Respiration, oxidative phosphorylation. and the corresponding changes in membrane potential (deltapsi) of Trypanosoma cruzi epimastigotes grown either in liver infusion-tryptose (LIT) or brain heart infusion (BHI) culture medium were assayed in situ using digitonin to render their plasma membrane permeable to succinate, ADP, safranine O, and other small molecules. When the cells were permeabilized with 64 microM digitonin, a concentration previously 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 by 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 of oxygen consumption by these permeabilized cells indicated that the rate of basal respiration, which was similar in both cell types, was significantly decreased by 64 microM digitonin. Addition of ADP to the permeabilized cells grown in LIT medium promoted an oligomycin-sensitive transition from resting to phosphorylating respiration in contrast to the cells grown in BHI medium, whose respiration decreased steadily and did not respond either to ADP or CCCP. Titration of the cells grown in BHI medium with different digitonin concentrations indicated that their mitochondria have higher sensitivity to digitonin than those grown in LIT medium. Analysis of the sterol composition of epimastigotes grown in the two different media showed a higher percentage of cholesterol in total and mitochondrial extracts of epimastigotes grown in BHI medium as compared to those grown in LIT medium, suggesting the involvement of this sterol in their increased sensitivity to digitonin-permeabilization.

Acidocalcisomes are acidic vacuoles present in trypanosomatids that contain most of the cellular calcium. The data presented here demonstrate that Leishmania donovani acidocalcisomes possess a Na + /H + exchanger. 3,5-Dibutyl-4-hydroxytoluene, in the concentration range of 0^20 W WM, inhibited the Na + /H + exchanger, and strongly stimulated the activity of the vacuolar H + -ATPase responsible for vacuolar acidification. As occurs with Na + , the cationic anti-leishmanial drugs pentamidine, WR-6026, and chloroquine promoted a fast and extensive alkalization of the L. donovani acidocalcisomes.z 2000 Federation of European Biochemical Societies.

In this communication, we show that the plant uncoupling mitochondrial protein (PUMP) present in potato tuber mitochondria is induced by aging at 28³C and that this induction is strongly stimulated when the potato tubers are stored at low temperature (4³C). PUMP activity was detected by the degree of linoleic acid (LA)-induced ATP-sensitive mitochondrial uncoupling measured as a function of the decrease in membrane potential (v v8 8). The PUMP content was evaluated by immunoblot analysis using polyclonal antibodies raised against potato PUMP that specifically detected a 32 kDa band. In agreement with the effect of LA on v v8 8, the content of the 32 kDa band increased during storage and was stimulated by low temperature. These results support the proposed role of PUMP in plant thermogenesis and possibly in fruit ripening and senescence.z 1999 Federation of European Biochemical Societies.

The placenta acts a regulator of nutrient composition and supply from mother to fetus and is the source of hormonal signals that affect maternal and fetal metabolism. Thus, appropriate development of the placenta is crucial for normal fetal development. We investigated the effect of gestational protein restriction (GPR) on placental morphology and mitochondrial function on day 19 of gestation. Pregnant dams were divided into two groups: normal (NP 17 % casein) or low-protein diet (LP 6 % casein). The placentas were processed for biochemical, histomorphometric and ultrastructural analysis. The integrity of rat placental mitochondria (RPM) isolated by conventional differential centrifugation was measured by oxygen uptake (Clark-type electrode). LP animals presented an increase in adipose tissue and triacylglycerol and a decrease in serum insulin levels. No alterations were observed in body, liver, fetus, or placenta weight. There was also no change in serum glucose, total protein, or lipid content. Gestational protein restriction had tissue-specific respiratory effects, with the observation of a small change in liver respiration (~13 %) and considerable respiratory inhibition in placenta samples (~37 %). The higher oxygen uptake by RPM in the LP groups suggests uncoupling between respiration and oxidative phosphorylation. In addition, ultrastructural analysis of junctional zone giant cells from LP placenta showed a disorganized cytoplasm, with loss of integrity of most organelles and intense vacuolization. The present results led us to hypothesize that GPR alters placental structure and morphology, induces sensitivity to insulin, mitochondrial abnormalities and suggests premature aging of the placenta. Further studies are needed to test this hypothesis.

No abstract

No abstract

The ability of low concentrations (5^15 W WM) of longchain fatty acids to open the permeability transition pore (PTP) in Ca 2+ -loaded mitochondria has been ascribed to their protonophoric effect mediated by mitochondrial anion carriers, as well as to a direct interaction with the pore assembly [M.R. Wieckowski and L. Wojtczak, FEBS Lett. 423 (1998) 339^342]. Here, we have compared the PTP opening ability of arachidonic acid (AA) with that of carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) at concentrations that cause similar quantitative dissipation of the membrane potential (v v8 8) in Ca 2 -loaded rat liver mitochondria respiring on succinate. The initial protonophoric effects of AA and FCCP were only slightly modified by carboxyatractyloside and were followed by PTP opening, as indicated by a second phase of v v8 8 disruption sensitive to EGTA, ADP, dithiothreitol and cyclosporin A. This second phase of v v8 8 dissipation could also be prevented by rotenone or NAD(P)H-linked substrates which decrease the pyridine nucleotide (PN) oxidation that follows the stimulation of oxygen consumption induced by AA or FCCP. These results suggest that, under the experimental conditions used here, the PTP opening induced by AA or FCCP was a consequence of PN oxidation. Exogenous catalase also inhibited both AA-and FCCP-induced PTP opening. These results indicate that a condition of oxidative stress associated with the oxidized state of PN underlies membrane protein thiol oxidation and PTP opening.z 1999 Federation of European Biochemical Societies.

SUMMARYEnteric parasitosis remains an important public health problem in many areas around the world including in Brazil, and it is frequently associated with poverty and lack of sanitation facilities. Research carried out over the course of a year revealed that 96.6% (28/29) of children randomly selected from a 'landless farm workers' settlement in Araras, São Paulo, aged 4 -15 years, presented Giardia intestinalis cysts. After referral to the neighborhood Health Office, all the children received tinidazole, given as a single dose of 50 mg/kg and 12 months later, new fecal samples were collected and analyzed. Despite the low adherence to the study, a high percentage (64.3% -9/14) of the children remained positive for the parasite. This study showed a high positivity of giardiasis in child residents of the settlement, even after treatment; adults were not sensitized to the study and did not collected and/or deliver children fecal samples. The precarious living conditions are consistent with a high susceptibility to parasitic diseases, suggesting that the treatment of the infected individuals without identifying and eradicating the means of contamination is simply a palliative measure.