Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments

AUZMENDI, JERÓNIMO A; FERNÁNDEZ DO PORTO, DARÍO; PALLAVECINI, CARLA; MOFFAT, LUCIANO

PLOS ONE

PUBLIC LIBRARY SCIENCE

Lugar: San Francisco ; Año: 2012 vol. 7 p. 42275 - 42275

1932-6203

AbstractBackground: Resolving the kinetics of agonist binding events separately from the subsequent channel gating processesrequires the ability of applying and removing the agonist before channel gating occurs. No reported system has yetachieved pulses shorter than 100 ms, necessary to study nicotinic ACh receptor or AMPA receptor activation.Methodology/Principal Findings: Solution exchange systems deliver short agonist pulses by moving a sharp interfacebetween a control and an experimental solution across a channel preparation. We achieved shorter pulses by means of anexchange system that combines a faster flow velocity, narrower partition between the two streams, and increased velocityand bandwidth of the movement of the interface. The measured response of the entire system was fed back to optimize thevoltage signal applied to the piezoelectric actuator overcoming the spurious oscillations arising from the mechanicalresonances when a high bandwidth driving function was applied. Optimization was accomplished by analyzing the transferfunction of the solution exchange system. When driven by optimized command pulses the enhanced system providedpulses lasting 26 6 1 ms and exchanging 93 6 1% of the solution, as measured in the open tip of a patch pipette.Conclusions/Significance: Pulses of this duration open the experimental study of the molecular events that occur betweenthe agonist binding and the opening of the channel.