Zbigniew Burdach

Effect of temperature on growth, proton extrusion and membrane potential in maize (Zea mays L.) coleoptile segments

The effects of temperature (5–45°C) on endogenous growth, growth in the presence of either indoleacetic acid (IAA) or fusicoccin (FC), and proton extrusion in maize coleoptile segments were studied. In addition, membrane potential changes at some temperatures were also determined. It was found that in this model system endogenous growth exhibits a clear maximum at 30°C, whereas growth in the presence of IAA and FC shows the maximum value in the range 30–35°C and 35–40°C, respectively. Simultaneous measurements of growth and external medium pH indicated that FC at stressful temperatures was not only much more active in the stimulation of growth, but was also more effective in acidifying the external medium than IAA. Also the addition of either IAA or FC to the bathing medium at 30 and 40°C did not change the kinetic characteristic of membrane potential changes observed for both substances at 25°C. However, the increased temperature significantly decreased IAA and FC-induced membrane hyperpolarization. IAA in the incubation medium, at 10°C, brought about additional membrane depolarization (apart from the one induced by low temperature). In contrast to IAA, FC at 10°C caused gradual repolarization of membrane potential, which correlated with both FC-induced growth and FC-induced proton extrusion. A plausible interpretation for temperature-induced changes in growth of maize coleoptile segments is that, at least in part, these changes were mediated via a PM H+-ATPase activity.

Interactive effects of temperature and heavy metals (Cd, Pb) on the elongation growth in maize coleoptiles.

The effect of Cd and Pb on endogenous and IAA-induced elongation growth and medium pH of maize coleoptile segments incubated at 20, 25 and 30 °C was studied. It was found that the elongation of coleoptile segments and proton extrusion increased with the temperature and reached its maximum at 30 °C. For Cd, the maximal inhibition of endogenous and IAA-induced growth as well as medium acidification of coleoptile segments was observed at 25 °C. Meanwhile, Pb, irrespective of the temperature, diminished the growth of the segments by ca. 20%, increasing the acidification of the incubation medium. It was also found that in contrast to Cd, Pb accumulation in maize coleoptile segments did not correlate with temperature. The results suggest that the toxic effect of Cd on elongation growth of coleoptile segments is connected with the decrease of the PM H(+)-ATPase activity and probably with Cd-induced high acivity of IAA oxidase, whereas the effect of Pb did not depend on activity of any of the enzymes.

Effect of Trimethyltin Chloride on Slow Vacuolar (SV) Channels in Vacuoles from Red Beet (Beta vulgaris L.) Taproots

In the present study, patch-clamp techniques have been used to investigate the effect of trimethyltin chloride (Met3SnCl) on the slow vacuolar (SV) channels in vacuoles from red beet (Beta vulgaris L.) taproots. Activity of SV channels has been measured in whole-vacuole and cytosolic side-out patch configurations. It was found that addition of trimethyltin chloride to the bath solution suppressed, in a concentration-dependent manner, SV currents in red beet vacuoles. The time constant, τ, increased significantly in the presence of the organotin. When single channel activity was analyzed, only little channel activity could be recorded at 100 μM Met3SnCl. Trimethyltin chloride added to the bath medium significantly decreased (by ca. threefold at 100 μM Met3SnCl and at 100 mV voltage, as compared to the control medium) the open probability of single channels. Single channel recordings obtained in the presence and absence of trimethyltin chloride showed that the organotin only slightly (by <10%) decreased the unitary conductance of single channels. It was also found that Met3SnCl significantly diminished the number of SV channel openings, whereas it did not change the opening times of the channels. Taking into account the above and the fact that under the here applied experimental conditions (pH = 7.5) Met3SnCl is a non-dissociated (more lipophilic) compound, we suggest that the suppression of SV currents observed in the presence of the organotin results probably from its hydrophobic properties allowing this compound to translocate near the selectivity filter of the channel.

Effect of trimethyllead chloride on slowly activating (SV) channels in red beet (Beta vulgaris L.) taproots

The patch-clamp technique was used to examine the effect of trimethyllead chloride (Met(3)PbCl) on SV channel activity in red beet (Beta vulgaris L.) taproot vacuoles. It was found that in the control bath the macroscopic currents showed the typical slow activation and a strong outward rectification of the steady-state currents. An addition of Met(3)PbCl to the bath solution blocked, in a concentration-dependent manner, SV currents in red beet vacuoles. The time constant τ increased several times in the presence of 100 μM trimethyllead chloride at all voltages tested. When single channel properties were analyzed, only little channel activity could be recorded in the presence of 100 μM Met(3)PbCl. Trimethyllead chloride decreased significantly (by about one order of magnitude) the open probability of single channels. The recordings of single channel activity obtained in the presence and absence of Met(3)PbCl showed that organolead only slightly (by ca. 10%) decreased the unitary conductance of single channels. It was also found that Met(3)PbCl diminished significantly the number of SV channel openings, whereas it did not change the opening times of the channels. Taken together, these results suggest that Met(3)PbCl binding site is located outside the channel selectivity filter.

Fusicoccin counteracts inhibitory effects of high temperature on auxin-induced growth and proton extrusion in maize coleoptile segments

Plant growth and development are tightly regulated by both plant growth substances and environmental factors such as temperature. Taking into account the above, it was reasonable to point out that indole-3-acetic acid (IAA), the most abundant type of auxin in plants, could be involved in temperature-dependent growth of plant cells. We have recently shown that growth of maize coleoptile segments in the presence of auxin (IAA) and fusicoccin (FC) shows the maximum value in the range 30-35˚C and 35-40˚C, respectively. Furthermore, simultaneous measurements of growth and external medium pH indicated that FC at stressful temperatures was not only much more active in the stimulation of growth, but was also more effective in acidifying the external medium than IAA. The aim of this addendum is to determine interrelations between the action of IAA and FC (applied together with IAA) on growth and medium pH of maize coleoptile segments incubated at high temperature (40˚C), which was optimal for FC but not for IAA. Addendum to: Karcz W, Burdach Z. Effect of temperature on growth, proton extrusion and membrane potential in maize (Zea mays L.) coleoptile segments. Plant Growth Regul 2007; 52:141-50.

Single and combined effects of Cd and Pb on the growth, medium pH, membrane potential and metal contents in maize (Zea mays L.) coleoptile segments

The mechanisms of the toxic effects of Cd and Pb on plant cell growth are still poorly understood. In particular, little is known about their interactive effects, which usually occur in the environment. Moreover, the data that do exist in the literature are controversial. This study describes experiments that were performed with maize (Zea mays) coleoptile segments, which is a classical model system for studies of plant cell elongation growth. Cadmium and lead, which were added at 0.1 mM, reduced the endogenous and IAA-induced elongation growth of maize coleoptile cells. When both metals were added together or in sequence, their effect on IAA-induced growth was more toxic. The medium pH changes, which were measured simultaneously with growth, indicated that while Pb stopped IAA-induced proton extrusion, Cd only partially diminished it. Although Cd was generally more accumulated than Pb in the maize coleoptile segments, when IAA was added together with Pb, it significantly increased the accumulation of the metal. The short-term electrophysiological experiments showed that the addition of Cd caused the depolarisation of the membrane potential (Em), whereas Pb caused membrane hyperpolarisation. In the long-term electrophysiological experiments, it was found that the Cd-induced Em changes are complex. In conclusion, these results suggest that the effects of Cd and Pb as well as their combination on the elongation growth of maize coleoptile cells and the accumulation of the metals result, among others, from different ionic mechanisms by which each metal change the membrane potential of the cells.