Numerical Simulation and Experimental Studies of a Ribbon Coil for Trans Spinal Magnetic Stimulation (TSMS) in Rats.

We propose a non-invasive Trans Spinal Magnetic Stimulation (TSMS) coil allowing for focal stimulation. The device is based on a new figure-8 ribbon design, ensuring low R0, and low heating. The two coils were designed and studied using the finite element method (FEM) coupled with NEURON and tested for efficacy on rats. The numerical simulations confirmed the generation of the observed action potentials when the coil was driven with 2.8kA.Clinical Relevance- Chronic neuropathic back and leg pain is one of the main indications for spinal cord stimulation in the United States. Chronic low back pain is one of the most common reasons patients seek medical care, and in 2013 resulted in 87.6 billion dollars in healthcare costs in the USA. Patients would most likely prefer a low-risk, non-invasive procedure, such as TSMS, to surgery with a significant rate of complications.

Electric field bridging-effect in electrified microfibrils' scaffolds.

Introduction: The use of biocompatible scaffolds combined with the implantation of neural stem cells, is increasingly being investigated to promote the regeneration of damaged neural tissue, for instance, after a Spinal Cord Injury (SCI). In particular, aligned Polylactic Acid (PLA) microfibrils' scaffolds are capable of supporting cells, promoting their survival and guiding their differentiation in neural lineage to repair the lesion. Despite its biocompatible nature, PLA is an electrically insulating material and thus it could be detrimental for increasingly common scaffolds' electric functionalization, aimed at accelerating the cellular processes. In this context, the European RISEUP project aims to combine high intense microseconds pulses and DC stimulation with neurogenesis, supported by a PLA microfibrils' scaffold. Methods: In this paper a numerical study on the effect of microfibrils' scaffolds on the E-field distribution, in planar interdigitated electrodes, is presented. Realistic microfibrils' 3D CAD models have been built to carry out a numerical dosimetry study, through Comsol Multiphysics software. Results: Under a voltage of 10 V, microfibrils redistribute the E-field values focalizing the field streamlines in the spaces between the fibers, allowing the field to pass and reach maximum values up to 100 kV/m and values comparable with the bare electrodes' device (without fibers). Discussion: Globally the median E-field inside the scaffolded electrodes is the 90% of the nominal field, allowing an adequate cells' exposure.

Astrocyte Responses Influence Local Effects of Whole-Brain Magnetic Stimulation in Parkinsonian Rats.

BACKGROUND: Excessive glutamatergic transmission in the striatum is implicated in Parkinson's disease (PD) progression. Astrocytes maintain glutamate homeostasis, protecting from excitotoxicity through the glutamate-aspartate transporter (GLAST), whose alterations have been reported in PD. Noninvasive brain stimulation using intermittent theta-burst stimulation (iTBS) acts on striatal neurons and glia, inducing neuromodulatory effects and functional recovery in experimental parkinsonism. OBJECTIVE: Because PD is associated with altered astrocyte function, we hypothesized that acute iTBS, known to rescue striatal glutamatergic transmission, exerts regional- and cell-specific effects through modulation of glial functions. METHODS: 6-Hydroxydopamine-lesioned rats were exposed to acute iTBS, and the areas predicted to be more responsive by a biophysical, hyper-realistic computational model that faithfully reconstructs the experimental setting were analyzed. The effects of iTBS on glial cells and motor behavior were evaluated by molecular and morphological analyses, and CatWalk and Stepping test, respectively. RESULTS: As predicted by the model, the hippocampus, cerebellum, and striatum displayed a marked c-FOS activation after iTBS, with the striatum showing specific morphological and molecular changes in the astrocytes, decreased phospho-CREB levels, and recovery of GLAST. Striatal-dependent motor performances were also significantly improved. CONCLUSION: These data uncover an unknown iTBS effect on astrocytes, advancing the understanding of the complex mechanisms involved in TMS-mediated functional recovery. Data on numerical dosimetry, obtained with a degree of anatomical details never before considered and validated by the biological findings, provide a framework to predict the electric-field induced in different specific brain areas and associate it with functional and molecular changes. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

A study of flex miniaturized coils for focal nerve magnetic stimulation.

