Time courses of cell electroporation as revealed by submicrosecond imaging of transmembrane potential

Abstract

Changes in the membrane conductance of sea urchin eggs, during the course of electroporation, were investigated over the time range of 0.5 microsecond to 1 ms by imaging the transmembrane potential at a submicrosecond resolution with the voltage-sensitive fluorescent dye RH292. When a rectangular electric pulse of moderate intensity was applied across an egg, a position-dependent potential developed synchronously with the pulse, as theory predicts for a cell with an insulating membrane. From the rise and fall times, the membrane capacitance of unfertilized eggs was estimated to be 0.95 microF/cm2 and the intracellular conductance 220 omega.cm. Under an electric pulse of much higher intensity, the rise of the induced potential stopped at a certain level and then slowly decreased on the microsecond time scale. This saturation and subsequent reversal of the potential development was ascribed to the introduction of finite membrane conductance, or permeabilization of the membrane, by the action of the intense pulse (electroporation). Detailed analysis indicated the following: already at 0.5 microsecond in the rectangular electric pulse, the two sides of the egg facing the positive and negative electrodes were porated and gave a high membrane conductance in the order of 1 S/cm2; the conductance on the positive side appeared higher. Thereafter, the conductance increased steadily, reaching the order of 10 S/cm2 by 1 ms. This increase was faster on the negative-electrode side; by 1 ms the conductance on the negative side was more than twice that on the positive side. The recovery of the porated membrane after the pulse treatment was assessed from the membrane conductance estimated in a second electric pulse of a small amplitude. At least two recovery processes were distinguished, one with a time constant of 7 microseconds and the other 0.5 ms, at the end of which the membrane conductance was already < 0.1 S/cm2.

Electrical stimulation of retinal neurons in epiretinal and subretinal configuration using a multicapacitor array

“…7 and 8). This hypothesis is reasonable since the resistance of the cell membrane is much higher than the cellular interior that quickly becomes equipotential after the onset of a voltage change (membrane time constant Ͻ1 s; Hibino et al 1993).…”

Therapeutic Applications of Electroporation

“…Chang et.al Chang and Reese (1990) were the first to observe them. Other aspects of electroporation, for example, vizualization of transmembrane potential and its evolution in space and time, resealing of pores and asymmetry in permeability of porated cells (sea urchin egg and liposomes) with the help of an optical microscope, were also reported Hibino et al (1993); Kinosita et al (1992). These microscopes have a time resolution of sub-microseconds suitable for studying electroporation.…”

Probing Lipid Bilayers under Ionic Imbalance

“…In neurons and muscle cells, the TMP can reach~130 mV during the propagation of action potentials, which is known as hyperpolarization (3). During electroporation, cells can experience TMPs of up to several volts (4)(5)(6)(7). Electroporation utilizes an external electric field to increase the permeability of the cell membrane, allowing the exchange of materials between the inside and outside of the cell.…”

“…The equator does not exhibit a considerable TMP. Temporally, the TMP gradually increases to its steady-state value over the course of tens of microseconds when an external electric field is turned on (5,22). Although it is not emphasized in the literature, a key point of electroporation is that the TMP results from the redistribution of ions in cell membranes, which corresponds to a local ionic imbalance.…”

“…Above this potential, membranes become conductive due to significant pore formation (5,23). The threshold potential is an intrinsic property of the membrane that is related to the composition of lipids and proteins in the membrane.…”