A vertical flow chamber for Xenopus oocyte electrophysiology and automated drug screening.
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.
Joshi
P R
PR
School of Pharmacy, The University of Louisiana at Monroe, Monroe, LA 71209, USA.
Suryanarayanan
A
A
Schulte
M K
MK
D000818:Animals; D001331:Automation / Q000295:instrumentation* / Q000379:methods; D017076:Computer-Aided Design; D015194:Diffusion Chambers, Culture / Q000295:instrumentation* / Q000379:methods; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004553:Electric Conductivity; D004594:Electrophysiology; D005260:Female; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008845:Microinjections / Q000379:methods; D009865:Oocytes / Q000187:drug effects* / Q000502:physiology; D018408:Patch-Clamp Techniques; D010477:Perfusion / Q000379:methods; D018075:RNA, Complementary / Q000096:biosynthesis; D044406:Receptors, Serotonin, 5-HT3 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D012701:Serotonin / Q000494:pharmacology; D014982:Xenopus laevis
D018075:RNA, Complementary; D044406:Receptors, Serotonin, 5-HT3; D012701:Serotonin