Biophysical and Biochemical Properties of the Large Conductance Potassium Channel in Fibroblast Mitochondria

Abstract

Potassium channels have been found in the inner mitochondrial membranes of various cells. The activation of these channels is cytoprotective. Hence, the identification of ion channels present in the inner mitochondrial membrane of fibroblast cells is important in distinguishing possible protective mechanisms in these cells.In our study single channel activity of a large conductance Ca2+-regulated potassium channel was measured by patch-clamp of mitoplasts isolated from fibroblast cell line. Mitoplast were prepared by addition to a hypotonic solution causing unfolding of the cristae of the inner membrane and consequently breaking of the outer membrane. Isotonicity was restored by adding a hypertonic solution. A potassium selective current was recorded with a mean conductance of 280 ± 2 pS in symmetrical 150 mM KCl solution. The channel was activated by Ca2+ at micromolar concentrations and inhibited irreversibly by paxilline, an selective inhibitor of the BKCa channel. The substances known to modulate BKCa channel activity were found to influence the bioenergetics of mitochondria, isolated from human dermal fibroblast cells. In isolated mitochondria, 10 μM NS1619 depolarized the mitochondrial membrane potential and stimulated nonphosphorylating respiration. This effect was blocked by 20 μM paxilline.Our findings indicate presence of the large conductance Ca2+-regulated potassium channels in the inner mitochondrial membranes of human fibroblast.This study was supported by a grant MERIS PBS1/B8/1/2012 from the National Centre of Research and Development.