The Journal of Membrane Biology

, Volume 250, Issue 5, pp 441–453 | Cite as

The Effect of Nanosecond, High-Voltage Electric Pulses on the Shape and Permeability of Polymersome GUVs

  • Tina Batista Napotnik
  • Gianluca Bello
  • Eva-Kathrin Sinner
  • Damijan Miklavčič


Polymersomes, vesicles composed of block copolymers, are promising candidates as membrane alternatives and functional containers, e.g., as potential carriers for functional molecules because of their stability and tunable membrane properties. In the scope of possible use for membrane protein delivery to cells by electrofusion, we investigated the cytotoxicity of such polymersomes as well as the effects of nanosecond electric pulses with variable repetition rate on the shape and permeability of polymersomes in buffers with different conductivities. The polymersomes did not show cytotoxic effects to CHO and B16-F1 cells in vitro in concentrations up to 250 µg/mL (for 48 h) or 1.35 mg/mL (for 60 min), which renders them suitable for interacting with living cells. We observed a significant effect of the pulse repetition rate on electrodeformation of the polymersomes. The electrodeformation was most pronounced in low conductivity buffer, which is favorable for performing electrofusion with cells. However, despite more pronounced deformation at higher pulse repetition rate, the electroporation performance of polymersomes was unaffected and remained in similar ranges both at 10 Hz and 10 kHz. This phenomenon is possibly due to the higher stability and rigidity of polymer vesicles, compared to liposomes, and can serve as an advantage (or disadvantage) depending on the aim in employing polymersomes such as stable membrane alternative architectures or drug vehicles.


Electroporation Polymersomes Electrodeformation Nanosecond electric pulses Membrane alternatives 



The study was supported by the Austrian Science Fund (FWF) and Slovenian Research Agency (ARRS)—Austrian-Slovenian Lead Agency Joint Project: Electroporation as Method for Inserting Functional Membrane Proteins in Mammalian Cells N2-0027 (2015-2017), and by Austrian-Slovenian Lead Agency Joint Project: Electroporation as Method for Inserting Functional Membrane Proteins in Mammalian Cells BI-AT/16-17-003 (2015-2017). It was conducted in the scope of the LEA EBAM: European Laboratory of Pulsed Electric Fields Applications in Biology and Medicine (2011-2018).


  1. Ahmed F, Discher DE (2004) Self-porating polymersomes of PEG-PLA and PEG-PCL: hydrolysis-triggered controlled release vesicles. J Control Release Off J Control Release Soc 96:37–53. doi: 10.1016/j.jconrel.2003.12.021 CrossRefGoogle Scholar
  2. Alberts B, Johnson A, Lewis J et al (2002) Ion channels and the electrical properties of membranes. In: Molecular biology of the cell, 4th edn. Garland Science, New York. Available from
  3. Angelova MI, Dimitrov DS (1986) Liposome electroformation. Faraday Discuss Chem Soc 81:303–311. doi: 10.1039/DC9868100303 CrossRefGoogle Scholar
  4. Aranda S, Riske KA, Lipowsky R, Dimova R (2008) Morphological transitions of vesicles induced by alternating electric fields. Biophys J 95:L19–21. doi: 10.1529/biophysj.108.132548 CrossRefPubMedPubMedCentralGoogle Scholar
