Patch-clamp techniques for time-resolved capacitance measurements in single cells

Abstract

Two methods are described for estimation of passive cell parameters such as membrane capacitance, membrane conductance and access resistance in tight-seal whole cell recording. Both methods are restricted in their application to cases where the cell under study can be approximated by a simple three-component network with linear properties over some voltage range. One method, referred to as the time domain technique, requires only standard electrophysiological equipment and a computer. Parameters are derived from an analysis of capacitive transients during square wave stimulation. It is readily adaptable to wide variations in experimental parameters. Particularly, it is equally applicable to the "slow whole-cell" configuration (access resistance in the range 100 M omega to 1 G omega) and to normal whole-cell measurements (access resistance typically 10 M omega). The other method applies a sine wave command signal to the cell and employs a lock-in amplifier to analyse the resulting current signal. Two modes of operating the lock-in amplifier are described. One mode provides an output signal directly proportional to small changes in capacitance at maximum resolution (1-10 fF). The other mode, in conjunction with a digital computer, supplies estimates of all passive cell parameters, as does the time domain technique, but with a large amount of data reduction performed by the lock-in amplifier itself. Due to the special hardware, however, this method is not as flexible as the time domain technique.

Dual effects of G‐protein activation on Ca‐dependent exocytosis in bovine lactotrophs

“…The d.c. current and the real and imaginary admittance signals were digitised with an A/D converter (CED 1401, Cambridge, England). The software programme (John Dempster, University of Strathclyde) used holding potential and reversal potential in the calculations [17,18]. Fig.1 shows a typical response of a lactotroph cell during exocytosis, stimulated to secrete with -1/zM Ca in the pipette solution, confirming our previous findings [19,20].…”

Mathematical modeling and statistical analysis of calcium-regulated insulin granule exocytosis in β-cells from mice and humans

“…When exocytosis is triggered by opening Ca 2+ -channels with depolarizing pulses using the voltage-clamp of the patch-clamp technique, the Ca 2+ -current can be measured during the depolarization, but only the total increase in cell membrane capacitance ∆C m is obtained, since C m can not be followed during the depolarization (Lindau and Neher, 1988;Ammälä et al, 1993b;Horrigan and Bookman, 1994). For this reason, depolarizations of different durations are often applied to follow the dynamics of the exocytotic response, assuming that the kinetics of exocytosis is unchanged from pulse to pulse (Ammälä et al, 1993b;Horrigan and Bookman, 1994;Barg et al, 2001;Braun et al, 2008).…”

Roles of the Fast-Releasing and the Slowly Releasing Vesicles in Synaptic Transmission at the Calyx of Held

“…Presynaptic capacitance measurements were performed using an EPC/9 amplifier and the SineϩDC technique (Lindau and Neher, 1988;Sun and Wu, 2001;Yamashita et al, 2005). A sine wave (30 mV in amplitude, 1000 Hz) was superimposed onto a holding potential of Ϫ80 mV.…”