Demystifying the Venus flytrap action potential.

All plants are electrically excitable, but only few are known to fire a well-defined, all-or-nothing action potential (AP). The Venus flytrap Dionaea muscipula displays APs with an extraordinarily high firing frequency and speed, enabling the capture organ of this carnivorous plant to catch small animals as fast as flies. The number of APs triggered by the prey is counted and serves as the basis for decisions within the flytrap's hunting cycle. The archetypical Dionaea AP lasts 1 s and consists of five phases: Starting from the resting state, an initial cytosolic Ca2+ transient is followed by depolarization, repolarization and a transient hyperpolarization (overshoot) before the original membrane potential is finally recovered. When the flytrap matures and becomes excitable, a distinct set of ion channels, pumps and carriers is expressed, each mastering a distinct AP phase.

Plant electrophysiology with conformable organic electronics: Deciphering the propagation of Venus flytrap action potentials.

Electrical signals in plants are mediators of long-distance signaling and correlate with plant movements and responses to stress. These signals are studied with single surface electrodes that cannot resolve signal propagation and integration, thus impeding their decoding and link to function. Here, we developed a conformable multielectrode array based on organic electronics for large-scale and high-resolution plant electrophysiology. We performed precise spatiotemporal mapping of the action potential (AP) in Venus flytrap and found that the AP actively propagates through the tissue with constant speed and without strong directionality. We also found that spontaneously generated APs can originate from unstimulated hairs and that they correlate with trap movement. Last, we demonstrate that the Venus flytrap circuitry can be activated by cells other than the sensory hairs. Our work reveals key properties of the AP and establishes the capacity of organic bioelectronics for resolving electrical signaling in plants contributing to the mechanistic understanding of long-distance responses in plants.

Cytokinins and auxins in organs of aquatic carnivorous plants: what do they reflect?

BACKGROUND AND AIMS: Aquatic carnivorous plants have typical rootless linear shoots bearing traps and exhibit steep physiological polarity with rapid apical growth. The aim was to analyse auxin and cytokinin metabolites in traps, leaves/shoots and shoot apices in several species of genera Aldrovanda and Utricularia to elucidate how the hormonal profiles reflect the specific organ functions and polarity. METHODS: The main auxin and cytokinin metabolites were analysed in miniature samples (>2 mg dry weight) of different organs of Aldrovanda vesiculosa and six Utricularia species using ultraperformance liquid chromatography coupled with triple quadrupole mass spectrometry. KEY RESULTS: Total contents of biologically active forms (free bases, ribosides) of all four main endogenously occurring cytokinin types were consistently higher in traps than in leaves in four Utricularia species with monomorphic shoots and/or higher than in shoots in two Utricularia species with dimorphic shoots. In Aldrovanda traps, the total content of different cytokinin forms was similar to or lower than that in shoots. In U. australis leaves, feeding on prey increased all cytokinin forms, while no consistent differences occurred in Aldrovanda. In four aquatic Utricularia species with monomorphic shoots, the content of four auxin forms was usually higher in traps than in leaves. Zero IAA content was determined in U. australis leaves from a meso-eutrophic site or when prey-fed. CONCLUSIONS: Different cytokinin and auxin profiles estimated in traps and leaves/shoots of aquatic carnivorous plants indicate an association with different dominant functions of these organs: nutrient uptake by traps versus photosynthetic function of traps. Interplay of cytokinins and auxins regulates apical dominance in these plants possessing strong polarity.

Searching for Optimal Substitute Habitats for Plants by Biological Experiments-A Case Study of the Endangered Species Aldrovanda vesiculosa L. (Droseraceae).