  5. Aranda-Espinoza H, Bermudez H, Bates FS, Discher DE (2001) Electromechanical limits of polymersomes. Phys Rev Lett 87:208301. doi: 10.1103/PhysRevLett.87.208301 CrossRefPubMedGoogle Scholar
  6. Bain J, Ruiz-Pérez L, Kennerley AJ et al (2015) In situ formation of magnetopolymersomes via electroporation for MRI. Sci Rep 5:14311. doi: 10.1038/srep14311 CrossRefPubMedPubMedCentralGoogle Scholar
  7. Barltrop JA, Owen TC, Cory AH, Cory JG (1991) 5-(3-carboxymethoxyphenyl)-2-(4,5-dimethylthiazolyl)-3-(4-sulfophenyl)tetrazolium, inner salt (MTS) and related analogs of 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide (MTT) reducing to purple water-soluble formazans As cell-viability indicators. Bioorg Med Chem Lett 1:611–614. doi: 10.1016/S0960-894X(01)81162-8 CrossRefGoogle Scholar
  8. Batista Napotnik T, Reberšek M, Vernier PT et al (2016) Effects of high voltage nanosecond electric pulses on eukaryotic cells (in vitro): a systematic review. Bioelectrochem Amst Neth 110:1–12. doi: 10.1016/j.bioelechem.2016.02.011 CrossRefGoogle Scholar
  9. Bermudez H, Brannan AK, Hammer DA et al (2002) Molecular weight dependence of polymersome membrane structure, elasticity, and stability. Macromolecules 35:8203–8208. doi: 10.1021/ma020669l CrossRefGoogle Scholar
  10. Bermúdez H, Aranda-Espinoza H, Hammer DA, Discher DE (2003) Pore stability and dynamics in polymer membranes. EPL Europhys Lett 64:550. doi: 10.1209/epl/i2003-00264-2 CrossRefGoogle Scholar
  11. Dimova R, Seifert U, Pouligny B et al (2002) Hyperviscous diblock copolymer vesicles. Eur Phys J E 7:241–250. doi: 10.1140/epje/i200101032 CrossRefGoogle Scholar
  12. Dimova R, Riske KA, Aranda S et al (2007) Giant vesicles in electric fields. Soft Matter 3:817–827. doi: 10.1039/B703580B CrossRefGoogle Scholar
  13. Dimova R, Bezlyepkina N, Jordö MD et al (2009) Vesicles in electric fields: some novel aspects of membrane behavior. Soft Matter 5:3201–3212. doi: 10.1039/B901963D CrossRefGoogle Scholar
  14. Discher BM, Won YY, Ege DS et al (1999) Polymersomes: tough vesicles made from diblock copolymers. Science 284:1143–1146CrossRefPubMedGoogle Scholar
  15. Discher BM, Bermudez H, Hammer DA et al (2002) Cross-linked polymersome membranes: vesicles with broadly adjustable properties. J Phys Chem B 106:2848–2854. doi: 10.1021/jp011958z CrossRefGoogle Scholar
  16. Erfani-Moghadam V, Nomani A, Zamani M et al (2014) A novel diblock of copolymer of (monomethoxy poly [ethylene glycol]-oleate) with a small hydrophobic fraction to make stable micelles/polymersomes for curcumin delivery to cancer cells. Int J Nanomed 9:5541–5554. doi: 10.2147/IJN.S63762 CrossRefGoogle Scholar
  17. Gabriel B, Teissié J (1995) Control by electrical parameters of short- and long-term cell death resulting from electropermeabilization of Chinese hamster ovary cells. Biochim Biophys Acta 1266:171–178CrossRefPubMedGoogle Scholar
  18. Gallon E, Matini T, Sasso L et al (2015) Triblock copolymer nanovesicles for pH-responsive targeted delivery and controlled release of siRNA to cancer cells. Biomacromol 16:1924–1937. doi: 10.1021/acs.biomac.5b00286 CrossRefGoogle Scholar
  19. Israelachvili JN (2011) Intermolecular and surface forces, 3rd edn. Academic Press, Santa BarbaraGoogle Scholar
  20. Jordan CA, Neumann E, Sowers AE (2013) Electroporation and electrofusion in cell biology. Springer Science & Business Media, New YorkGoogle Scholar
