Effects of Feeding Spodoptera littoralis on Lima Bean Leaves. I. Membrane Potentials, Intracellular Calcium Variations, Oral Secretions, and Regurgitate Components

Abstract

Membrane potentials (V m ) and intracellular calcium variations were studied in Lima bean (Phaseolus lunatus) leaves when the Mediterranean climbing cutworm (Spodoptera littoralis) was attacking the plants. In addition to the effect of the feeding insect the impact of several N-acyl Glns (volicitin, N-palmitoyl-Gln, N-linolenoyl-Gln) from the larval oral secretion was studied. The results showed that the early events upon herbivore attack were: a) a strong V m depolarization at the bite zone and an isotropic wave of V m depolarization spreading throughout the entire attacked leaf; b) a V m depolarization observed for the regurgitant but not with volicitin {N-(17-hydroxy-linolenoyl)-Gln} alone; c) an enhanced influx of Ca 21 at the very edge of the bite, which is halved, if the Ca 21 channel blocker Verapamil is used. Furthermore, the dose-dependence effects of N-acyl Gln conjugatestriggered influx of Ca 21 studied in transgenic aequorin-expressing soybean (Glycine max) cells, showed: a) a concentrationdependent influx of Ca 21 ; b) a configuration-independent effect concerning the stereochemistry of the amino acid moiety; c) a slightly reduced influx of Ca 21 after modification of the fatty acid backbone by functionalization with oxygen and; d) a comparable effect with the detergent SDS. Finally, the herbivore wounding causes a response in the plant cells that cannot be mimicked by mechanical wounding. The involvement of Ca 21 in signaling after herbivore wounding is discussed.

Effects of Feeding Spodoptera littoralis on Lima Bean Leaves. III. Membrane Depolarization and Involvement of Hydrogen Peroxide

“…6A). As reported earlier (Maffei et al, 2004), HW tissues showed a more positive V m with respect to MD leaves, and the response to H 2 O 2 was a net lowering of the response (Fig. 6B), even though a direct comparison of MD and HW V m responses to H 2 O 2 showed some sort of threshold lowering.…”

“…(Maffei et al, 2004). Preincubation of lima bean leaves with 100 mM Verapamil was found to completely suppress H 2 O 2 -dependent V m depolarization in MD leaves (Fig.…”

“…One of the early signals in plant-herbivore interaction is the variation in the V m (Maffei et al, 2004). Having assessed the dose-response effect of H 2 O 2 release on [Ca 21 ] cyt , and because of the possible unbalance of charges across the plasma membrane depending on [Ca 21 ] cyt variations, we analyzed changes in V m when increasing H 2 O 2 concentrations (from 0.3 mM to 24 mM) were applied to MD and HW tissues.…”

“…Price et al, 1994;Low and Merida, 1996;Cazalé et al, 1998). Electrophysiological studies revealed the existence of plasma membrane Ca 21 -permeable channels activated by membrane depolarization or hyperpolarization in response to environmental stimuli (for review, see White, 2000;Sanders et al, 2002) and at the very edge of herbivore wounding in plantinsect interactions (Maffei et al, 2004). ROS can interact with ion channel activity, and oxidase-dependent electron transfer could drive a transmembrane potential (V m ) depolarization; both effects lead to channel activation.…”

“…By taking advantage of the model system consisting of Spodoptera littoralis larvae feeding on lima bean (Phaseolus lunatus) leaves (Maffei et al, 2004;Mithö fer et al, 2005), we used several microscopic, physiological, biochemical, and molecular biological tools to evaluate the role of H 2 O 2 in plant-herbivore interactions. Thus, we investigated (1) the cellular and subcellular H 2 O 2 localization in mechanically damaged (MD) and S. littoralis herbivore-wounded (HW) tissues by means of fluorescence light microscopy, confocal laser-scanning microscopy (CLSM), and transmission electron microscopy; (2) the effect of H 2 O 2 on [Ca 21 ] cyt using CLSM and apoaequorin-transformed plants; (3) the effect of H 2 O 2 on V m and cytosolic calcium ([Ca 21 ] cyt ) variations using specific ion channel inhibitors in MD and HW tissues; and (4) the enzyme activity and quantitative gene expression of some ROS scavenging systems.…”