Study on Plant Electric Potential Variation in Cucumber Growing under Water Stress

Transcription

1 Study on Plant Electric Potential Variation in Cucumber Growing under Water Stress LI Guochen 1, YU Haiye 2 1 Engineering college, Jinggangshan University, P.R.China, ; 2 College of Biological and Agricultural Engineering, Jilin University, P.R.China, guochen_li@163.com Abstract Action potentials (AP) and variation potentials (VP) were measured in four-week old cucumber under different watering disposals. External stimuli (e.g. changing temperature, wounding or chemical compounds, and the density of light) cause an action potential to be generated and change the variation potential from the original steady state level. The paper discussed that the effect of water stress on plant physiological signals (AP and VP) which were induced by environmental factors such as heat wounding, light and electrical stimulation. Under environmental stimulations, the effect of water stress on variations of plant potentials mainly focus on APs, water stress enlarged the amplitude of APs which reached 50-60mv and speed up the propagation speed of APs, which increased from 0.2m/s to 0.5mm/s. Key words Plant potentials, Variation potential (VP), Action potential (AP), Water stress 1 Introduction The most rapid methods of long-distance communication between plant tissues and organs are bioelectrochemical or electrophysiological signals [1], about 130 years ago, the bulk of experiments demonstrated the existence of electrical signals in plant [2,3]. Paszewski and Zawadzki concluded that AP of lupine possessed the same characteristic as the nerval potentials [4,5]. Houwink studied the conduction of excitation in Mimosa pudica, and considered that wounding could excite AP,no-wounding stimulated VP [6]. Then came the breakthrough-the paper by Wildon et al. showing that electrical signals were involved in turning on protease inhibitor (PIN) genes in tomato [7], and flame wound evoked rapid changes in membrane potential in distant tissue, while transport of chemical signals in the phloem took an hour or more, the signal they were monitoring was action potential (AP). Meantime, we have already known that a flame wound invariably evoke another type of electrical signal known as a variation potential (VP). Bratislav Stankovic and Eric Davies [8,9] also studied plant potentials induced proteinase inhibitor gene expression in tomato, and concluded that hot injure and electrical stimulation could trigger VPs and electrically the triggered APs were capable of evoking pin2 gene expression. In china, LOU Cheng-Hou has researched the transmitting and inducing of mechanism of plant potentials [10]. Until now, different measuring methods and materials have been used, and many papers has mainly focused on discussing the responses evoked by external stimuli (mechanical or chemical) in upper ground parts of plant, but rarely talking about the effect of water deficit on plant electrophysiology. At present, the research on water-saving based on the feedback of plant physiology, such as leaf transpiration, the evolution of stem diameter or plant water potential, have been developing. This paper provided a way not only to study the change of electrical signals induced by water stress, but also to show the role of electrical signals in diagnosing water deficit of plant. 2 Materials and method 2.1 Plants The cucumber (Chang-Qing 1#, Jilin Province) is grown in a greenhouse for days at 20-30, and those with similar height (about 12cm) and appearance are selected for the experiments and transferred into plant growing chamber which well is shielded, where the air-temperature is kept at 22-25, and the air-humidity is 40-50%. The plants are illuminated with white fluorescent lights furnishing about µmol/m 2 s -1 which is active radiation at plant level. The soil was sandy soil, and there are two disposals for plant water status, namely well-irrigated plant (Soil Moisture: 22-25%) and 560