  21. Katz JS, Levine DH, Davis KP et al (2009) Membrane stabilization of biodegradable polymersomes. Langmuir ACS J Surf Colloids 25:4429–4434. doi: 10.1021/la803769q CrossRefGoogle Scholar
  22. Knorr RL, Staykova M, Gracià RS, Dimova R (2010) Wrinkling and electroporation of giant vesicles in the gel phase. Soft Matter 6:1990–1996. doi: 10.1039/B925929E CrossRefGoogle Scholar
  23. Kotnik T, Miklavcic D (2000) Second-order model of membrane electric field induced by alternating external electric fields. IEEE Trans Biomed Eng 47:1074–1081. doi: 10.1109/10.855935 CrossRefPubMedGoogle Scholar
  24. Kotnik T, Miklavčič D, Slivnik T (1998) Time course of transmembrane voltage induced by time-varying electric fields—a method for theoretical analysis and its application. Bioelectrochem Bioenerg 45:3–16. doi: 10.1016/S0302-4598(97)00093-7 CrossRefGoogle Scholar
  25. Le Meins J-F, Sandre O, Lecommandoux S (2011) Recent trends in the tuning of polymersomes’ membrane properties. Eur Phys J E Soft Matter 34:14. doi: 10.1140/epje/i2011-11014-y CrossRefPubMedGoogle Scholar
  26. Lee JS, Feijen J (2012) Polymersomes for drug delivery: design, formation and characterization. J Control Release Off J Control Release Soc 161:473–483. doi: 10.1016/j.jconrel.2011.10.005 CrossRefGoogle Scholar
  27. Li S, Byrne B, Welsh J, Palmer AF (2007) Self-assembled poly(butadiene)-b-poly(ethylene oxide) polymersomes as paclitaxel carriers. Biotechnol Prog 23:278–285. doi: 10.1021/bp060208+ CrossRefPubMedPubMedCentralGoogle Scholar
  28. Lieber AD, Yehudai-Resheff S, Barnhart EL et al (2013) Membrane tension in rapidly moving cells is determined by cytoskeletal forces. Curr Biol CB 23:1409–1417. doi: 10.1016/j.cub.2013.05.063 CrossRefPubMedGoogle Scholar
  29. Liu L, Mao Z, Zhang J et al (2016) The influence of vesicle shape and medium conductivity on possible electrofusion under a pulsed electric field. PLoS ONE 11:e0158739. doi: 10.1371/journal.pone.0158739 CrossRefPubMedPubMedCentralGoogle Scholar
  30. Lodish H, Berk A, Zipursky SL et al (2000) Molecular cell biology, 4th edn. W. H. Freeman, New YorkGoogle Scholar
  31. Mauroy C, Portet T, Winterhalder M et al (2012) Giant lipid vesicles under electric field pulses assessed by non invasive imaging. Bioelectrochem Amst Neth 87:253–259. doi: 10.1016/j.bioelechem.2012.03.008 CrossRefGoogle Scholar
  32. Mayer LD, Hope MJ, Cullis PR (1986) Vesicles of variable sizes produced by a rapid extrusion procedure. Biochim Biophys Acta 858:161–168CrossRefPubMedGoogle Scholar
  33. Müller LK, Landfester K (2015) Natural liposomes and synthetic polymeric structures for biomedical applications. Biochem Biophys Res Commun 468:411–418. doi: 10.1016/j.bbrc.2015.08.088 CrossRefPubMedGoogle Scholar
  34. Napotnik TB, Reberšek M, Kotnik T et al (2010) Electropermeabilization of endocytotic vesicles in B16 F1 mouse melanoma cells. Med Biol Eng Comput 48:407–413. doi: 10.1007/s11517-010-0599-9 CrossRefPubMedPubMedCentralGoogle Scholar
  35. Needham D, Hochmuth RM (1989) Electro-mechanical permeabilization of lipid vesicles. Role of membrane tension and compressibility. Biophys J 55:1001–1009CrossRefPubMedPubMedCentralGoogle Scholar
  36. Neumann E, Kakorin S (2000) Electroporation of curved lipid membranes in ionic strength gradients. Biophys Chem 85:249–271CrossRefPubMedGoogle Scholar
