Vercesi A E, Docampo R
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801.
Biochem J. 1992 Jun 1;284 ( Pt 2)(Pt 2):463-7. doi: 10.1042/bj2840463.
The use of low concentrations of digitonin allowed the quantitative determination of the mitochondrial membrane potential of Leishmania donovani promastigotes in situ using safranine O. L. donovani mitochondria were able to build up and retain a membrane potential of a value comparable with that of mammalian mitochondria. The response of promastigotes mitochondrial membrane potential to phosphate, carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP), valinomycin and Ca2+ indicates that these mitochondria behave similarly to vertebrate mitochondria with regard to the properties of their electrochemical proton gradient. When L. donovani promastigotes were permeabilized with digitonin in a reaction medium containing MgATP, succinate and 3.5 microM free Ca2+, they lowered the medium Ca2+ concentration to the submicromolar level (0.05-0.1 microM). The presence of 1 microM-FCCP decreased by about 75% the initial rate of Ca2+ sequestration by these permeabilized cells. This FCCP-insensitive Ca2+ uptake, probably by the endoplasmic reticulum, was completely inhibited by 500 microM-vanadate. On the other hand, when vanadate instead of FCCP was present, the initial rate of Ca2+ accumulation was decreased by about 25% and the Ca2+ set point was increased to 0.7 microM. The succinate-dependence and FCCP-and Ruthenium Red-sensitivity of the Ca2+ uptake detected in the presence of vanadate indicate that this uptake is probably by the mitochondria. This interpretation was further supported by the Ruthenium Red-sensitive decrease in the mitochondrial membrane potential caused by Ca2+ addition. The anti-leishmanial cationic drugs pentamidine and WR-6026 also induced a rapid collapse of the mitochondrial inner membrane potential of L. donovani promastigotes.
使用低浓度的洋地黄皂苷，可利用番红O原位定量测定杜氏利什曼原虫前鞭毛体的线粒体膜电位。杜氏利什曼原虫线粒体能够建立并维持与哺乳动物线粒体相当的膜电位值。前鞭毛体线粒体膜电位对磷酸盐、羰基氰化物p -（三氟甲氧基）苯腙（FCCP）、缬氨霉素和Ca2 +的反应表明，就其电化学质子梯度的性质而言，这些线粒体的行为与脊椎动物线粒体相似。当杜氏利什曼原虫前鞭毛体在含有MgATP、琥珀酸和3.5微摩尔游离Ca2 +的反应介质中用洋地黄皂苷通透后，它们将介质中的Ca2 +浓度降低至亚微摩尔水平（0.05 - 0.1微摩尔）。1微摩尔FCCP的存在使这些通透细胞摄取Ca2 +的初始速率降低约75%。这种可能由内质网介导的对FCCP不敏感的Ca2 +摄取被500微摩尔钒酸盐完全抑制。另一方面，当存在钒酸盐而非FCCP时，Ca2 +积累的初始速率降低约25%，Ca2 +设定点增加至0.7微摩尔。在钒酸盐存在下检测到的Ca2 +摄取对琥珀酸的依赖性以及对FCCP和钌红的敏感性表明，这种摄取可能是由线粒体进行的。添加Ca2 +导致线粒体膜电位对钌红敏感的降低进一步支持了这一解释。抗利什曼原虫阳离子药物喷他脒和WR - 6026也能迅速使杜氏利什曼原虫前鞭毛体的线粒体内膜电位崩溃。