The selection of appropriate locations for the reintroduction of endangered plant species is an important process, because it usually influences the success of the conservation. The aim of this study was to select the optimal substitute habitats for Aldrovanda vesiculosa, taking into account the influence of physical-chemical factors (light intensity, temperature, pH, concentration of dissolved forms of nitrogen and cyanobacterial toxin microcystin-LR) on the efficiency of plant growth. Water analysis and field observations of the habitats of six lakes in Eastern Poland typified as potential substitute habitats for aldrovanda were carried out. The results of the experiments showed that both the concentration and the form in which nitrogen compounds are present in the environment were the factors limiting the growth rate and condition of plants. The second factor that caused the inhibition of aldrovanda growth was microcystin-LR. It was found that the habitat conditions in Lake Brzeziczno were within the ecological tolerance of the species. Particularly important was the low content of mineral compounds and the available forms of nitrogen and phosphorus in the water. Therefore, the probability of development of toxic cyanobacteria, the metabolites of which may affect the growth of A. vesiculosa, is also minimal.

Plant-inspired multi-stimuli and multi-temporal morphing composites.

Plants are inspiring models for adaptive, morphing systems. In addition to their shape complexity, they can respond to multiple stimuli and exhibit both fast and slow motion. We attempt to recreate these capabilities in synthetic structures, proposing a fabrication and design scheme for multi-stimuli and multi-temporal responsive plant-inspired composites. We leverage a hierarchical, spatially tailored microstructural and compositional scheme to enable both fast morphing through bistability and slow morphing through diffusion processes. The composites consisted of a hydrogel layer made of gelatine and an architected particle-reinforced epoxy bilayer. Using magnetic fields to achieve spatially distributed orientations of magnetically responsive platelets in each epoxy layer, complex bilayer architectural patterns in various geometries were realised. This feature enabled the study of plant-inspired complex designs,viafinite element analysis and experiments. We present the design and fabrication strategy utilizing the material properties of the composites. The deformations and temporal responses of the resulting composites are analysed using digital image correlation. Finally, we model and experimentally demonstrate plant-inspired composite shells whose stable shapes closely mimic those of the Venus flytrap, while maintaining the multi-stimuli and multi-temporal responses of the materials. The key to achieving this is to tune the local in-plane orientations of the reinforcing particles in the bilayer shapes, to induce distributed in-plane mechanical properties and shrinkage. How these particles should be distributed is determined using finite element modelling. The work presented in this study can be applied to autonomous applications such as robotic systems.

Ether anesthetics prevents touch-induced trigger hair calcium-electrical signals excite the Venus flytrap.

Plants do not have neurons but operate transmembrane ion channels and can get electrical excited by physical and chemical clues. Among them the Venus flytrap is characterized by its peculiar hapto-electric signaling. When insects collide with trigger hairs emerging the trap inner surface, the mechanical stimulus within the mechanosensory organ is translated into a calcium signal and an action potential (AP). Here we asked how the Ca2+ wave and AP is initiated in the trigger hair and how it is feed into systemic trap calcium-electrical networks. When Dionaea muscipula trigger hairs matures and develop hapto-electric excitability the mechanosensitive anion channel DmMSL10/FLYC1 and voltage dependent SKOR type Shaker K+ channel are expressed in the sheering stress sensitive podium. The podium of the trigger hair is interface to the flytrap's prey capture and processing networks. In the excitable state touch stimulation of the trigger hair evokes a rise in the podium Ca2+ first and before the calcium signal together with an action potential travel all over the trap surface. In search for podium ion channels and pumps mediating touch induced Ca2+ transients, we, in mature trigger hairs firing fast Ca2+ signals and APs, found OSCA1.7 and GLR3.6 type Ca2+ channels and ACA2/10 Ca2+ pumps specifically expressed in the podium. Like trigger hair stimulation, glutamate application to the trap directly evoked a propagating Ca2+ and electrical event. Given that anesthetics affect K+ channels and glutamate receptors in the animal system we exposed flytraps to an ether atmosphere. As result propagation of touch and glutamate induced Ca2+ and AP long-distance signaling got suppressed, while the trap completely recovered excitability when ether was replaced by fresh air. In line with ether targeting a calcium channel addressing a Ca2+ activated anion channel the AP amplitude declined before the electrical signal ceased completely. Ether in the mechanosensory organ did neither prevent the touch induction of a calcium signal nor this post stimulus decay. This finding indicates that ether prevents the touch activated, glr3.6 expressing base of the trigger hair to excite the capture organ.

