset of a hyperpolarizing pulse showed inward rectification and was largely blocked by Ba2+. This Ba2+‐sensitive current corresponded well to the inward rectifier K+ current (IK1), although it was much smaller in amplitude than in the ventricle. A sustained inward current was activated on depolarization from ‐80 mV to the voltage range of slow diastolic depolarization. The current was blocked by nicardipine, enlarged by isoprenaline and was insensitive to removal of external Ca2+. These characteristics were similar to the sustained inward current, Ist, previously described in the rabbit and guinea‐pig SA node cells. The role of Ist was considered by constructing empirical equations, which were applied to the experimental record of the action potential. It is demonstrated that the voltage‐dependent activation of Ist constitutes a positive feedback loop with the depolarization of the membrane.

Abstract-Several time-and voltage-dependent ionic currents have been identified in cardiac pacemaker cells, including Na ϩ current, L-and T-type Ca 2ϩ currents, hyperpolarization-activated cation current, and various types of delayed rectifier K ϩ currents. Mathematical models have demonstrated that spontaneous action potentials can be reconstructed by incorporating these currents, but relative contributions of individual currents vary widely between different models. In 1995, the presence of a novel inward current that was activated by depolarization to the potential range of the slow diastolic depolarization in rabbit sinoatrial (SA) node cells was reported. Because the current showed little inactivation during depolarizing pulses, it was called the sustained inward current (I st ). A similar current is also found in SA node cells of the guinea pig and rat and in subsidiary pacemaker atrioventricular node cells. Recently, single-channel analysis has revealed a nicardipine-sensitive, 13-pS Na ϩ current, which is activated by depolarization to the diastolic potential range in guinea pig SA node cells. This channel differs from rapid voltage-gated Na ϩ or L-type Ca 2ϩ channels both in unitary conductance and gating kinetics. Because I st was observed only in spontaneously beating SA node cells, ie, it was absent in quiescent cells dissociated from the same SA or atrioventricular node, an important role of I st for generation of intrinsic cardiac automaticity was suggested. (Circ Res. 2000;87:88-91.) Key Words: cardiac pacemaker Ⅲ sinoatrial node Ⅲ sustained inward current V oltage-and time-dependent gating mechanisms of ionic channels play a primary role in driving membrane potential changes during the action potential. In cardiac pacemaker sinoatrial (SA) node cells, the rising phase of the action potential is due to activation of the L-type Ca 2ϩ channels, and the delayed rectifier K ϩ channels (I Kr or I Ks ) are activated by this depolarization. 1,2 After repolarization, I K slowly deactivates, 3 which results in a gradual increase in the permeability ratio of P Na /P K of the membrane. In the presence of background inward currents, 4 -8 this induces gradual depolarization of the membrane during diastole (pacemaker depolarization).In addition to the above basic mechanisms, involvement of voltage-dependent inward currents, such as I f , is necessary 9 to explain dynamic changes in the pacemaker depolarization under physiological conditions, such as during the positive chronotropic effect of ␤-adrenergic stimulation. Verheijck et al 10 demonstrated that a small I Ca, L can be activated on depolarization to Ϫ60 mV from a holding potential of Ϫ90 mV, suggesting a role for I Ca, L in the pacemaker modulation, although most studies report that I Ca, L is activated only at potentials less negative than Ϫ40 mV. 11,12 Recently, a novel inward current, I st , was identified in spontaneously active SA node cells of rabbits, 13 guinea pigs, 14 and rats 15 and in the atrioventricular (AV) node cells of rabbits. 16,17 Beca...

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The spin densities of the radical anions from 4-methylcatechol, 4-carboxylic catechol, and 3-tiicthylcatechol were calculated by means of the I N D O method with the geometrical parameter adjusting method. The observed proton hyperfine coupling constants were exactly reproduced.

Whole-cell and single-channel currents of the inward rectifier K+ channels from guinea-pig ventricular myocytes were recorded over the range between 5 and 37 degrees C. The conductance for inward currents was decreased by lowering the temperature with a Q10 of 1.28 (whole cell), or 1.41 (single channel) between 20 and 30 degrees C. The open probability of the channel at -100 mV remained high (> 0.9). The distribution of open times was single exponential at all temperatures, confirming a single open state. The entropy change (delta S) for the closing rate of the channel obtained from open-time distribution was -14.0 e.u. (cal/mol/K), and enthalpy change (delta H) was 11.9 kcal/mol. The configuration of closed-time distribution varied markedly by altering the temperature, and three exponentials were necessary to fit the histogram. The slowest component showed higher temperature dependency (delta S = 13.6 e.u. and delta H = 19.0 kcal/mol) than the other two faster components. By assuming a reduced model of C-O at 37 degrees C, the difference in Gibb's free energy (GOC) between the open and closed states was approximately 2 kcal/mol, and the height of the energy barrier for the C-O transition was estimated to be approximately 15 kcal/mol.