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Cancellation of nerve excitation by the reversal of nanosecond stimulus polarity and its relevance to the gating time of sodium channels

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Abstract

The initiation of action potentials (APs) by membrane depolarization occurs after a brief vulnerability period, during which excitation can be abolished by the reversal of the stimulus polarity. This vulnerability period is determined by the time needed for gating of voltage-gated sodium channels (VGSC). We compared nerve excitation by ultra-short uni- and bipolar stimuli to define the time frame of bipolar cancellation and of AP initiation. Propagating APs in isolated frog sciatic nerve were elicited by cathodic pulses (200 ns–300 µs), followed by an anodic (canceling) pulse of the same duration after a 0–200-µs delay. We found that the earliest and the latest boundaries for opening the critical number of VGSC needed to initiate AP are, respectively, between 11 and 20 µs and between 100 and 200 µs after the onset of depolarization. Stronger depolarization accelerated AP initiation, apparently due to faster VGSC opening, but not beyond the 11-µs limit. Bipolar cancellation was augmented by reducing pulse duration, shortening the delay between pulses, decreasing the amplitude of the cathodic pulse, and increasing the amplitude of the anodic one. Some of these characteristics contrasted the bipolar cancellation of cell membrane electroporation (Pakhomov et al. in Bioelectrochemistry 122:123–133, 2018; Gianulis et al. in Bioelectrochemistry 119:10–19, 2017), suggesting different mechanisms. The ratio of nerve excitation thresholds for a unipolar cathodic pulse and a symmetrical bipolar pulse increased as a power function as the pulse duration decreased, in remarkable agreement with the predictions of SENN model of nerve excitation (Reilly and Diamant in Health Phys 83(3):356–365, 2002).

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Abbreviations

AP:

Action potential

CAP:

Compound action potential

CPW:

Coplanar waveguide

CPD:

Cathodic pulse duration

MSA:

Maximum stimulation amplitude

nsEP:

Nanosecond electric pulses

VGSC:

Voltage-gated sodium channels

References

  1. van den Honert C, Mortimer JT (1979) The response of the myelinated nerve fiber to short duration biphasic stimulating currents. Ann Biomed Eng 7(2):117–125

    Article  PubMed  Google Scholar 

  2. Gorman PH, Mortimer JT (1983) The effect of stimulus parameters on the recruitment characteristics of direct nerve stimulation. IEEE Trans Biomed Eng 30(7):407–414

    Article  CAS  PubMed  Google Scholar 

  3. Rubinstein JT, Miller CA, Mino H, Abbas PJ (2001) Analysis of monophasic and biphasic electrical stimulation of nerve. IEEE Trans Biomed Eng 48(10):1065–1070. https://doi.org/10.1109/10.951508

    Article  CAS  PubMed  Google Scholar 

  4. Butikofer R, Lawrence PD (1978) Electrocutaneous nerve stimulation—I: model and experiment. IEEE Trans Biomed Eng 25(6):526–531

    Article  CAS  PubMed  Google Scholar 

  5. Miller CA, Robinson BK, Rubinstein JT, Abbas PJ, Runge-Samuelson CL (2001) Auditory nerve responses to monophasic and biphasic electric stimuli. Hear Res 151(1–2):79–94

    Article  CAS  PubMed  Google Scholar 

  6. Cappaert NL, Ramekers D, Martens HC, Wadman WJ (2013) Efficacy of a new charge-balanced biphasic electrical stimulus in the isolated sciatic nerve and the hippocampal slice. Int J Neural Syst 23(1):1250031. https://doi.org/10.1142/S0129065712500311

    Article  PubMed  Google Scholar 

  7. Macherey O, van Wieringen A, Carlyon RP, Deeks JM, Wouters J (2006) Asymmetric pulses in cochlear implants: effects of pulse shape, polarity, and rate. J Assoc Res Otolaryngol 7(3):253–266. https://doi.org/10.1007/s10162-006-0040-0

    Article  PubMed  PubMed Central  Google Scholar 

  8. Horne CD, Sumner CJ, Seeber BU (2016) A phenomenological model of the electrically stimulated auditory nerve fiber: temporal and biphasic response properties. Front Comput Neurosci 10:8. https://doi.org/10.3389/fncom.2016.00008

