Elementary Ca2+ and Ba2+ currents were recorded from cell-attached membrane patches of ventricular myocytes from adult guinea pig hearts using the improved patch-clamp technique (Hamill et al. 1981). High concentrations of Ba2+ or Ca2+ (50 or 90 mM) were used in the pipettes to increase the signal-to-noise ratio. All data were derived from elementary current analyses in patches containing only one channel. 1) In response to voltage steps, channel openings occurred singly or in bursts of closely spaced unitary current pulses separated by wider shut intervals. During depolarizations of small amplitude from the resting potential, channel openings occurred almost randomly, whereas during larger depolarizations the events were grouped preferentially at the beginning. 2) Channel openings became more probable with increased depolarization; simultaneously, unitary current amplitudes declined in an ohmic manner. Elementary current amplitudes were slightly larger, when 50 mM Ba2+ replaced 50 mM Ca2+ in the pipettes (slope conductances 9 and 10 pS, respectively), but more than doubled, when Ba2+ was increased to 90 mM (slope conductance 18 pS). Clear outward currents through Ca2+ channels were not observed under these conditions. 3) Peak amplitudes of reconstructed mean currents doubled when 50 mM Ba2+ replaced 50 mM Ca2+ and were larger still when 90 mM Ba2+ was used in the pipettes. The current-voltage relations of the reconstructed mean currents showed a positive shift along the voltage axis as Ba2+ was increased or substituted equimolarly by Ca2+. correspondingly, the open state probability-voltage relations (activation curves) showed a parallel shift as Ba2+ was increased, which was less pronounced when Ba2+ was replaced equimolarly by Ca2+. 4) Determination of Ca2+ channel inactivation using 90 mM Ba2+ in the pipettes indicated an overlap with channel activation in a limited voltage range, resulting in a steady-state "window" current. Inactivation can occur without divalent cation influx. 5) Formation of an inside-out patch resulted in a fast rundown of elementary Ca2+ channel currents. 6) Channel openings were often grouped in bursts. The lifetimes of the open state, the bursts, and the closed states were estimated for Ba2+ and Ca2+ as permeating ions. At least two exponentials were needed to fit the histogram of the lifetimes of all closed states. The lifetimes of the individual openings and bursts were mono-exponentially distributed. The kinetics of the Ca+ channel depended on the voltage and the permeating ion. During +30 mV depolarizations, no significant effect on the permeating ion on channel gating could be detected.(ABSTRACT TRUNCATED AT 400 WORDS)

No abstract

1. The activation kinetics of the IRK1 channel stably expressed in L cells (a murine fibroblast cell line) were studied under the whole-cell voltage clamp. Without polyamines or Mg2+ in the pipettes, inward currents showed an exponential activation on hyperpolarization. The steep inward rectification of the currents around the reversal potential (Erev) could bedescribed by the open-close transition of the channel with first-order kinetics. 2. When the tetravalent organic cation spermine (Spm) was added in the pipettes, the activation kinetics changed; this was explicable by the increase in the closing rate constant.The activation of the currents observed without Spm or Mg2+ in the pipettes was ascribed to the unblocking of the 'endogenous-Spm block'.3. In the presence of the divalent cation putrescine (Put) (Hagiwara, Miyazaki & Rosenthal, 1976;Leech & dependent activation following an instantaneous current Stanfield, 1981;Kurachi, 1985). As the mechanism jump on hyperpolarization. The steep inward rectification accounting for this inward rectification, the block of the around the reversal potential (Erev), and the time-channel by an internal molecule has been proposed dependent property of the currents have been described (Hagiwara & Takahashi, 1974;Hille & Schwarz, 1978). The by the open-close transition of the activation gate, which intracellular cations Mg2+ (Horie,

No abstract

SUMMARY1. The mechanism of enhancement of Ca2+ current by isoprenaline was studied by recording single-channel activity from cell-attached patches on isolated guinea-pig ventricular cells using patch pipettes containing 50 or 100 mM-Ba2 .2. Isoprenaline (100 nM) increased the amplitude of ensemble average currents by increasing the rate of non-blank sweeps (availability). The current decay during 400 ms steps was significantly slowed by isoprenaline. However, the open probability for the non-blank sweeps elicited by 100 ms steps was only slightly increased by the application of isoprenaline.3. The durations of the available state (T.) and the unavailable state (TF) were estimated by the number of non-blank and blank sweeps per run, respectively, applying repetitively 100 ms steps at 2 Hz.4. At large negative holding potentials the distribution of Ts was well fitted by an exponential curve, whose time constant was increased from 1-6 to 3-1 sweeps by 100 nM-isoprenaline, while TF distributed approximately single exponentially with a time constant of 2 0 sweeps in control and 1-3 sweeps in the presence of the drug.5. At depolarized holding potentials a slow voltage-dependent component appeared in the histogram of TF and its time constant was markedly decreased by 100 nM-isoprenaline.6. The availability-voltage relationship was simulated by the Boltzmann equation with a maximal value of 0-4 in the control. The maximal value was increased to 0 7 and the curve was shifted to a depolarizing direction by 7 mV by 100 nmisoprenaline.7. Isoprenaline increased the availability of cardiac Ca2+ channels by increasing the forward rate constant and decreasing the backward rate constant in both voltage-dependent and independent slow state transitions.