Review: 98 refs.

Abstract:Interrelations between the action of acetylcholine (ACh) and cadmium ions (Cd 2+ ) on bioelectrogenesis of Nitellopsis obtusa cells were investigated. We analyzed repetitively triggered action potentials (AP), their reproducibility, shape and dynamics of membrane potential after AP induction. ACh significantly increased membrane permeability only at high concentrations (1 mM and 5 mM). Repolarisation level of action potential after the first stimulus was much more positive in all cells treated with ACh as compared to the control. Differences of membrane potentials between points just before the first and the second stimuli were 23.4±.0 mV (control); 40.4±5.9 mV (1 mM ACh solution) and 57.7 ± 8.5 mV (5 mM ACh solution). Cd 2+ at 20 μM concentration was examined as a possible inhibitor of acetylcholinesterase (AChE) in vivo. We found that cadmium strengthens depolarizing effect of acetylcholine after the first stimulus. The highest velocity of AP repolarization was reduced after ACh application and Cd 2+ strengthened this effect. There were no differences in dynamics of membrane potential after repetitively triggered action potentials in ACh or ACh and Cd 2+ solutions. This shows that cadmium in small concentration acts as inhibitor of acetylcholinesterase.

Action potentials (AP) of characean cells were the first electrical transients identified in plants. APs provide information about plethora of environmental cues. Salinity stress is critical for plants and impacts on excitability. The AP of brackish Characeae Nitellopsis obtusa , obtained in artificial pond water (APW) and under osmotic stress of 90 or 180 mM sorbitol APW or saline stress of 50 or 100 mM NaCl APW, were simulated by the Thiel-Beilby model (Beilby and Al Khazaaly, 2016 ). The model is based on a paradigm from animal systems, featuring the second messenger inositol 1,4,5-triphosphate (IP 3 ) mediating the opening of Ca 2+ channels on internal stores. In plants the IP 3 receptors have not been identified, so other second messengers might translate the threshold plasma membrane depolarization to Ca 2+ release. The increased Ca 2+ concentration in the cytoplasm activates Cl − channels, which lead to the depolarizing phase of the AP. The repolarization to normal resting potential difference (PD) results from the Ca 2+ being re-sequestered by the Ca 2+ pumps, the closure of the Cl − channels, efflux of K + through the depolarization-activated outward rectifier channels and the continuing activity of the proton pump. The Nitellopsis AP form is longer in APW compared to that of Chara , with more gradual repolarization. The tonoplast component of the AP is larger than that in Chara australis . The plasma membrane AP is prolonged by the exposure to saline to a “rectangular” shape, similar to that in Chara . However, the changes are more gradual, allowing more insight into the mechanism of the process. It is possible that the cells recover the original AP form after prolonged exposure to brackish conditions. Some cells experience tonoplast APs only. As in Chara , the proton pump is transiently inhibited by the high cytoplasmic Ca 2+ and gradually declines in saline media. However, if the cells are very hyperpolarized at the start of the experiment, the pump inhibition both by the AP and by the saline medium is mitigated. The model parameters and their changes with salinity are comparable to those in Chara .

The Nitellopsis obtusa (N.A.Desvaux) J.Groves cell provides a model system for complex investigation of instantaneous effects of various biologically active compounds (BC) on the generation of plant bioelectrical signals in vivo. Experimental evidence using multiple electrical signals as biomarkers of the effects of BC (acetylcholine, asparagine, glutamate, nicotine, aluminium, nickel and cadmium ions) is provided. The effect of BC on membrane transport systems involved in the cell excitability were tested by current clamp, voltage clamp and patch clamp methods. Membrane potential (MP) alterations and action potential (AP) patterns in response to BC were shown to represent the cell state. High discretisation frequency allows precise, high time resolution analysis of real-time processes measuring changes in excitation threshold, AP amplitude and velocity of repolarisation values after application of BC indicating the effect on ion channels involved in AP generation. Application of voltage clamp revealed that changes in AP peak value were caused not only by increment in averaged maximum amplitude of the Clcurrent, but in prolonged Clchannels' opening time also. The cytoplasmic droplet can serve as a model system in which the effects of BC on single tonoplast ion channel can be studied by patch clamping. Investigation of electrical cell-to-cell communication revealed evidence on the electrical signal transduction through plasmodesmata.

