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Voltage-dependent properties of macroscopic and elementary calcium channel currents in guinea pig ventricular
Insights
This study investigated calcium (Ca) channel currents in guinea pig ventricular myocytes using barium (Ba) solutions. Higher Ba concentrations shifted current-voltage relationships, supporting surface charge neutralization and ion permeation models, and allowing Ca channel density estimation.
Area of Science:
- Cardiovascular Physiology
- Ion Channel Electrophysiology
- Membrane Biophysics
Background:
- Understanding calcium (Ca) channel function is crucial for cardiac electrophysiology.
- Previous models, like Hess and Tsien (1984), proposed mechanisms for ion permeation and surface charge effects.
- Accurate estimation of Ca channel density in cardiac myocytes is essential for interpreting electrophysiological data.
Purpose of the Study:
- To characterize whole-cell and single Ca channel currents in guinea pig ventricular myocytes under varying divalent cation concentrations.
- To investigate the influence of extracellular barium (Ba) concentration on Ca channel gating and permeation.
- To estimate the density of Ca channels in ventricular myocytes using electrophysiological recordings.
Main Methods:
- Whole-cell patch-clamp recordings of Ca channel currents were performed using Cs-internal solution.
- Myocytes were bathed in solutions containing 3.6 mM Ca, 3.6 mM Ba, or 90 mM Ba at 34°C.
- Single Ca channel currents were recorded from cell-attached patches with a 90 mM Ba pipette solution.
Main Results:
- The peak inward current (Vpeak) shifted approximately 30 mV positive in 90 mM Ba compared to 3.6 mM Ba, indicating surface charge effects.
- Activation and inactivation kinetics in 90 mM Ba were shifted ~30 mV rightward versus 3.6 mM Ba.
- Current densities were similar in 3.6 mM Ca and 3.6 mM Ba but approximately 10-fold higher in 90 mM Ba; single-channel data supported whole-cell observations, yielding a myocyte Ca channel density of 3-5 channels/µm².
Conclusions:
- High extracellular Ba concentrations significantly alter Ca channel voltage-dependence, consistent with surface charge neutralization.
- The ion permeation properties observed align with the Hess and Tsien (1984) model.
- The estimated Ca channel density of 3-5 channels/µm² in guinea pig ventricular myocytes is consistent with other studies but contrasts with predictions from single-channel observation frequencies.
Abstract:
Whole-cell Ca channel currents were recorded from guinea pig ventricular myocytes that were internally perfused with Cs solution and bathed in solutions containing 3.6 mM Ca, 3.6 mM Ba or 90 mM Ba (34 degrees C). Single Ca channel currents were recorded from cell-attached membrane patches of similar myocytes; the patch pipettes contained a 90 mM Ba solution. 1. Although the shape of the whole-cell I-V relation was independent of the bathing solution, this was not the case with the location of the inward current maximum (Vpeak); Vpeak in 90 mM Ba was about 30 mV positive to Vpeak in 3.6 mM Ba. 2. The activation and inactivation of whole-cell currents were voltage dependent. Compared to the voltage dependencies in 3.6 mM Ba, those in 90 mM Ba were shifted by about 30 mV to the right, suggesting a neutralization of surface charges. 3. Observations compatible with the ion permeation model proposed by Hess and Tsien (1984) included (a) a depression of current during Ca/Ba solution exchange, (b) a high divalent to monovalent ion permeability, and (c) rectification of the outward limb of the I-V relation. 4. Estimated current densities at Vpeak were similar for myocytes in 3.6 mM Ca and 3.6 mM Ba, and about 10 times larger in 90 mM Ba. 5. Average currents (I) calculated from ensembles of records of single Ca channel current had voltage-dependent time courses resembling those of whole-cell IBa (90 mM). 6. Single-channel I-V relations were superimposable on whole-cell I-V curves suggesting that voltage-dependent single-channel parameters (probability of opening, elementary current amplitude) can be related to the voltage-dependent macroscopic current parameters (activation, instantaneous I-V relation) when scaled by channel number. 7. The density of Ca channels in myocytes was calculated from whole-cell IBa (90 mM) and average current through single channels. The outcome, 3-5 channels/micron 2, agrees with two other recent estimates (Tsien et al. 1983; Lux and Brown 1984). However, it is difficult to reconcile with the much lower density that one would forecast from the frequency of functional channel observation in myocyte membrane patches (Pelzer et al. 1985c).