Electroporation of Unilamellar Vesicles Studied by Using a Pore-Mediated Electron-Transfer Reaction

Abstract

The dynamics of electric field-induced pore formation (electroporation) is investigated in 186 nm diameter, unilamellar bilayer vesicles (liposomes) prepared from the phospholipid surfactant dioleoylphosphatidylcholine (DOPC). Formation of reversible transient pores is detected via the spectral changes associated with the electron-transfer reaction between Fe(CN)6 4- ions entrapped in the interior compartment of the vesicle and Ir(Cl)6 2- ions placed in the bulk medium. Upon application of a high-voltage square pulse (E = 8 kV/cm) to the suspension, poration occurs only if the pulse length is ≥200 μs. The field-induced elongation of the time average spherical vesicles to prolate ellipsoid in the direction of the applied field E is detected through the transient birefringence of the system. The onset of elongation precedes the availability of pores large enough for Fe(CN)6 4- to pass through, with an estimated radius of ∼9.6 Å. The average rate of radius growth of the reversible pores (4 × 10-6 m s-1) is found to be 5 orders of magnitude smaller than that of irreversible electropores in planar black lipid membranes of similar phospholipids.

An Alternative Approach to Quantify Partition Processes in Confined Environments: The Electrochemical Behavior of PRODAN in Unilamellar Vesicles

“…The monodispersity of the LUV size achieved through this technique was previously checked. [6,7,37] The unilamellar nature of the pure DOPC vesicles prepared by using the extrusion method [37] was previously confirmed [4,5,7,38] by measuring the extent of quenching by Mn + 2 of their 31 P NMR signals. [39] All samples were used immediately after preparation.…”

“…The lipid 1,2-di-oleoyl-sn-glycero-3-phosphatidylcholine (DOPC T c = À17.3 8C [6,36] ) in chloroform solution, obtained from Avanti Polar Lipids, Inc. (Alabaster Al, USA), and the fluorescent and electroactive probe 6-propionyl-2-dimethylaminonaphthalene, PRODAN, from Molecular Probes (Eugene Or), were used without further purification. At the temperature used in the experiences (20 AE 0.1 8C), the DOPC LUV system is in a pure liquid-crystalline phase.…”

“…For this reason, it is common to use large unilamellar vesicles (LUVs) to get closer to cell-like structures. [5][6][7][8][9] On the other hand, interactions of small molecules with membranes are important issues in membrane biology. Understanding their role in modulating the structure and function of biological membranes requires the knowledge of the location of the molecules in the membranes.…”

A System for the Determination of Planar Lipid Bilayer Breakdown Voltage and Its Applications

“…The exact molecular mechanisms of the phenomenon are still not fully elucidated; however, it is believed that "pores" in the cell membrane are mostly formed by the rearrangement of lipid molecules. Therefore, artificial lipid bilayer structures in the form of planar lipid bilayers or lipid bilayer vesicles can be used as a model for experimental [10], [11] and theoretical studies of electroporation [12].…”

“…In some recent papers, it was reported that the field induces elongation of vesicles in the direction of the applied field prior to electroporation [10], and that the average rate of radius growth of the reversible pores is about five orders of magnitude smaller than that of irreversible electropores in planar lipid membranes of similar phospholipids [11].…”