Changes in Electroporation Thresholds of Lipid Membranes by Surfactants and Peptides LESLIE TUNG, a GREG C. TROIANO, a VINOD SHARMA, a ROBERT M. RAPHAEL, a AND KATHLEEN J. STEBE b Departments of a Biomedical Engineering and b Chemical Engineering, The Johns Hopkins University, Baltimore, Maryland 21205 ABSTRACT: This article reviews recent work from our laboratory that ex- plores how chemical additives may alter the threshold of electroporation of syn- thetic lipid bilayers. The addition of the nonionic block copolymeric surfactant, poloxamer 188 (P188), at a concentration of 1 mM increased the electropora- tion thresholds of planar lipid bilayer membranes made of azolectin. For a 10- s rectangular pulse, P188-treated membranes were found to have a statistical- ly higher threshold voltage, longer latency time to rupture, and lower postpulse conductance. Addition of the nonionic surfactant, octaethyleneglycol-mono-n- dodecyl-ether (C 12 E 8 ), decreased the electroporation threshold of bilayer mem- branes made of 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC) by 10–40% for 10-s- to 10-s-duration pulses, in a concentration-dependent manner over concentrations ranging from 0.1 to 10 mM. Postpulse membrane conductance also increased. The opposite effects of the two surfactants on electroporation thresholds may result from their very different structures, which would encour- age different modes of surfactant-lipid interactions. To examine protein-lipid interactions and their effects on the electroporation threshold, the effects of a channel-forming polypeptide, gramicidin D (gD), was studied on membrane conductance and electroporation threshold. Electropora- tion thresholds for 15-ms pulses were unaffected by addition of gramicidin to POPC at a peptide:lipid concentration estimated to be 1:10,000, but increased significantly at ratios of 1:500 and 1:15, while membrane conductance in- creased monotonically with peptide concentration. A micropipette aspiration technique was applied to giant unilamellar POPC vesicles to measure changes in the membrane physical properties.When gD was added to give an estimated peptide:lipid ratio of 1:15, the membrane area expansivity modulus increased, indicating that the increase in electroporation threshold is correlated with a change in membrane stiffness. Thus, these findings demonstrate that surfac- tants or peptides can mediate the electroporation threshold of lipid bilayers. INTRODUCTION The application of high intensity electric fields to cells or tissues permeabilizes the cell membrane and is believed to produce aqueous-filled pores in the otherwise impermeant lipid phase of the cell membrane, a process referred to as electropora- tion. 1 Factors that mediate electroporation and recovery from electroporation are known to be waveform parameters, temperature, and the ionic and osmotic condi- tions. 1–7 A thorough understanding of these factors is important to minimize the elec- 249