Optimizing planar lipid bilayer single-channel recordings for high resolution with rapid voltage steps.

摘要

We describe two enhancements of the planar bilayer recording method which enable low-noise recordings of single-channel currents activated by voltage steps in planar bilayers formed on apertures in partitions separating two open chambers. First, we have refined a simple and effective procedure for making small bilayer apertures (25-80 micrograms diam) in plastic cups. These apertures combine the favorable properties of very thin edges, good mechanical strength, and low stray capacitance. In addition to enabling formation of small, low-capacitance bilayers, this aperture design also minimizes the access resistance to the bilayer, thereby improving the low-noise performance. Second, we have used a patch-clamp headstage modified to provide logic-controlled switching between a high-gain (50 G omega) feedback resistor for high-resolution recording and a low-gain (50 M omega) feedback resistor for rapid charging of the bilayer capacitance. The gain is switched from high to low before a voltage step and then back to high gain 25 microseconds after the step. With digital subtraction of the residual currents produced by the gain switching and electrostrictive changes in bilayer capacitance, we can achieve a steady current baseline within 1 ms after the voltage step. These enhancements broaden the range of experimental applications for the planar bilayer method by combining the high resolution previously attained only with small bilayers formed on pipette tips with the flexibility of experimental design possible with planar bilayers in open chambers. We illustrate application of these methods with recordings of the voltage-step activation of a voltage-gated potassium channel.