    Article  PubMed  PubMed Central  Google Scholar 

  9. Crago PE, Peckham PH, Mortimer JT, Van der Meulen JP (1974) The choice of pulse duration for chronic electrical stimulation via surface, nerve, and intramuscular electrodes. Ann Biomed Eng 2(3):252–264

    Article  CAS  PubMed  Google Scholar 

  10. Pudenz RH, Bullara LA, Dru D, Talalla A (1975) Electrical stimulation of the brain. II. Effects on the blood–brain barrier. Surg Neurol 4(2):265–270

    CAS  PubMed  Google Scholar 

  11. Freites JA, Tobias DJ (2015) Voltage sensing in membranes: from macroscopic currents to molecular motions. J Membr Biol 248(3):419–430. https://doi.org/10.1007/s00232-015-9805-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Gonzalez C, Contreras GF, Peyser A, Larsson P, Neely A, Latorre R (2012) Voltage sensor of ion channels and enzymes. Biophys Rev 4(1):1–15. https://doi.org/10.1007/s12551-011-0061-8

    Article  CAS  PubMed  Google Scholar 

  13. Armstrong CM, Bezanilla F (1974) Charge movement associated with the opening and closing of the activation gates of the Na channels. J Gen Physiol 63(5):533–552

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Catterall WA (2010) Ion channel voltage sensors: structure, function, and pathophysiology. Neuron 67(6):915–928. https://doi.org/10.1016/j.neuron.2010.08.021

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Bezanilla F (2005) Voltage-gated ion channels. IEEE Trans Nanobiosci 4(1):34–48

    Article  Google Scholar 

  16. Bezanilla F (2018) Gating currents. J Gen Physiol 150(7):911–932. https://doi.org/10.1085/jgp.201812090

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Sigg D, Bezanilla F, Stefani E (2003) Fast gating in the Shaker K+ channel and the energy landscape of activation. Proc Natl Acad Sci USA 100(13):7611–7615. https://doi.org/10.1073/pnas.1332409100

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Delemotte L, Kasimova MA, Klein ML, Tarek M, Carnevale V (2015) Free-energy landscape of ion-channel voltage-sensor—domain activation. Proc Natl Acad Sci USA 112(1):124–129. https://doi.org/10.1073/pnas.1416959112

    Article  CAS  PubMed  Google Scholar 

  19. Conti F, Stuhmer W (1989) Quantal charge redistributions accompanying the structural transitions of sodium channels. Eur Biophys J 17(2):53–59

    Article  CAS  PubMed  Google Scholar 

  20. Xiao S, Semenov I, Petrella R, Pakhomov AG, Schoenbach KH (2016) A subnanosecond electric pulse exposure system for biological cells. Med Biol Eng Comput. https://doi.org/10.1007/s11517-016-1516-7

    Article  PubMed  PubMed Central  Google Scholar 

  21. Semenov I, Xiao S, Pakhomov AG (2016) Electroporation by subnanosecond pulses. Biochem Biophys Rep 6:253–259. https://doi.org/10.1016/j.bbrep.2016.05.002

    Article  PubMed  PubMed Central  Google Scholar 

  22. Xiao S, Guo S, Nesin V, Heller R, Schoenbach KH (2011) Subnanosecond electric pulses cause membrane permeabilization and cell death. IEEE Trans Biomed Eng 58(5):1239–1245. https://doi.org/10.1109/TBME.2011.2112360

    Article  PubMed  Google Scholar 

  23. Casciola M, Xiao S, Pakhomov AG (2017) Damage-free peripheral nerve stimulation by 12-ns pulsed electric field. Sci Rep 7(1):10453. https://doi.org/10.1038/s41598-017-10282-5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Guo S, Jing Y, Burcus NI, Lassiter BP, Tanaz R, Heller R, Beebe SJ (2018) Nano-pulse stimulation induces potent immune responses, eradicating local breast cancer while reducing distant metastases. Int J Cancer 142(3):629–640. https://doi.org/10.1002/ijc.31071

    Article  CAS  PubMed  Google Scholar 

  25. Guo J, Dong F, Ding L, Wang K, Zhang J, Fang J (2018) A novel drug-free strategy of nano-pulse stimulation sequence (NPSS) in oral cancer therapy: in vitro and in vivo study. Bioelectrochemistry 123:26–33. https://doi.org/10.1016/j.bioelechem.2018.04.010