  37. Neumann E, Kakorin S, Toensing K (1998) Membrane electroporation and electromechanical deformation of vesicles and cells. Faraday Discuss 111:125–157Google Scholar
  38. Oliveira H, Pérez-Andrés E, Thevenot J et al (2013) Magnetic field triggered drug release from polymersomes for cancer therapeutics. J Control Release Off J Control Release Soc 169:165–170. doi: 10.1016/j.jconrel.2013.01.013 CrossRefGoogle Scholar
  39. Perrier DL, Rems L, Boukany PE (2017) Lipid vesicles in pulsed electric fields: fundamental principles of the membrane response and its biomedical applications. Adv Colloid Interface Sci. doi: 10.1016/j.cis.2017.04.016 PubMedGoogle Scholar
  40. Photos PJ, Bermudez H, Aranda-Espinoza H et al (2007) Nuclear pores and membrane holes: generic models for confined chains and entropic barriers in pore stabilization. Soft Matter 3:364–371. doi: 10.1039/B611412C CrossRefGoogle Scholar
  41. Qiao Z-Y, Ji R, Huang X-N et al (2013) Polymersomes from dual responsive block copolymers: drug encapsulation by heating and acid-triggered release. Biomacromol 14:1555–1563. doi: 10.1021/bm400180n CrossRefGoogle Scholar
  42. Ramos C, Bonato D, Winterhalter M et al (2002) Spontaneous lipid vesicle fusion with electropermeabilized cells. FEBS Lett 518:135–138CrossRefPubMedGoogle Scholar
  43. Raz-Ben Aroush D, Yehudai-Resheff S, Keren K (2015) Electrofusion of giant unilamellar vesicles to cells. Methods Cell Biol 125:409–422. doi: 10.1016/bs.mcb.2014.11.005 CrossRefPubMedGoogle Scholar
  44. Rebersek M, Kranjc M, Pavliha D et al (2009) Blumlein configuration for high-repetition-rate pulse generation of variable duration and polarity using synchronized switch control. IEEE Trans Biomed Eng 56:2642–2648. doi: 10.1109/TBME.2009.2027422 CrossRefPubMedGoogle Scholar
  45. Rems L, Usaj M, Kanduser M et al (2013) Cell electrofusion using nanosecond electric pulses. Sci Rep 3:3382. doi: 10.1038/srep03382 CrossRefPubMedPubMedCentralGoogle Scholar
  46. Riske KA, Dimova R (2005) Electro-deformation and poration of giant vesicles viewed with high temporal resolution. Biophys J 88:1143–1155. doi: 10.1529/biophysj.104.050310 CrossRefPubMedGoogle Scholar
  47. Riske KA, Dimova R (2006) Electric pulses induce cylindrical deformations on giant vesicles in salt solutions. Biophys J 91:1778–1786. doi: 10.1529/biophysj.106.081620 CrossRefPubMedPubMedCentralGoogle Scholar
  48. Riske KA, Knorr RL, Dimova R (2009) Bursting of charged multicomponent vesicles subjected to electric pulses. Soft Matter 5:1983–1986. doi: 10.1039/B900548J CrossRefGoogle Scholar
  49. Sadik MM, Li J, Shan JW et al (2011) Vesicle deformation and poration under strong DC electric fields. Phys Rev E 83:66316. doi: 10.1103/PhysRevE.83.066316 CrossRefGoogle Scholar
  50. Saito AC, Ogura T, Fujiwara K et al (2014) Introducing micrometer-sized artificial objects into live cells: a method for cell-giant unilamellar vesicle electrofusion. PLoS ONE 9:e106853. doi: 10.1371/journal.pone.0106853 CrossRefPubMedPubMedCentralGoogle Scholar
  51. Salipante PF, Vlahovska PM (2014) Vesicle deformation in DC electric pulses. Soft Matter 10:3386–3393. doi: 10.1039/C3SM52870G CrossRefPubMedGoogle Scholar
