Use of Xenopus Oocytes to Measure Ionic Selectivity of Pore-Forming Peptides and Ion Channels
The Xenopus laevis oocyte is a widely used system for heterologous expression of exogenous ion channel proteins (1, 2). Among other advantages, these easy to obtain, mechanically and electrically stable, large-sized cells enable multiple types of electrophysiological recordings: two-electrode voltage-clamp, single-cell attached or cell-free patch-clamp, and macropatch recordings. The size of an oocyte (1 mm in diameter) also allows the use of additional electrodes (1–3) for injection of diverse materials (Ca2+ chelators, peptides, chemicals, antibodies, proteic-partners, and so on) before or during the course of the electrophysiological experiment. We have successfully used this system to analyze the biophysical properties of pore-forming peptides. Simple perfusion of these peptides induced the formation of channels in the oocyte plasma membrane; these channels can then be studied and characterized in diverse ionic conditions. The ease of the perfusion and the stability of the voltage-clamped oocyte make it a powerful tool for such analyses. Compared with artificial bilayers, oocytes offer a real animal plasma membrane where biophysical properties and toxicity can be studied in the same environment.
Key WordsVoltage clamp reversal potential calcium channels anomalous mole fraction
This work was supported by CNRS, INSERM, Association Française contre les Myopathies, Association pour la Recherche contre le Cancer, Fondation pour la Recherche sur le Cerveau, and Fondation Simone et Cino del Duca. The authors thank Dr. I. Lefevre for critical reading of the manuscript, Dr. V. Lullien-Pellerin for Discussion and J-M Donnay for oocyte preparation.
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