Ethylazinphos interaction with membrane lipid organization induces increase of proton permeability and impairment of mitochondrial bioenergetic functions

Ethylazinphos increases the passive proton permeability of lipid bilayers r econstituted with dipalmitoylphosphatidylcholine (DPPC) and mitochondrial l ipids. A sharp increase of proton permeability is detected at insecticide/l ipid molar ratios identical to those inducing phase separation in the plane of DPPC bilayers, as revealed by differential scanning calorimetry (I)SQ. Ethylazinphos progressively depresses the transmembrane potential (AW) of m itochondria supported by piruvate/malate, succinate, or ascorbate/TMPD. Add itionally, a decreased depolarization induced by ADP depends on ethylazinph os concentration, reflecting a phosphorylation depression. This loss of pho sphorylation is a consequence of a decreased AV. A decreased respiratory co ntrol ratio is also observed, since ethylazinphos stimulates state 4 respir ation and inhibits ADP-stimulated respiration (state 3). Ethylazinphos conc entrations up to 100 nmol/mg mitochondrial protein increase the rate of sta te 4 together with a decrease in Delta Psi, without significant perturbatio n of state 3 and carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP)- uncoupled respiration. For increased insecticide concentrations, the state 3 and FCCP-uncoupled respiration are inhibited to approximately the same ex tent. The perturbations are more pronounced when the energization is suppor ted by pyruvate/malate and less effective when succinate is used as substra te. The present data, in association with previous DSC studies, indicate th at ethylazinphos, at concentrations up to 100 nmol/mg mitochondrial protein , interacts with the lipid bilayer of mitochondrial membrane, changing the lipid organization and increasing the proton permeability of the inner memb rane. The increased proton permeability explains the decreased oxidative ph osphorylation coupling. Resulting disturbed ATP synthesis may significantly underlie the mechanisms of ethylazinphos toxicity, since most of cell ener gy in eukaryotes is provided by mitochondria. (C) 2001 Academic Press.