A vertical flow chamber for Xenopus oocyte electrophysiology and automated drug screening

J Neurosci Methods. 2004 Jan 15;132(1):69-79. doi: 10.1016/j.jneumeth.2003.09.002.

Abstract

Xenopus laevis oocytes are used extensively in the study of ion channel coupled receptors. Efficient use of oocytes for ion channel characterization requires a system that is inherently stable, reproducible, minimizes drug volumes, and maximizes oocyte longevity. We have constructed a vertical flow oocyte recording chamber to address the aforesaid issues, where the oocyte is placed in a funnel-shaped chamber and perfused from the bottom of the funnel. The vertical rather than horizontal flow of perfusate results in an unusually stable environment for oocyte recording. Two-electrode voltage clamp recordings from a single oocyte are acquired easily and routinely over several hours while maintaining stable baseline currents and reproducible response profiles. Chamber characteristics were tested using a serotonin ligand-gated ion channel receptor (5-HT3R). Data obtained from this system corresponds well with published data. To further test the stability and reliability of this perfusion chamber, we constructed an automated oocyte perfusion system utilizing a commonly available HPLC autosampler. We were able to obtain dose-response curves for various 5-HT3AR ligands using the automated perfusion system with minimal user intervention. Such a system can easily satisfy need for automated oocyte electrophysiology in academic settings, especially small to medium sized laboratories.

Publication types

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

MeSH terms

  • Animals
  • Automation / instrumentation*
  • Automation / methods
  • Computer-Aided Design
  • Diffusion Chambers, Culture / instrumentation*
  • Diffusion Chambers, Culture / methods
  • Dose-Response Relationship, Drug
  • Drug Evaluation, Preclinical / methods*
  • Electric Conductivity
  • Electrophysiology*
  • Female
  • Humans
  • Membrane Potentials / drug effects
  • Membrane Potentials / physiology
  • Microinjections / methods
  • Oocytes / drug effects*
  • Oocytes / physiology
  • Patch-Clamp Techniques
  • Perfusion / methods
  • RNA, Complementary / biosynthesis
  • Receptors, Serotonin, 5-HT3 / drug effects
  • Receptors, Serotonin, 5-HT3 / genetics
  • Receptors, Serotonin, 5-HT3 / metabolism
  • Serotonin / pharmacology
  • Xenopus laevis

Substances

  • RNA, Complementary
  • Receptors, Serotonin, 5-HT3
  • Serotonin