Search

The effect of sublethal concentrations of potassium dichromate and fenitrothion on sodium-leucine cotransport in brush border membrane vesicles from Chironomus riparius larvae has been investigated. Exposure to potassium dichromate and fenitrothion caused a dose- and time-dependent inhibition of leucine uptake. Transport inhibition is easily detectable at doses 100-fold lower than LD50. Kinetic experiments showed that inhibition was mainly caused by a decrease of the Vmax (680 ± 53 vs. 382 ± 23 and 555 ± 27 nmol/15s/mg protein in control and exposed larvae to K2Cr2O7 and fenitrothion, respectively). Inhibition is possibly related to a variation of sodium ions permeability as evidenced by increased membrane lipid peroxidation. Appropriate control experiments ruled out that the observed differences could be due to changes in general features of membrane preparations. Transport inhibition observed in larvae exposed to potassium dichromate was accompanied by changes in ascorbate peroxidase and dehydroascorbate reductase activities, whereas those exposed to fenitrothion displayed an increase in transaminase activity. The possible value of leucine uptake as biochemical biomarker is briefly discussed. Arch. Insect Biochem. Physiol. 55:90–101, 2004. © 2004 Wiley-Liss, Inc.

Changes in enzyme activities, metabolite concentrations, and membrane transport activity underlying the Chironomus riparius larvae adaptive response to anoxia were investigated. Trehalose, malate, and aspartate degradation and alanine accumulation were recorded. During anoxia exposure, there was a boost of antioxidant defenses as shown by an increase of the specific activity of the enzymes catalase, glutathione-S-transferase, glutathione peroxidase, glutathione-synthase, malic enzyme, and NADP-dependent isocitrate dehydrogenase. The ratio, glutathione reduced over glutathione oxidized, decreased. Except for alanine and catalase, the parameters return to their basal value when larvae are transferred to normoxic conditions. To test whether antioxidant defenses had protective effects on membrane functionality, L-leucine uptake into brush border membrane vesicles and membrane lipid peroxidation was measured. No difference between membranes prepared from larvae exposed to anoxia and control larvae was found. The amino acid alanine, when present inside the vesicles, trans-stimulated leucine uptake. This effect could represent a mechanism to stimulate amino acid uptake and catabolism in vivo when free alanine concentration increases during hypoxic periods.

Leucine transport across brush border membrane vesicles prepared from four insect species common to European freshwater streams has been characterized. The species studied were: Ephemera danica (Ephemeroptera: Ephemeridae), Isoperla grammatica (Plecoptera: Perlodidae), Hydropsyche pellucidula (Trichoptera: Hydropsychidae), and Hybomitra bimaculata (Diptera: Tabanidae). The transport differed among the studied taxa for several features, including pH and sodium dependence, substrate affinity and specificity, and efficiency. In H. pellucidula and E. danica, leucine uptake was higher at pH 7.4 than at more alkaline or acidic pH values, whereas in I. grammatica and H. bimaculata, the uptake was rather constant when pH varied from 5.0 to 7.4, then strongly decreased at pH 8.8. All but E. danica displayed a transient intravescicular leucine accumulation in the presence of sodium, suggesting the existence of a cation-leucine symport mechanism. The sodium dependence ranged according to the following order: H. pellucidula > I. grammatica > H. bimaculata > E. danica. Moreover, in H. pellucidula and I. grammatica, the sodium-dependence was stronger at pH 8.8 than at pH 7.4. In E. danica, leucine uptake was sodium-independent at all pH values. The highest value of Vmax (45.3 pmol·s–1·mg proteins–1) was in E. danica, which, however, displayed the lowest affinity (Km 137 μM) when compared to the kinetic parameters of other taxa. The Vmax and Km values were: 40 and 52.5, 32.1 and 12.5, and 4.5 and 230 for H. bimaculata, H. pellucidula, and I. grammatica, respectively. The obtained results are discussed within our current knowledge of amino acid transport systems in insects. Arch. Insect Biochem. Physiol. 63:110–122, 2006. © 2006 Wiley-Liss, Inc.

The freshwater scud Echinogammarus stammeri is a very abundant benthic crustacean in riverine systems of northern Italy. The molecular physiology of L-[3H]leucine transport across cell membranes of E. stammeri was investigated. At pH 7.4, the presence of a NaCl gradient stimulated the leucine transport rate, when compared with KCl gradient. By contrast, alkaline pH strongly inhibited transport activity. In NaCl medium at acidic pH, leucine uptake displayed a specific activity 2.1-fold lower than that measured at neutral pH. Analysis of the uptake as a function of external leucine concentration revealed saturation kinetics. Leucine uptake appeared broadly unspecific and it was much strongly inhibited by the non-polar amino acids phenylalanine, methionine, and BCH, as well as by hydrophobic hydroxyacids. These results revealed important differences with respect to amino acid transport systems described in marine crustaceans. Their relevance in the feeding adaptation of the scud is briefly discussed.

Benthic macroinvertebrates are an essential component of freshwater environments. The ecotoxicological risk of benthic communities, estimated through the assay of biochemical markers, can be used as an early warning signal for environment alterations. In this work the activities of a number of enzymes, regarded as potential biomarker of exposure to pollutants (catalase, acetylcholinesterase, glutathione-S-transferase) was determined in homogenates of whole organisms. Specimens were collected in the rivers Taro and Ticino, northern Italy, in stony bottom reaches in five and three stations respectively. The orders of aquatic Insects Diptera, Plecoptera, Odonata, Ephemeroptera and Trichoptera, the Crustacea Amphipoda, and Annelida Oligochaeta were examined. Additional enzymes, such as NADP-dependent malate dehydrogenase, isocitric dehydrogenase, glucose-6-phosphate dehydrogenase, alcohol dehydrogenase, and lactate dehydrogenase were considered in the study. Results emphasize significant differences among taxa concerning the specific activity of most enzymes.