2 water stressed plant (Soil Moisture: 11-13%). 2.2 Electric potential Measurements The experimental set-up for measuring the AP and VP is shown in Figure1 [11,12]. An IBM-compatible microcomputer with S series simultaneous sampling Multifunction DAQ-NI PCI-6143 is interfaced through a multiplexed screw terminal accessory board with 0.2 mm non-polarizable reversible Ag/AgCl electrodes and used to record the digital data. Based on the property of the plant electrical potentials [13], being of biological information, the resistance of plant s signals source is very high (about Ω). Therefore, electrodes outputs are passed through a custom-made high-impedance (more than 1010Ω) instrumental amplifier and low-pass filter (for minimizing the power-line interference-50hz), the frequency of scanning is 4000 samples per second. Ag/AgCl electrodes are inserted into the plant at different position, which is for long-term monitoring. The reference electrode is usually inserted in the stem or in the root of a cucumber, and the working electrode is inserted in the leaf or the upper part of the stem, differential input between electrodes can well avoid co-module noise. 2.3 Application of Stimuli The plant potential belongs to ultra-weak signal; generally, it can be evoked by external stimuli [2]. In the experiment, hot wounding, light and electrical stimulation are taken for inducing AP and VP. As for hot wounding, plants are locally wounded on leaf 3 (next-to-youngest) by passing a lit match for about 3s underneath a region about 3-4cm 2 ; for light stimulus, the intensity of natural light in plant growing chamber is about 400µmol/m 2 s -1, the strong light is PAR=1200µmol/m 2 s -1, The cucumber s upper-leaf is illuminated by turns light on (strong light) and off (nature light); for electrical stimulus, a custom-made generator (voltage divider) furnishing a squared DC pulse applied for about 3s between a pair of inserted silver electrodes spaced about 1cm apart, the stimulating voltage is about 3v. 3 Results and Discussion Figure 1 Arrangement for Measuring Plant Potentials The generation and propagation of action potentials and electrical impulses between the tissues can be measured by reversible non-polarizable electrodes. Following insertion of the electrodes, the plants are allowed to rest until a stable potential difference is obtained between the measuring and reference electrodes. Insertion of electrodes in plants evokes slow fluctuations of the variation potential in the phloem and action potential across the stem, the resting potential stabilizes and action potentials disappear after approximately 1 hour. 3.1 Hot wounding Hot wounding can evoke the generation and propagation of AP (Bratislav Stankovic, 1996). Similar plants are given a mild flame treatment, the cucumber plants generate complex electrical responses, 561

3 these signals are slow waves which have been called VPs (Malone and Stankovic, 1991) [14], and the VPs are accompanied by some spikes which are putative APs. The plant potentials are measured as shown in Figure 2 and Figure 3, the datum in Figure 2 is about the cucumbers under well-watered, the figure 3 about the cucumbers under water stress. Generally, the VP change slowly, AP fluctuates very quickly [13]. Hot wounding can evoke the VP and AP in figure 2 and figure 3, the changes of VP in figure 2 is similar with that in figure 3, its variation ranges within 50-60mv. The APs are very different between two water disposal plants. As for well-watered cucumber in Figure 2, the magnitude of the AP is just about 3mv, its duration time of s; as for water stressed cucumber in Figure 3, the magnitude of AP can reach 10mv. Furthermore, the AP in Figure 2 ordinarily lasts about 2 minutes; But in Figure 3, the AP is transitorily excited and it appear only 4~6 waves. Figure 2 Potentials in Cucumber under Well-watered Figure 3 Potentials in Cucumber under Water Stress 3.2 Light stimulation In this experiment, the plant cucumber s upper-leaf is illuminated by light in turn. The changes of potentials are measured in Figure 4 and Figure 5. Figure 4 indicates the successive electrical signals of plant under well-watered, Figure 5 is for water-stressed plant. On the horizontal multi-line, the strong indicates light on whose intensity is about 1200µmol/m 2 s -1, the other part indicates light off. When the light changes from nature to strong status, the waves in Figure 4 and Figure 5 can clearly show the variation of potentials. In Figure 4, the range of VPs change is about 15mv; the duration of APs is between 0.005s, its positive and negative spikes appear during the measurement, this phenomena is explain by Volkov A.G. and R.A.Haack.(1995), the action potential propagates along the stem, and it reaches the reference electrode after some millisecond, which gives a mirror image of the potential peak with an opposite sign from that of the first peak. The speed of AP s propagation is measured as the distance between Ag/AgCl electrodes divided by time between positive and negative peaks of action potentials, and it is about 200mm/s (10mm/0.05s). In Figure 5, the range of VPs change is not more than 5mv, but the propagating speed of APs are very rapid, it can reach 500mm/s (10mm/0.02s). Figure 4 AP in Cucumber under Well-watered Figure 5 AP in Cucumber under Water stress 562