Structural insights into the Venus flytrap mechanosensitive ion channel Flycatcher1.

Flycatcher1 (FLYC1), a MscS homolog, has recently been identified as a candidate mechanosensitive (MS) ion channel involved in Venus flytrap prey recognition. FLYC1 is a larger protein and its sequence diverges from previously studied MscS homologs, suggesting it has unique structural features that contribute to its function. Here, we characterize FLYC1 by cryo-electron microscopy, molecular dynamics simulations, and electrophysiology. Akin to bacterial MscS and plant MSL1 channels, we find that FLYC1 central core includes side portals in the cytoplasmic cage that regulate ion preference and conduction, by identifying critical residues that modulate channel conductance. Topologically unique cytoplasmic flanking regions can adopt 'up' or 'down' conformations, making the channel asymmetric. Disruption of an up conformation-specific interaction severely delays channel deactivation by 40-fold likely due to stabilization of the channel open state. Our results illustrate novel structural features and likely conformational transitions that regulate mechano-gating of FLYC1.

Mechanical factors contributing to the Venus flytrap's rate-dependent response to stimuli.

The sensory hairs of the Venus flytrap (Dionaea muscipula Ellis) detect mechanical stimuli imparted by their prey and fire bursts of electrical signals called action potentials (APs). APs are elicited when the hairs are sufficiently stimulated and two consecutive APs can trigger closure of the trap. Earlier experiments have identified thresholds for the relevant stimulus parameters, namely the angular displacement [Formula: see text] and angular velocity [Formula: see text]. However, these experiments could not trace the deformation of the trigger hair's sensory cells, which are known to transduce the mechanical stimulus. To understand the kinematics at the cellular level, we investigate the role of two relevant mechanical phenomena: viscoelasticity and intercellular fluid transport using a multi-scale numerical model of the sensory hair. We hypothesize that the combined influence of these two phenomena and [Formula: see text] contribute to the flytrap's rate-dependent response to stimuli. In this study, we firstly perform sustained deflection tests on the hair to estimate the viscoelastic material properties of the tissue. Thereafter, through simulations of hair deflection tests at different loading rates, we were able to establish a multi-scale kinematic link between [Formula: see text] and the cell wall stretch [Formula: see text]. Furthermore, we find that the rate at which [Formula: see text] evolves during a stimulus is also proportional to [Formula: see text]. This suggests that mechanosensitive ion channels, expected to be stretch-activated and localized in the plasma membrane of the sensory cells, could be additionally sensitive to the rate at which stretch is applied.

Cold atmospheric pressure He-plasma jet and plasma ball interactions with the Venus flytrap: Electrophysiology and side effects.

Cold atmospheric pressure radio frequency plasma (CAPP) can play an important role in agriculture, medicine, biophysical and bioelectrochemical applications, disinfection and sterilization, synthesis of different compounds, nitrogen fixation, and treatment of surfaces. Here we found that reactive oxygen and nitrogen species, UV-Vis photons, and high-frequency strong electromagnetic fields with an amplitude of a few kV produced by a cold plasma jet can interact with bio-tissue and damage it if the plasma treatment is long enough. The electrophysiological effects of CAPP treatment of bio-tissue and electrical signals transmission were measured in the Venus flytrap. The plasma ball does not produce any visible side effects on the Venus flytrap, but induces electrical signals in bio-tissue with very high amplitude. Plasma (Kirlian) photography shows the existence of a blue aura around the plasma ball due to a corona discharge. Understanding the mechanisms of interactions between CAPP and bio-tissue and preventing side effects can contribute to the application of plasma technology in medicine and agriculture. The use of cold plasma in medicine and agriculture should be monitored for side effects from strong high-frequency electro-magnetic fields, UV photons, and reactive oxygen and nitrogen species to protect against undesirable consequences.

Excitability and propagation of the electrical impulse in Venus flytrap; a comparative electrophysiological study of unipolar electrograms with myocardial tissue.