    Article  CAS  PubMed  Google Scholar 

  26. Nuccitelli R, McDaniel A, Anand S, Cha J, Mallon Z, Berridge JC, Uecker D (2017) Nano-pulse stimulation is a physical modality that can trigger immunogenic tumor cell death. J Immunother Cancer 5:32. https://doi.org/10.1186/s40425-017-0234-5

    Article  PubMed  PubMed Central  Google Scholar 

  27. Pakhomov AG, Semenov I, Xiao S, Pakhomova ON, Gregory B, Schoenbach KH, Ullery JC, Beier HT, Rajulapati SR, Ibey BL (2014) Cancellation of cellular responses to nanoelectroporation by reversing the stimulus polarity. Cell Mol Life Sci 71(22):4431–4441. https://doi.org/10.1007/s00018-014-1626-z

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Ibey BL, Ullery JC, Pakhomova ON, Roth CC, Semenov I, Beier HT, Tarango M, Xiao S, Schoenbach KH, Pakhomov AG (2014) Bipolar nanosecond electric pulses are less efficient at electropermeabilization and killing cells than monopolar pulses. Biochem Biophys Res Commun 443(2):568–573. https://doi.org/10.1016/j.bbrc.2013.12.004

    Article  CAS  PubMed  Google Scholar 

  29. Gianulis EC, Casciola M, Xiao S, Pakhomova ON, Pakhomov AG (2017) Electropermeabilization by uni- or bipolar nanosecond electric pulses: the impact of extracellular conductivity. Bioelectrochemistry 119:10–19. https://doi.org/10.1016/j.bioelechem.2017.08.005

    Article  CAS  PubMed  Google Scholar 

  30. Gianulis EC, Lee J, Jiang C, Xiao S, Ibey BL, Pakhomov AG (2015) Electroporation of mammalian cells by nanosecond electric field oscillations and its inhibition by the electric field reversal. Sci Rep 5:13818. https://doi.org/10.1038/srep13818

    Article  PubMed  PubMed Central  Google Scholar 

  31. Valdez CM, Barnes RA Jr, Roth CC, Moen EK, Throckmorton GA, Ibey BL (2017) Asymmetrical bipolar nanosecond electric pulse widths modify bipolar cancellation. Sci Rep 7(1):16372. https://doi.org/10.1038/s41598-017-16142-6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Schoenbach KH, Pakhomov AG, Semenov I, Xiao S, Pakhomova ON, Ibey BL (2015) Ion transport into cells exposed to monopolar and bipolar nanosecond pulses. Bioelectrochemistry 103:44–51. https://doi.org/10.1016/j.bioelechem.2014.08.015

    Article  CAS  PubMed  Google Scholar 

  33. Merla C, Pakhomov AG, Semenov I, Vernier PT (2017) Frequency spectrum of induced transmembrane potential and permeabilization efficacy of bipolar electric pulses. Biochim Biophys Acta 1859(7):1282–1290. https://doi.org/10.1016/j.bbamem.2017.04.014

    Article  CAS  Google Scholar 

  34. Pakhomov AG, Grigoryev S, Semenov I, Casciola M, Jiang C, Xiao S (2018) The second phase of bipolar, nanosecond-range electric pulses determines the electroporation efficiency. Bioelectrochemistry 122:123–133. https://doi.org/10.1016/j.bioelechem.2018.03.014

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Gowrishankar TR, Stern JV, Smith KC, Weaver JC (2018) Nanopore occlusion: a biophysical mechanism for bipolar cancellation in cell membranes. Biochem Biophys Res Commun 503(3):1194–1199. https://doi.org/10.1016/j.bbrc.2018.07.024

    Article  CAS  PubMed  Google Scholar 

  36. Pakhomov AG, Pakhomova ON (2010) Nanopores: a distinct transmembrane passageway in electroporated cells. In: Pakhomov AG, Miklavcic D, Markov MS (eds) Advanced electroporation techniques in biology in medicine. CRC Press, Boca Raton, pp 178–194

    Chapter  Google Scholar 

  37. Pakhomov AG, Bowman AM, Ibey BL, Andre FM, Pakhomova ON, Schoenbach KH (2009) Lipid nanopores can form a stable, ion channel-like conduction pathway in cell membrane. Biochem Biophys Res Commun 385(2):181–186. https://doi.org/10.1016/j.bbrc.2009.05.035

