Whole-cell and single-channel alpha1 beta1 gamma2S GABAA receptor currents elicited by a "multipuffer" drug application device

Pflugers Arch. 1996 Oct;432(6):1080-90. doi: 10.1007/s004240050238.

Abstract

Pharmacological characterization of ion channels and receptors in cultured neurons or transfected cell lines requires microapplication of multiple drug solutions during electrophysiological recording. An ideal device could apply a large number of solutions to a limited area with rapid arrival and removal of drug solutions. We describe a novel "multipuffer" rapid application device, based on a modified T-tube with a nozzle made from a glass micropipette tip. Drug solutions are drawn via suction from open reservoirs mounted above the recording chamber through the device into a waste trap. Closure of a solenoid valve between the device and the waste trap causes flow of drug solution though the T-tube nozzle. Any number of drug solutions can be applied with rapid onset (50-100 ms) after a brief fixed delay (100-200 ms). Recombinant alpha1beta1gamma2S GABAA receptors (GABARs) transfected into L929 fibroblasts were recorded using whole-cell and single-channel configurations. Application of GABA resulted in chloride currents with an EC50 of 12.2 microM and a Hill slope of 1.27, suggesting more than one binding site for GABA. GABAR currents were enhanced by diazepam and pentobarbital and inhibited by bicuculline and picrotoxin. Single-channel recordings revealed a main conductance state of 26-28 pS. This device is particularly suitable for rapid, spatially controlled drug applications onto neurons or other cells recorded in the whole-cell configuration, but is also appropriate for isolated single-channel or multichannel membrane patch recordings.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Line
  • Chloride Channels / drug effects
  • Chloride Channels / physiology
  • Electric Conductivity
  • Equipment and Supplies*
  • Ion Channels / drug effects
  • Ion Channels / physiology*
  • Kinetics
  • Mice
  • Pharmaceutical Preparations / administration & dosage*
  • Potassium / administration & dosage
  • Potassium / pharmacology
  • Receptors, GABA / genetics
  • Receptors, GABA / physiology*
  • Recombinant Proteins
  • Solutions
  • Transfection
  • gamma-Aminobutyric Acid / administration & dosage
  • gamma-Aminobutyric Acid / pharmacology

Substances

  • Chloride Channels
  • Ion Channels
  • Pharmaceutical Preparations
  • Receptors, GABA
  • Recombinant Proteins
  • Solutions
  • gamma-Aminobutyric Acid
  • Potassium