The role of acetylcholine (ACh) as a signalling molecule in plants was investigated using a model system of Characeae cells. The effect of ACh on conductance of K+ channels in Nitella flexilis cells and on the action potential generation in Nitellopsis obtusa cells after H+-ATPase inhibition, where repolarization occurs after the opening of outward rectifying K+ channels, was investigated. Voltage-clamp method based on only one electrode impalement was used to evaluate the activity of separate potassium ion transport system at rest. We found that ACh at high concentrations (1 mM and 5 mM) activates K+ channels as the main membrane transport system at the resting state involved in electrogenesis of Characeaen membrane potential. We observed that ACh caused an increase in duration of AP repolarization of cells in K+ state when plasmalemma electrical characteristics are determined by large conductance K+ channels irrespective of whether AP were spontaneous or electrically evoked. These results indicate interference of ACh with electrical cellular signalling pathway in plants.

Background In this review, we summarize data concerning action potentials (APs) – long-distance electrical signals in Characean algae and liverworts. These lineages are key in understanding the mechanisms of plant terrestrialization. Liverworts are postulated to be pioneer land plants, whereas aquatic Charophytes are considered the closest relatives to land plants. The drastic change of the habitat was coupled with the adaptation of signalling systems to the new environment. Scope APs fulfil the “all-or-nothing” law, exhibit refractory periods, and propagate with a uniform velocity. Their ion mechanism in the algae and liverworts consists of a Ca 2+ influx (from external and internal stores) followed by/coincident with a Cl - efflux, which both evoke the membrane potential depolarization, and a K + efflux leading to the repolarization. The molecular identity of ion channels responsible for these fluxes remains unknown. Publication of the Chara braunii and Marchantia polymorpha genomes opened new possibilities for studying the molecular basis of APs. Here we present the list of genes which can participate in AP electrogenesis. We also point out the differences between these plant species, e.g. the absence of Ca 2+-permeable glutamate receptors (GLRs) and Cl --permeable SLAC1 channel homologues in the Chara genome. Both these channels play a vital role in long-distance signalling in liverworts and vascular plants. Among the common properties of APs in liverworts and higher plants is the duration of APs (dozens of seconds) and the speed of propagation (mm s -1), which are much slower than in the algae (seconds, and dozens of mm s -1, respectively). Conclusions Future studies with combined application of electrophysiological and molecular techniques should unravel ion channel proteins responsible for AP generation, their regulation, and transduction of those signals to physiological responses. This should also help to understand the adaptation of the signalling systems to the land environment and further evolution of APs in vascular plants.

Inhibitors of human two-pore channels (TPC1 and TPC2), i.e., verapamil, tetrandrine, and NED-19, are promising medicines used in treatment of serious diseases. In the present study, the impact of these substances on action potentials (APs) and vacuolar channel activity was examined in the aquatic characean algae Nitellopsis obtusa and in the terrestrial liverwort Marchantia polymorpha. In both plant species, verapamil (20–300 µM) caused reduction of AP amplitudes, indicating impaired Ca2+ transport. In N. obtusa, it depolarized the AP excitation threshold and resting potential and prolonged AP duration. In isolated vacuoles of M. polymorpha, verapamil caused a reduction of the open probability of slow vacuolar SV/TPC channels but had almost no effect on K+ channels in the tonoplast of N. obtusa. In both species, tetrandrine (20–100 µM) evoked a pleiotropic effect: reduction of resting potential and AP amplitudes and prolongation of AP repolarization phases, especially in M. polymorpha, but it did not alter vacuolar SV/TPC activity. NED-19 (75 µM) caused both specific and unspecific effects on N. obtusa APs. In M. polymorpha, NED-19 increased the duration of repolarization. However, no inhibition of SV/TPC channels was observed in Marchantia vacuoles, but an increase in open probability and channel flickering. The results indicate an effect on Ca2+ -permeable channels governing plant excitation.

Aluminium induced membrane potential (E m ) changes and potential changes during repolarization phase of the action potential (AP) in the internodal cells of Nitellopsis obtusa after blocking H + -ATPase activity by DCCD were investigated. Micromolar concentrations of DCCD are sufficient togive complete and irreversible inhibition of proton pumping. The membrane potential was measured by conventional glass-microelectrode technique. We found that the half-amplitude pulse duration differs significantly between standard conditions, after DCCD application, and after H + -ATPase blocking and subsequent Al 3+ treatment: 4.9, 7.7 and 17.2 seconds, respectively. We propose that in the short term (2 hours) treatment of Al 3+ , the decrease in membrane potential was compensated for by H + -ATPase activity. Blocking H + -ATPase activity by DCCD can enhance the influence of Al 3+ on the bioelectrical activity of cell membranes.