Membrane Electroporation in High Electric Fields

The main topics to be covered build upon our knowledge of the mechanical and rheological properties of membranes and their response to perturbations. In the remaining parts of this introductory section, some of these properties are briefly described to set the basis for a discussion of the behavior and response of membranes to external forces. The following sections consider in detail the morphological changes and poration electric fields can induce in vesicles made of membranes in different phases, and the effects of media environment and various molecular inclusions in the lipid bilayer, that is, the specific membrane composition. Finally, some application aspects of the work are discussed. 

The fleld of membrane structure and characterization is attracting the attention of a growing number of researchers. The basic research in this area builds upon studies performed on the simplest and minimal systems mimicking cell membranes, namely model membranes. Examples of such model membranes are 

Advances in Electrochemical Science and Engineering. Edited by Richard C. Alkire. Dieter M. Kolb, and Jacek Lipkowski 

The two most popular techniques for the formation of giant vesicles (other avail- 

These two methods were further developed and improved by several groups (e.g., 

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Cell lysis under a high electric field is referred to as electroporation [6], Under these conditions, the cell membrane experiences dramatic changes in permeability to macromolecules. The main applications of the electroporation include the electrotransformation of cells and the electroporative gene transfer by the uptake of foreign DNA or RNA (in plants, animals, bacteria, and yeast). The electric field generates permeable microspores at the cell membrane, so that the nucleic acid can be introduced by electroosmosis or diffusion. 

The first method employs the ballistic gun (2,3), where cells are exposed to ballistic bombardment by microparticles coated with the molecules of choice (e g., DNA). The second method is based on exposing the cells to ultrasound leading to an increased transmembrane transport (4). The third approach is based on an electrically driven process (electroporation), where cells are exposed to high-electric fields for short durations of micro- to milliseconds (5). This exposure leads to induction of short-lived permeability changes in the membrane ( pores ) enabling the diffusion of molecules across the membrane along their electrochemical gradients. 

About 10 years ago, a new, easy and versatile technique for the introduction of larger macromolecules into eukaryotic and prokaryotic cells was established (Neumann et al., 1982 Knight, 1981) it is now commonly known as electroporation (Weaver, 1993). It is mainly a physical process, based on the transient permeabiliza-tion of cell membranes by pulses of sufficiently high electric fields. The underlying membrane phenomenon, called reversible electrical breakdown (REB) followed by transient pore formation, occurs if the transmembrane potential reaches values of 0.5 -1.5 V. Membrane pores are generated and molecules are transported through these pores by diffusion, electrical drift, and electroosmosis. Electroporation seems to be a rather universal process in most natural membranes. 

Application of an electric field to lipid bilayers such as those found in cellular membranes causes short-term depolarization of the membrane and formation of pores and other structural changes [17]. These so-called electropores allow the uptake of hydrophilic macromolecules such as plasmid DNA, siRNA, or proteins that are otherwise unable to diffuse passively through this highly regulated barrier. The use of high-voltage electrical pulses to permeabilize cell membranes was first described as a tool to deliver DNA into mammalian cells in 1982 (Wong and Neumann 1982 Neumann et al. 1982). In cuvette-based methods, cells are... 

When cells are placed in external apphed electric fields, they experience an electric force. Electroporation involves the use of short, high voltage pulses to overcome barrier of the cell membrane. When a cell is submitted to an external electric field of high intensity and short duration (kV/cm, p,s), transient and dramatic increase in the permeability of the plasma membrane occurs beyond a point. This phenomenon is popularly called electroporation or electropermeabilization, which allows entry of otherwise impermeable exogenous molecules into the cell interior. This phenomenon has been an active area of research in biology and bioelectrochemistry for more than three decades [3,4] and has found many apphcations in cell biology. 

The creation of very dense electropores (supra electroporation) is probably the initial step to a complete disintegration of the membrane. If the electric field is sufficiently high, micelles instead of membrane structures become stable. Because of the higher mobility of ions in the vicinity of the membrane, a significant increase in conductivity happens. 

It is already 40 years since the first, theoretical papers have been published on the possibility of high-frequency EMF to initiate reversible pores in plasma membranes. The recent advances in theoretical and experimental work, as well as application with therapeutic purposes, were summarized by Pakhomov et al. [16]. This method allows transport through the membrane of small ions and large molecules, which is otherwise impossible. The electric field required to achieve electroporation depends on duration of the pulse and the amplitude of the applied electric field. 

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