  52. Salipante PF, Knorr RL, Dimova R, Vlahovska PM (2012) Electrodeformation method for measuring the capacitance of bilayer membranes. Soft Matter 8:3810–3816. doi: 10.1039/C2SM07105C CrossRefGoogle Scholar
  53. Shirakashi R, Sukhorukov VL, Reuss R et al (2012) Effects of a pulse electric field on electrofusion of giant unilamellar vesicle (GUV)-Jurkat cell. J Therm Sci Technol 7:589–602. doi: 10.1299/jtst.7.589 CrossRefGoogle Scholar
  54. Singer SJ, Nicolson GL (1972) The fluid mosaic model of the structure of cell membranes. Science 175:720–731. doi: 10.1126/science.175.4023.720 CrossRefPubMedGoogle Scholar
  55. Taubert A, Napoli A, Meier W (2004) Self-assembly of reactive amphiphilic block copolymers as mimetics for biological membranes. Curr Opin Chem Biol 8:598–603. doi: 10.1016/j.cbpa.2004.09.008 CrossRefPubMedGoogle Scholar
  56. Teissie J, Tsong TY (1981) Electric field induced transient pores in phospholipid bilayer vesicles. Biochemistry (Mosc) 20:1548–1554CrossRefGoogle Scholar
  57. Tekle E, Astumian RD, Friauf WA, Chock PB (2001) Asymmetric pore distribution and loss of membrane lipid in electroporated DOPC vesicles. Biophys J 81:960–968. doi: 10.1016/S0006-3495(01)75754-2 CrossRefPubMedPubMedCentralGoogle Scholar
  58. Tekle E, Oubrahim H, Dzekunov SM et al (2005) Selective field effects on intracellular vacuoles and vesicle membranes with nanosecond electric pulses. Biophys J 89:274–284. doi: 10.1529/biophysj.104.054494 CrossRefPubMedPubMedCentralGoogle Scholar
  59. Usaj M, Kanduser M (2012) The systematic study of the electroporation and electrofusion of B16-F1 and CHO cells in isotonic and hypotonic buffer. J Membr Biol 245:583–590. doi: 10.1007/s00232-012-9470-2 CrossRefPubMedGoogle Scholar
  60. Usaj M, Flisar K, Miklavcic D, Kanduser M (2013) Electrofusion of B16-F1 and CHO cells: the comparison of the pulse first and contact first protocols. Bioelectrochem Amst Neth 89:34–41. doi: 10.1016/j.bioelechem.2012.09.001 CrossRefGoogle Scholar
  61. Wang L, Chierico L, Little D et al (2012) Encapsulation of biomacromolecules within polymersomes by electroporation. Angew Chem Int Ed 51:11122–11125. doi: 10.1002/anie.201204169 CrossRefGoogle Scholar
  62. Yamamoto T, Aranda-Espinoza S, Dimova R, Lipowsky R (2010) Stability of spherical vesicles in electric fields. Langmuir ACS J Surf Colloids 26:12390–12407. doi: 10.1021/la1011132 CrossRefGoogle Scholar
  63. Zhang J, Wu L, Meng F et al (2012) pH and reduction dual-bioresponsive polymersomes for efficient intracellular protein delivery. Langmuir ACS J Surf Colloids 28:2056–2065. doi: 10.1021/la203843m CrossRefGoogle Scholar
  64. Zhelev DV, Needham D (1993) Tension-stabilized pores in giant vesicles: determination of pore size and pore line tension. Biochim Biophys Acta 1147:89–104CrossRefPubMedGoogle Scholar
  65. Zimmermann U (1982) Electric field-mediated fusion and related electrical phenomena. Biochim Biophys Acta 694:227–277CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  • Tina Batista Napotnik
    • 1
  • Gianluca Bello
    • 2
  • Eva-Kathrin Sinner
    • 2
  • Damijan Miklavčič
    • 1
  1. 1.Faculty of Electrical EngineeringUniversity of LjubljanaLjubljanaSlovenia
  2. 2.Institute of Synthetic BioarchitecturesUniversity of Natural Resources and Life Sciences (BOKU)ViennaAustria

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