Robust, high-resolution, whole cell patch-clamp capacitance measurements using square wave stimulation.
High-resolution, whole cell capacitance measurements are usually performed using sine wave stimulation using a single frequency or a sum of two frequencies. We present here a high-resolution technique for whole-cell capacitance measurements based on square-wave stimulation. The square wave represents a sum of sinusoidal frequencies at odd harmonics of the base frequency, the amplitude of which is highest for the base frequency and decreases as the frequency increases. The resulting currents can be analyzed by fitting the current relaxations with exponentials, or by a phase-sensitive detector technique. This method provides a resolution undistinguishable from that of single-frequency sine wave stimulation, and allows for clear separation of changes in capacitance, membrane conductance, and access resistance. In addition, it allows for the analysis of more complex equivalent circuits as associated with the presence of narrow fusion pores during degranulation, tracking many equivalent circuit parameters simultaneously. The method is insensitive to changes in the reversal potential, pipette capacitance, or widely varying cell circuit parameters. It thus provides important advantages in terms of robustness for measuring cell capacitances, and allows analysis of complicated changes of the equivalent circuits.
Thompson
R E
RE
Cornell University, School of Applied and Engineering Physics, Ithaca, New York 14853, USA.
Lindau
M
M
Webb
W W
WW
D000818:Animals; D004553:Electric Conductivity; D004594:Electrophysiology / Q000379:methods*; D008564:Membrane Potentials; D018408:Patch-Clamp Techniques / Q000379:methods*; D051381:Rats; D014407:Tumor Cells, Cultured