title: Atovaquone, a broad spectrum antiparasitic drug, collapses mitochondrial membrane potential in a malarial parasite.
abstract: At present, approaches to studying mitochondrial functions in malarial parasites are quite limited because of the technical difficulties in isolating functional mitochondria in sufficient quantity and purity. We have developed a flow cytometric assay as an alternate means to study mitochondrial functions in intact erythrocytes infected with Plasmodium yoelii, a rodent malaria parasite. By using a very low concentration (2 nM) of a lipophilic cationic fluorescent probe, 3,3'dihexyloxacarbocyanine iodide, we were able to measure mitochondrial membrane potential(DeltaPsim) in live intact parasitized erythrocytes through flow cytometry. The accumulation of the probe into parasite mitochondria was dependent on the presence of a membrane potential since inclusion of carbonyl cyanide m-chlorophenylhydrazone, a protonophore, dissipated the membrane potential and abolished the probe accumulation. We tested the effect of standard mitochondrial inhibitors such as myxothiazole, antimycin, cyanide and rotenone. All of them except rotenone collapsed the DeltaPsim and inhibited respiration. The assay was validated by comparing the EC50 of these compounds for inhibiting DeltaPsim and respiration. This assay was used to investigate the effect of various antimalarial drugs such as chloroquine, tetracycline and a broad spectrum antiparasitic drug atovaquone. We observed that only atovaquone collapsed DeltaPsim and inhibited parasite respiration within minutes after drug treatment. Furthermore, atovaquone had no effect on mammalian DeltaPsim. This suggests that atovaquone, shown to inhibit mitochondrial electron transport, also depolarizes malarial mitochondria with consequent cellular damage and death.
authors.affiliation: Department of Microbiology and Immunology, MCP Hahnemann School of Medicine, Allegheny University of the Health Sciences, Philadelphia, Pennsylvania 19102-1192, USA.
authors.forename: I K
authors.initials: IK
authors.lastname: Srivastava
authors.forename: H
authors.initials: H
authors.lastname: Rottenberg
authors.forename: A B
authors.initials: AB
authors.lastname: Vaidya
mesh_terms: D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D053626:Atovaquone; D004912:Erythrocytes / Q000187:drug effects / Q000469:parasitology / Q000502:physiology; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects*; D051379:Mice; D008807:Mice, Inbred BALB C; D008928:Mitochondria / Q000187:drug effects* / Q000502:physiology; D009285:Naphthoquinones / Q000494:pharmacology*
chemical_list: D000962:Antimalarials; D009285:Naphthoquinones; D053626:Atovaquone