FIG. 5.

Effect of l-proline on ATP production and infectivity of T. cruzi metacyclic forms (G strain) subjected to nutritional stress. (A) Parasites previously maintained in PBS for 36 h were incubated for 1 h with glucose (1 mM), l-proline (0.75 mM or 3.0 mM), or l-glutamic acid (1 mM). After washings, the parasite ATP content and the ability to enter HeLa cells were examined. Intracellular parasite numbers were counted in 250 cells stained with Giemsa. A reference value of 100% was assigned to controls. Values represent the means ± standard deviations of the results of three independent experiments performed in duplicate. The differences in the levels of ATP content between 36-h-starved and l-proline-supplemented parasites were significant (P < 0.005). (B) Transport of glucose, l-proline, or glutamate was assayed by using radioactive compounds as described in Materials and Methods. At different time points after the addition of the radioactive reagent, the uptake was stopped by adding cold glucose, l-proline, or l-glutamic acid in PBS. Values represent the means ± standard deviations of the results of triplicate experiments. (C) Parasites were treated with inhibitors of energy production (rotenone [200 μM] and antimycin A [0.5 μM]) for 30 min, washed in PBS, and then used for cell invasion assays in the absence or presence of 1 mM glucose, 0.75 mM l-proline, or 1 mM l-glutamate. Intracellular parasite numbers were counted in 200 Giemsa-stained cells. Values represent the means ± standard deviations of the results of three independent experiments performed in duplicate.