Single channel currents at six microsecond resolution elicited by acetylcholine in mouse myoballs

1. A patch-clamp set-up was optimized for low noise and high time resolution. An Axoclamp 200B amplifier was modified to incorporate a Teflon connector to the electrode. An electrode puller was equipped with a hydrogen-oxygen burner to produce quartz-glass pipettes with optimally 0.2 mu m openings and 20 M Omega resistance. 2. The r.m.s. (root mean square) noise of sealed pipettes in the bath ranged from 3.6 fA with 100 Hz filter cut-off to 1.5 pA with 61 kHz filter cut-off. At these extremes currents of 17 fA and more than 3 ms, or 9 pA and more than 6 mu s could be resolved with a negligible error rate. 3. The system was tested on mouse myoballs, recording 9-10 pA single channel currents on-cell at -200 mV polarization which were elicited by 0.1-5000 mu M acetylcholine (ACh). 4. Distributions of open and closed times and of correlations of open times to the preceding closed time defined several open states: single openings with mean durations of 1.2 and 25 mu s, from single-liganded receptors, and bursts of 10 ms mean duration containing on average 800 mu s openings and 16 mu s closings, from double liganded receptors. Above 0.1 mM ACh these openings are interrupted increasingly by on average 18 mu s and 72 mu s channel blocks by ACh.