4 3.3 Electrical stimulation The experiment show that electrical stimulation can make well-watered cucumber evoke slow waves (VPs), whose magnitude is about 5mv, but for water-stressed plants, no clear VPs generate. According to previously published work using sunflower plants (Bratislav Stankovic, 1998) [15], electrical stimulus induce sunflowers to evoke action potentials, there are no visible action potentials occur during the experiment, in order to analyze the characteristics of the electrical stimulus-generated APs, multi-resolution wavelet analysis is taken for decomposing signal. A wavelet transformation is local transformation of the time and the frequency, it can withdraw strange of variety composition of the signals, well localized in both time and frequency domains. Figure 6 is the result of four-level wavelet decomposition based on basis functions-coif4. A, B, C and D are the singular point in waves; the details-d2 can reflect the valuable information of singular point. Figure 7 shows the enlarged signals which is at A, B, C and D, it is obvious that there are obvious action potentials, which amplitude is about 300µV, its period is about s. Therefore, the action potentials are evoked, but the changed range is not more than 1mv, which is too slight to be seen. Figure 6 The Details of Four-level Wavelet Decomposition of Plant Potentials Figure 7 Enlarged Signals in Singular Point A, B, C and D 563

5 4 Conclusions Environmental factors could induce plant electrical signals. When cucumber plants are stimulated by hot wounding, the AP and VP can be evoked, the changes of APs have clear differences in plants under two water disposals, as for well-watered cucumber, its amplitude is about 3mv, but the plant under water stress could generate AP whose amplitude could reach 50-60mv. When the intensity of light increases, there occur evidently spiking electrical signals-aps, the speed of propagation in plant under water stress is about 500m/s, which is very faster than that that in the well-watered plant (just 200mm/s). Electrical stimulation just evoke slight plant potentials, there are no clear difference between cucumber plants under different water disposals. Acknowledgments The financial support from the Natural Science Foundation of JiLin Province (No ). References [1] A.G.Volkov, R.A.Haack. Insect-induced bioelectrochemical signals in potato plants[j]. Bioelectrochemistry and Bioenergetics, 37(1995), p55~60 [2] Eric Davies. New functions for electrical signals in plants[j]. New Phytologist, 161(2004), p607~610 [3] Alexander G. Volkov, Green plants: electrochemical interfaces[j], Journal of Electroanalytical Chemistry, 483(2000), p150~156 [4] Paszewski A, Zawadzki T. Action potentials in Lupinus angustifolius shoots 3 Determinatio of the refractory periods[j]. J Exp Bot, 27(1976), p369~374 [5] Zawadzki T, Trebacz K. Action potentials in Lupinus angustifolius 6 Propagation of action potential in the stem after the application of mechanical block[j]. J Exp Bot, 33(1982), p100~110 [6] Howink A L. The conduction of excition in Mimosa pudica[j]. Rec Trav Bot Neel, 32(1935), p51~91 [7] Wildon D C, Thain JF, Minchin PEH, et al. Electrical signalling and systemic proteinase inhibitor induction in the wounded plant. Nature, 360(1992), p62~65 [8] Bratislav Stankovic, Eric Davies. Both action potentials and variation potentials induce proteinase inhibitor gene expression in tomato, FEBS Letter, 390(1996), p275~279 [9] Stankovic, E. Davies. The wound response in tomato involves rapid growth and electrical responses, systemically up-regulated transcription of proteinase inhibitor and calmodulin and down- regulated translation[j]. Plant & Cell Physiology, 39(1998),pp268~274 [10] Lou Chenghou, Hua Baoguang. Plant signaling system[j]. Chinese Bulletin of Life Sciences. 12(2000), p49~50 [11] Li Guochen, Yu Haiye, Ma Chenglin. Development of measuring system for weak electrical potential in plants and its application[j]. Transactions of the CSAE, 21(2005), p6~10 [12] Li Guochen, Yu Haiye,Li Qiang-zheng. Discussion on method of measuring and proceeding plant physiological signals [J]. J of agricultural mechanization research, 6(2006), p145~148 [13] Zhao Zikai. A study on ultra-weak electronic signal in plants[j]. Journal of China Institute of Metrology, 03(2002), p243~257 [14] Malone M, Stankovic B. Surface potentials and hydraulic signals in wheat leaves following localized wounding by heat[j]. Plant Cell Environ, 14(1991), p431~436 [15] Bratislav Stankovic, Daniel L.Witter, Tadeusz Zawadzki and Eric Davies. Action potentials and variation potentials in sunflower: An analysis of their relationships and distinguishing characteristics[j]. Physiologia Plantarum, 103(1998), p51~58 564