Mammalian heart cells and cells of leaves of Dionaea muscipula share the ability to generate propagated action potentials, because the excitable cells are electrically coupled. In the heart the propagated action potential causes synchronized contraction of the heart muscle after automatic generation of the impulse in the sinus node. In Dionaea propagation results in closure of the trap after activation of trigger hairs by an insect. The electrical activity can be recorded in the extracellular space as an extracellular electrogram, resulting from transmembrane currents. Although the underlying physiological mechanism that causes the electrogram is similar for heart and Dionaea cells, the contribution of the various ions to the transmembrane current is different. We recorded extracellular electrograms from Dionaea leaves and compared the recorded signals with those known from the heart. The morphology of the electrograms differed considerably. In comparison to activation in mammalian myocardium, electrograms of Dionaea are more temporally and spatially variable. Whereas electrograms in healthy myocardium recorded at some distance from the site of activation reveal a simple biphasic pattern, Dionaea activation showed positive, negative or biphasic deflections. Comparison of patch clamp data from plant cells and cardiomyocytes suggests a role of temperature and ion concentrations in extracellular space for the diversity of morphologies of the Dionaea electrograms.

Action potentials induce biomagnetic fields in carnivorous Venus flytrap plants.

Upon stimulation, plants elicit electrical signals that can travel within a cellular network analogous to the animal nervous system. It is well-known that in the human brain, voltage changes in certain regions result from concerted electrical activity which, in the form of action potentials (APs), travels within nerve-cell arrays. Electro- and magnetophysiological techniques like electroencephalography, magnetoencephalography, and magnetic resonance imaging are used to record this activity and to diagnose disorders. Here we demonstrate that APs in a multicellular plant system produce measurable magnetic fields. Using atomic optically pumped magnetometers, biomagnetism associated with electrical activity in the carnivorous Venus flytrap, Dionaea muscipula, was recorded. Action potentials were induced by heat stimulation and detected both electrically and magnetically. Furthermore, the thermal properties of ion channels underlying the AP were studied. Beyond proof of principle, our findings pave the way to understanding the molecular basis of biomagnetism in living plants. In the future, magnetometry may be used to study long-distance electrical signaling in a variety of plant species, and to develop noninvasive diagnostics of plant stress and disease.

Gaussian-preserved, non-volatile shape morphing in three-dimensional microstructures for dual-functional electronic devices.

Motile plant structures such as Mimosa pudica leaves, Impatiens glandulifera seedpods, and Dionaea muscipula leaves exhibit fast nastic movements in a few seconds or less. This motion is stimuli-independent mechanical movement following theorema egregium rules. Artificial analogs of tropistic motion in plants are exemplified by shape-morphing systems, which are characterized by high functional robustness and resilience for creating 3D structures. However, all shape-morphing systems developed so far rely exclusively on continuous external stimuli and result in slow response. Here, we report a Gaussian-preserved shape-morphing system to realize ultrafast shape morphing and non-volatile reconfiguration. Relying on the Gaussian-preserved rules, the transformation can be triggered by mechanical or thermal stimuli within a microsecond. Moreover, as localized energy minima are encountered during shape morphing, non-volatile configuration is preserved by geometrically enhanced rigidity. Using this system, we demonstrate a suite of electronic devices that are reconfigurable, and therefore, expand functional diversification.

Functional organisation of the endomembrane network in the digestive gland of the Venus flytrap: revisiting an old story with a new microscopy toolbox.

