The sterol composition of Trypanosoma cruzi changes after growth in different culture media and results in different sensitivity to digitonin-permeabilization

Respiration, oxidative phosphorylation, and the corresponding changes in me mbrane potential (Delta Psi) of Trypanosoma cruzi epimastigotes grown eithe r in liver infusion-tryptose (LIT) or brain heart infusion (BHI) culture me dium were assayed in situ using digitonin to render their plasma membrane p ermeable to succinate, ADP, safranine O, and other small molecules. When th e cells were permeabilized with 64 muM digitionin, a concentration previous ly used with epimastigotes, the ability of the cells grown in LIT medium to sustain oxidative phosphorylation was demonstrated by the detection of an oligomycin-sensitive decrease in mitochondrial membrane potential induced b y ADP. In contrast, the cells grown in BHI medium were not able to sustain a stable membrane potential and did not respond to ADP addition. Analyses o f oxygen consumption by these permeabilized cells indicated that the rate o f basal respiration, which was similar in both cell types, was significantl y decreased by 64 muM digitonin. Addition of ADP to the permeabilized cells grown in LIT medium promoted an oligomycin-sensitive transition from resti ng to phosphorylating respiration in contrast to the cells grown in BHI med ium, whose respiration decreased steadily and did not respond either to ADP or CCCP. Titration of the cells grown in BIR medium with different digiton in concentrations indicated that their mitochondria have higher sensitivity to digitonin than those grown in LIT medium. Analysis of the sterol compos ition of epimastigotes grown in the two different media showed a higher per centage of cholesterol in total and mitochondrial extracts of epimastigotes grown in BIH medium as compared to those grown in LIT medium, suggesting t he involvement of this sterol in their increased sensitivity to digitonin-p ermeabilization.