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Ibey BL, Mixon DG, Payne JA, Bowman A, Sickendick K, Wilmink GJ, Roach WP, Pakhomov AG (2010) Plasma membrane permeabilization by trains of ultrashort electric pulses. Bioelectrochemistry 79(1):114–121. https://doi.org/10.1016/j.bioelechem.2010.01.001

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Ibey BL, Xiao S, Schoenbach KH, Murphy MR, Pakhomov AG (2009) Plasma membrane permeabilization by 60- and 600-ns electric pulses is determined by the absorbed dose. Bioelectromagnetics 30:92–99

    Article  PubMed  PubMed Central  Google Scholar 

  40. Gianulis EC, Pakhomov AG (2015) Gadolinium modifies the cell membrane to inhibit permeabilization by nanosecond electric pulses. Arch Biochem Biophys 570:1–7. https://doi.org/10.1016/j.abb.2015.02.013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Kastrinaki G, Samsouris C, Kosmidis EK, Papaioannou E, Konstandopoulos AG, Theophilidis G (2015) Assessing the axonal translocation of CeO2 and SiO2 nanoparticles in the sciatic nerve fibers of the frog: an ex vivo electrophysiological study. Int J Nanomed 10:7089–7096. https://doi.org/10.2147/IJN.S93663

    Article  CAS  Google Scholar 

  42. Pakhomov AG, Prol HK, Mathur SP, Akyel Y, Campbell CB (1997) Search for frequency-specific effects of millimeter-wave radiation on isolated nerve function. Bioelectromagnetics 18(4):324–334

    Article  CAS  PubMed  Google Scholar 

  43. Merla C, Liberti M, Marracino P, Muscat A, Azan A, Apollonio F, Mir LM (2018) A wide-band bio-chip for real-time optical detection of bioelectromagnetic interactions with cells. Sci Rep 8(1):5044. https://doi.org/10.1038/s41598-018-23301-w

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Ryan HA, Hirakawa S, Yang E, Zhou C, Xiao S (2018) High-voltage, multiphasic, nanosecond pulses to modulate cellular responses. IEEE Trans Biomed Circuits Syst 12(2):338–350. https://doi.org/10.1109/TBCAS.2017.2786586

    Article  PubMed  Google Scholar 

  45. Paffi A, Pellegrino M, Beccherelli R, Apollonio F, Liberti M, Platano D, Aicardi G, D’Inzeo G (2007) A real-time exposure system for electrophysiological recording in brain slices. IEEE Trans Microw Theory Tech 55(11):2463–2471. https://doi.org/10.1109/Tmtt.2007.908657

    Article  Google Scholar 

  46. Liberti M, Apollonio F, Paffi A, Pellegrino M, D’Inzeo G (2004) A coplanar-waveguide system for cells exposure during electrophysiological recordings. IEEE Trans Microw Theory Tech 52(11):2521–2528. https://doi.org/10.1109/Tmtt.2004.837155

    Article  Google Scholar 

  47. Paffi A, Apollonio F, Lovisolo GA, Marino C, Pinto R, Repacholi M, Liberti M (2010) Considerations for developing an RF exposure system: a review for in vitro biological experiments. IEEE Trans Microw Theory Tech 58(10):2702–2714. https://doi.org/10.1109/Tmtt.2010.2065351

    Article  Google Scholar 

  48. Butikofer R, Lawrence PD (1979) Electrocutaneous nerve stimulation-II: stimulus waveform selection. IEEE Trans Biomed Eng 26(2):69–75. https://doi.org/10.1109/TBME.1978.326286

    Article  CAS  PubMed  Google Scholar 

  49. Reilly JP, Freeman VT, Larkin WD (1985) Sensory effects of transient electrical stimulation–evaluation with a neuroelectric model. IEEE Trans Biomed Eng 32(12):1001–1011. https://doi.org/10.1109/TBME.1985.325509

    Article  CAS  PubMed  Google Scholar 

  50. Reilly JP, Diamant AM (2002) Neuroelectric mechanisms applied to low frequency electric and magnetic field exposure guidelines—part II: non sinusoidal waveforms. Health Phys 83(3):356–365

    Article  CAS  PubMed  Google Scholar 

  51. Pakhomov AG, Semenov I, Casciola M, Xiao S (2017) Neuronal excitation and permeabilization by 200-ns pulsed electric field: an optical membrane potential study with FluoVolt dye. Biochim Biophys Acta 1859(7):1273–1281. https://doi.org/10.1016/j.bbamem.2017.04.016

