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pubmed:abstractText |
We characterized the influence of conductance changes on whole-cell patch clamp capacitance measurements with a lock-in amplifier and the limitations of the phase-tracking method by numerical computer simulations, error formulas, and experimental tests. At correct phase setting, the artifacts in the capacitance measurement due to activation of linear conductances are small. The cross talk into the capacitance trace is well approximately by the second-order term in the Taylor expansion of the admittance. In the case of nonlinear current-voltage relationships, the measured conductance corresponds to the slope conductance in the range of the sine wave amplitude, and the cross talk into the capacitance trace corresponds to the second-order effect of the slope conductance. The finite gating kinetics of voltage-dependent channels generate phase-shifted currents. These lead to major artifacts in the capacitance measurements when the angular frequency of the sine wave is close to the kinetic rate constant of the channel. However, when the channel kinetics are sufficiently slow, or sufficiently fast, the cross talk is still close to the second-order effect of the measured conductance. The effects of activation of voltage-dependent currents on the capacitance measurements may be estimated, provided a detailed characterization of the kinetics and voltage dependence is available. A phase error of the lock-in amplifier of a few degrees leads to significant projections. The phase-tracking method can be used to keep the phase aligned only during periods of low membrane conductance. However, nonideal properties of the equivalent circuit, in particular the fast capacitance between the pipette and the bath solutions, may lead to large phase errors when the phase-tracking method is used, depending on the electrical properties of the cell. In this article we provide practical values, setting the range where possible artifacts are below defined limits. For proper evaluation of capacitance measurements, the capacitance and conductance traces should always be displayed together.
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