ELECTROPORATION OF BILAYER LIPID MEMBRANES UNDER CURRENT-CLAMP CONDITIONS

Stanislawa Koronkiewicz*, Slawomir Kalinowski** , Krzysztof Bryl*

* University of Agriculture and Technology,10-957 Olsztyn Kortowo, Poland
** University of Warsaw, Bialystok Branch, Al. J. Pilsudskiego 11/4, 15 443 Bialystok, Poland.

Electroporation has a variety application in biotechnology and medicine. It seems to be the first step for electrofusion in cell membranes. This motivated the presented study, in which chronopotentiometric measurements under current-clamp condition were performed. Bilayer lipid membranes (BLM) have been used as model systems to investigate the process of electroporation.

The aim of our study was to examine the influence of the membrane composition and the membrane external medium on the process of pores formation in BLM. We have studied influence of Al3+, Ca2+ on the process of pore formation. Pores ware generated by a constant-intensity current flowing through the bilayer membrane. The membranes were formed by Mueller-Rudin technique from egg yolk, soybean lecithin and soybean lecithin with derivatives of vitamin A in n decane. Typical Ca2+ and Al3+ concentration in electrolyte (0.1 M KCl) was 0.1 mM.

The chronopotentiometric characteristics showed that the pore open and close cyclically. Shape of these curves were strongly dependend on the current intensity applied. Constant-intensity current of 0.2 nA flowing through the soybean lecithin membranes caused their rupture. Yolk lecithin formed membranes more resistant to electric stress. Destruction of egg yolk lecithin membranes was observed at about 4 nA. Addition of vitamin A derivatives stabilised the membranes. Ca2+ increased BLM electrical stability and preserved very high measurements reproducibility. Addition of Al3+ caused the increase of membrane reversible breakdown voltage from 250 mV till 350 mV. The simple mathematical method of pore size calculations based on chronopotentiometric curves was elaborated. Calculating pore diameter were dependent on constant current intensity applied: 5 nm for 0.2 nA, 8 nm for 0.5 nA, 11 nm for 1,0 nA of the assumed cylindrical single pore with a membrane thickness of 7 nm.

Supported by grant 03030.204