    Article  CAS  PubMed Central  Google Scholar 

  52. Rogers WR, Merritt JH, Comeaux JA, Kuhnel CT, Moreland DF, Teltschik DG, Lucas JH, Murphy MR (2004) Strength-duration curve for an electrically excitable tissue extended down to near 1 nanosecond. IEEE Trans Plasma Sci 32(4):1587–1599

    Article  Google Scholar 

  53. Hristov K, Mangalanathan U, Casciola M, Pakhomova ON, Pakhomov AG (1860) Expression of voltage-gated calcium channels augments cell susceptibility to membrane disruption by nanosecond pulsed electric field. Biochim Biophys Acta (BBA) Biomembr 11:2175–2183. https://doi.org/10.1016/j.bbamem.2018.08.017

    Article  CAS  Google Scholar 

  54. Semenov I, Xiao S, Pakhomova ON, Pakhomov AG (2013) Recruitment of the intracellular Ca2+ by ultrashort electric stimuli: the impact of pulse duration. Cell Calcium 54(3):145–150. https://doi.org/10.1016/j.ceca.2013.05.008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Azarov JE, Semenov I, Casciola M, Pakhomov AG (2019) Excitation of murine cardiac myocytes by nanosecond pulsed electric field. J Cardiovasc Electrophysiol 30(3):392–401. https://doi.org/10.1111/jce.13834

    Article  PubMed  PubMed Central  Google Scholar 

  56. Sözer EB, Vernier PT (2019) Modulation of biological responses to 2 ns electrical stimuli by field reversal. Biochim Biophys Acta (BBA) Biomembr. https://doi.org/10.1016/j.bbamem.2019.03.019

    Article  Google Scholar 

  57. Sozer EB, Levine ZA, Vernier PT (2017) Quantitative limits on small molecule transport via the electropermeome—measuring and modeling single nanosecond perturbations. Sci Rep 7(1):57. https://doi.org/10.1038/s41598-017-00092-0

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Semenov I, Casciola M, Ibey BL, Xiao S, Pakhomov AG (2018) Electropermeabilization of cells by closely spaced paired nanosecond-range pulses. Bioelectrochemistry 121:135–141. https://doi.org/10.1016/j.bioelechem.2018.01.013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The study was supported by an AFOSR MURI Grant FA9550-15-1-0517 (to AGP) on Nanoelectropulse-Induced Electromechanical Signaling and Control of Biological Systems, administered through Old Dominion University, and by a Grant from Pulse Biosciences (Grant no. 500226-014). Authors want to thank Agnese Denzi for her contribution to the design of the exposure system and solving technical aspects of system realization, and Nicola Lovecchio for his support in proposing optimal engineering solutions.

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Authors and Affiliations

  1. Frank Reidy Research Center for Bioelectrics, Old Dominion University, 4211 Monarch Way, Suite 300, Norfolk, VA, 23508, USA

    Maura Casciola, Shu Xiao, Claudia Muratori & Andrei G. Pakhomov

  2. Department of Electrical and Computer Engineering, Old Dominion University, Norfolk, VA, USA

    Shu Xiao

  3. Department of Information Engineering, Electronics and Telecommunications (D.I.E.T.), Sapienza University of Rome, Rome, Italy

    Francesca Apollonio, Alessandra Paffi & Micaela Liberti

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  1. Maura Casciola
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  2. Shu Xiao
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  3. Francesca Apollonio
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  4. Alessandra Paffi
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  5. Micaela Liberti
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  6. Claudia Muratori
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  7. Andrei G. Pakhomov
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Contributions

MC and AGP conceived the study; MC performed experiments; SX designed and manufactured nsEP generator; MC, FA, AP, and ML designed, characterized, and manufactured the grounded coplanar waveguide and wrote Supplementary material; CM contributed to data discussions and editing the paper; MC and AGP analyzed the data and wrote the paper.

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Correspondence to Andrei G. Pakhomov.

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AGP holds stock in Pulse Biosciences. Other authors declared no conflicts of interest.

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Casciola, M., Xiao, S., Apollonio, F. et al. Cancellation of nerve excitation by the reversal of nanosecond stimulus polarity and its relevance to the gating time of sodium channels. Cell. Mol. Life Sci. 76, 4539–4550 (2019). https://doi.org/10.1007/s00018-019-03126-0

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