InCatharanthus roseuscells, recent data have indicated that glyphosate uptake is mediated by a phosphate carrier for concentrations lower than 100 μM (F. Morinet al., Pestic. Biochem. Physiol.158,13, 1997). Evidence is presented in this study which indicates that two mechanisms interplay in the energization of cellular glyphosate uptake: (i) the classical proton/anion symport, as demonstrated by the increased glyphosate absorption when transport experiments were carried out with an imposed pH gradient, by buffering the medium at acidic pH values; and (ii) another mechanism independent of the formation of a pH gradient but involving the combined action of calcium and iron, which may be energized by the functioning of a redox pump at the plasmalemma level. Ca/Fe-dependent glyphosate uptake is not strictly related to the presence of calcium in the medium, since other bi- and trivalent cations such as lanthanum, in combination with Fe2+, also increase glyphosate absorption. In contrast, Fe2+, which plays a specific role in the increased glyphosate uptake, appears to be one of the major elements of the energization process. Experiments carried out with various chemical agents which decrease the availability of intracellular NAD(P)H as the electron donor of the plasmalemma redox pump, or using a competitive inhibitor of molybdate in electron transfer (Na2WO4), and an artificial electron acceptor (FeCN), suggest that transplasmalemma electron transport is involved both in Fe2+and in glyphosate uptake. Using the above data, we propose a model which explains the combined action of calcium and iron and the coupling reactions between the redox pump, iron uptake, and the cellular absorption of glyphosate.