Up-to-date imaging approaches were used to address the spatiotemporal organisation of the endomembrane system in secretory cells of Dionaea muscipula. Different 'slice and view' methodologies were performed on resin-embedded samples to finally achieve a 3D reconstruction of the cell architecture, using ultrastructural tomography, array tomography, serial block face-scanning electron microscopy (SBF-SEM), correlation, and volume rendering at the light microscopy level. Observations of cryo-fixed samples by high-pressure freezing revealed changes of the endomembrane system that occur after trap activation and prey digestion. They provide evidence for an original strategy that adapts the secretory machinery to a specific and unique case of stimulated exocytosis in plant cells. A first secretion peak is part of a rapid response to deliver digestive fluids to the cell surface, which delivers the needed stock of digestive materials 'on site'. The second peak of activity could then be associated with the reconstruction of the Golgi apparatus (GA), endoplasmic reticulum (ER) and vacuolar machinery, in order to prepare for a subsequent round of prey capture. Tubular continuum between ER and Golgi stacks observed on ZIO-impregnated tissues may correspond to an efficient transfer mechanism for lipids and/or proteins, especially for use in rapidly resetting the molecular GA machinery. The occurrence of one vacuolar continuum may permit continuous adjustment of cell homeostasy. The subcellular features of the secretory cells of Dionaea muscipula outline key innovations in the organisation of plant cell compartmentalisation that are used to cope with specific cell needs such as the full use of the GA as a protein factory, and the ability to create protein reservoirs in the periplasmic space. Shape-derived forces of the pleiomorphic vacuole may act as signals to accompany the sorting and entering flows of the cell.

A single touch can provide sufficient mechanical stimulation to trigger Venus flytrap closure.

The carnivorous Venus flytrap catches prey by an ingenious snapping mechanism. Based on work over nearly 200 years, it has become generally accepted that two touches of the trap's sensory hairs within 30 s, each one generating an action potential, are required to trigger closure of the trap. We developed an electromechanical model, which, however, suggests that under certain circumstances one touch is sufficient to generate two action potentials. Using a force-sensing microrobotic system, we precisely quantified the sensory-hair deflection parameters necessary to trigger trap closure and correlated them with the elicited action potentials in vivo. Our results confirm the model's predictions, suggesting that the Venus flytrap may be adapted to a wider range of prey movements than previously assumed.

Snapping mechanics of the Venus flytrap (Dionaea muscipula).

The mechanical principles for fast snapping in the iconic Venus flytrap are not yet fully understood. In this study, we obtained time-resolved strain distributions via three-dimensional digital image correlation (DIC) for the outer and inner trap-lobe surfaces throughout the closing motion. In combination with finite element models, the various possible contributions of the trap tissue layers were investigated with respect to the trap's movement behavior and the amount of strain required for snapping. Supported by in vivo experiments, we show that full trap turgescence is a mechanical-physiological prerequisite for successful (fast and geometrically correct) snapping, driven by differential tissue changes (swelling, shrinking, or no contribution). These are probably the result of the previous accumulation of internal hydrostatic pressure (prestress), which is released after trap triggering. Our research leads to an in-depth mechanical understanding of a complex plant movement incorporating various actuation principles.

A soft pneumatic bistable reinforced actuator bioinspired by Venus Flytrap with enhanced grasping capability.

Soft actuators, as an important part of soft robotics, have attracted significant attention due to their inherent compliance, flexibility and safety. However, low capacity in force and load limits their applications. Prestored elastic energy can improve the capacity in output force and load of soft actuators. This work introduces a soft pneumatic bistable reinforced actuator inspired by the Venus Flytrap's bistable mechanism that allows for the storage of elastic energy. The proposed actuator consists of two separated 3D-printed actuation chambers that are attached to a central prestressed steel shell to achieve bistability. The pressure triggering the pneumatic bistable reinforced actuator from one stable state to the other is derived and validated by experiments. Further experimental comparisons between the proposed actuator and a dual chamber actuator show that the proposed design significantly improves the block tip force, load capacity and stiffness. The pneumatic bistable reinforced actuator also demonstrates superior performance in the actuation speed and bending angle under the same input pressure. Finally, a two-finger gripper is developed using the proposed actuator, which is demonstrated to grasp and hold various objects.

Fine structure of Aldrovanda vesiculosa L: the peculiar lifestyle of an aquatic carnivorous plant elucidated by electron microscopy using cryo-techniques.

The aquatic carnivorous plant Aldrovanda vesiculosa L. is critically endangered worldwide; its peculiar lifestyle raises many questions and poses problems both intriguing on their own and relevant to conservation. While establishing a culture system for its propagation and restoring its natural habitat in Hozoji pond in Saitama, Japan, we conducted ultrastructural observations to examine the various aspects of Aldrovanda's way of life. Electron microscopic observation in combination with cryo-techniques produced novel information which could not be obtained by other methods. Some of the results are: phosphorous is stored in petiole cells of turions during winter; mucilaginous guides are provided for pollen tubes in parietal placental ovaries; storage of potassium in the vicinity of the midrib of carnivorous leaves may contribute to the rapid closing of the carnivorous leaves; dynamic sequential changes of the ultrastructure of digestive glands are involved in the synthesis and secretion of digestive enzymes, including protease and acid phosphatase. These results should contribute significantly to our understanding of Aldrovanda and the detailed mechanisms of its life.

Kinematics Governing Mechanotransduction in the Sensory Hair of the Venus flytrap.

Insects fall prey to the Venus flytrap (Dionaea muscipula) when they touch the sensory hairs located on the flytrap lobes, causing sudden trap closure. The mechanical stimulus imparted by the touch produces an electrical response in the sensory cells of the trigger hair. These cells are found in a constriction near the hair base, where a notch appears around the hair's periphery. There are mechanosensitive ion channels (MSCs) in the sensory cells that open due to a change in membrane tension; however, the kinematics behind this process is unclear. In this study, we investigate how the stimulus acts on the sensory cells by building a multi-scale hair model, using morphometric data obtained from µ-CT scans. We simulated a single-touch stimulus and evaluated the resulting cell wall stretch. Interestingly, the model showed that high stretch values are diverted away from the notch periphery and, instead, localized in the interior regions of the cell wall. We repeated our simulations for different cell shape variants to elucidate how the morphology influences the location of these high-stretch regions. Our results suggest that there is likely a higher mechanotransduction activity in these 'hotspots', which may provide new insights into the arrangement and functioning of MSCs in the flytrap.

Prey capture analyses in the carnivorous aquatic waterwheel plant (Aldrovanda vesiculosa L., Droseraceae).

We investigated the predator-prey interactions between an Australian ecotype of the carnivorous waterwheel plant (Aldrovanda vesiculosa, Droseraceae) and its potential natural prey, the water flea Daphnia longicephala (Daphniidae), which also occurs in Australia. A. vesiculosa develops snap-traps, which close within ~10-100 ms after mechanical triggering by zooplankton prey. Prey capture attempts (PCAs) were recorded via high-speed cinematography in the laboratory. From 14 recorded PCAs, nine were successful for the plant (the prey was caught), and five were unsuccessful (prey could escape), resulting in a capture rate of ~64%. The prey animals' locomotion behaviour (antenna beat frequency and movement type) in trap vicinity or inside the open traps is very variable. Traps were mainly triggered with the second antennae. During trap closure, the animals moved only very little actively. A flight response in reaction to an initiated trap closure was not observed. However, several animals could escape, either by having a "lucky" starting position already outside the triggered trap, by freeing themselves after trap closure, or by being pressed out by the closing trap lobes. According to our observations in the successful PCAs, we hypothesize that the convex curvature of the two trap lobes (as seen from the outside) and the infolded trap rims are structural means supporting the capture and retention of prey. Our results are discussed in a broader biological context and promising aspects for future studies are proposed.

Venus flytrap trigger hairs are micronewton mechano-sensors that can detect small insect prey.

Venus flytraps detect moving insects via highly sensitive, action potential (AP)-producing trigger hairs, which act as high-sensitivity levers, crucial for prey capture and digestion. Controlled stimulation revealed that they can trigger APs for deflections >2.9°, angular velocities >3.4° s-1 and forces >29 µN. Hairs became desensitized and subsequently responded to fast consecutive stimulations; desensitization increased at lower temperatures. Recording of ant trigger hair contact events revealed that even small insects exceed the hairs' sensitivity threshold.