BACKGROUND: Peripheral magnetic stimulation (PMS) is emerging as a complement to standard electrical stimulation (ES) of the peripheral nervous system (PNS). PMS may stimulate sensory and motor nerve fibers without the discomfort associated with the ES used for standard nerve conduction studies. The PMS coils are the same ones used in transcranial magnetic stimulation (TMS) and lack focality and selectiveness in the stimulation. PURPOSE: This study presents a novel coil for PMS, developed using Flexible technologies, and characterized by reduced dimensions for a precise and controlled targeting of peripheral nerves. METHODS: We performed hybrid electromagnetic (EM) and electrophysiological simulations to study the EM exposure induced by a novel miniaturized coil (or mcoil) in and around the radial nerve of the neuro-functionalized virtual human body model Yoon-Sun, and to estimate the current threshold to induce magnetic stimulation (MS) of the radial nerve. Eleven healthy subjects were studied with the mcoil, which consisted of two 15 mm diameter coils in a figure-of-eight configuration, each with a hundred turns of a 25 µm copper-clad four-layer foil. Sensory nerve action potentials (SNAPs) were measured in each subject using two electrodes and compared with those obtained from standard ES. The SNAPs conduction velocities were estimated as a performance metric. RESULTS: The induced electric field was estimated numerically to peak at a maximum intensity of 39 V/m underneath the mcoil fed by 70 A currents. In such conditions, the electrophysiological simulations suggested that the mcoil elicits SNAPs originating at 7 mm from the center of the mcoil. Furthermore, the numerically estimated latencies and waveforms agreed with those obtained during the PMS experiments on healthy subjects, confirming the ability of the mcoil to stimulate the radial nerve sensory fibers. CONCLUSION: Hybrid EM-electrophysiological simulations assisted the development of a miniaturized coil with a small diameter and a high number of turns using flexible electronics. The numerical dosimetric analysis predicted the threshold current amplitudes required for a suprathreshold peripheral nerve sensory stimulation, which was experimentally confirmed. The developed and now validated computational pipeline will be used to improve the performances (e.g., focality and minimal currents) of new generations of mcoil designs.

Changes in hydration of liposome membranes exposed to nanosecond electric pulses detected by wide-field Coherent anti-Stokes Raman microspectroscopy.

Electropulsation has become a powerful technological platform for electromanipulation of cells and tissues for various medical and biotechnological applications, but the molecular changes that underlay the very first initiation step of this process have not been experimentally observed. Here, we endowed a wide-field Coherent anti-Stokes Raman Scattering platform with an ad-hoc electromagnetic exposure device and we demonstrated, using artificial lipid vesicles (i.e. liposomes), that electropulsation is initiated by the increase of interstitial water content in liposome membranes. A pulse-dependent accumulation of the interstitial water molecules is observed in the membranes and a plausible mechanism supported by a computational electrochemical model is presented and discussed.

Systematic numerical assessment of occupational exposure to electromagnetic fields of transcranial magnetic stimulation.

PURPOSE: This study aims to perform a classification and rigorous numerical evaluation of the risks of occupational exposure in the health environment related to the administration of transcranial magnetic stimulation (TMS) treatment. The study investigates the numerically estimated induced electric field that occurs in the human tissues of an operator caused by exposure to the variable magnetic field produced by TMS during treatments. This could be a useful starting point for future risk assessment studies and safety indications in this context. METHODS: We performed a review of the actual positions assumed by clinicians during TMS treatments. Three different TMS coils (two circular and one figure-of-eight) were modeled and characterized numerically. Different orientations and positions of each coil with respect to the body of the operator were investigated to evaluate the induced electric (-E) field in the body tissues. The collected data were processed to allow comparison with the safety standards for occupational exposure, as suggested by the International Commission on Non-Ionizing Radiation Protection (ICNIRP) 2010 guidelines. RESULTS: Under the investigated conditions, exposure to TMS shows some criticalities for the operator performing the treatment. Depending on the model of the TMS coil and its relative position with respect to the operator's body, the numerically estimated E-field could exceed the limits suggested by the ICNIRP 2010 guidelines. We established that the worst-case scenario for the three coils occurs when they are placed in correspondence of the abdomen, with the handle oriented parallel to the body (II orientation). Working at a maximum TMS stimulator output (MSO), the induced E-field is up to 7.32 V/m (circular coil) and up to 1.34 V/m (figure-of-eight coil). The induced E-field can be modulated by the TMS percentage of MSO (%MSO) and by the distance between the source and the operator. At %MSO equal to or below 80%, the figure-of-eight coil was compliant with the ICNIRP limit (1.13 V/m). Conversely, the circular coil causes an induced E-field above the limits, even when powered at a %MSO of 30%. Thus, in the investigated worst-case conditions, an operator working with a circular coil should keep a distance from its edge to be compliant with the guidelines limit, which depends on the selected %MSO: 38 cm at 100%, 32 cm at 80%, 26.8 cm at 50%, and 19.8 cm at 30%. Furthermore, attention should be paid to the induced E-field reached in the operator's hand as the operator typically holds the coil by hand. In fact in the hand, we estimated an induced E-field up to 10 times higher than the limits. CONCLUSIONS: Our numerical results indicate that coil positions, orientations, and distances with respect to the operator's body can determine the levels of induced E-field that exceed the ICNIRP limits. The induced E-field is also modulated by the choice of %MSO, which is related to the TMS application. Even under the best exposure conditions, attention should be paid to the exposure of the hand. These findings highlight the need for future risk assessment studies to provide more safety information for the correct and safe use of TMS devices.

Pulsed Electromagnetic Fields: A Novel Attractive Therapeutic Opportunity for Neuroprotection After Acute Cerebral Ischemia.

OBJECTIVES: Acute cerebral ischemia is characterized by several pathological processes evolving during time, which contribute to the final tissue damage. Secondary processes, such as prolonged inflammatory response, impaired mitochondrial function, and oxidative stress, are responsible for the progression of brain injury to the peri-infarct area, called "penumbra." Adenosine has been shown to play a crucial role in regulating the inflammatory cascade following brain ischemia. Pulsed electromagnetic fields (PEMFs) act as modulators of adenosine receptors, increasing the functionality of the endogenous adenosine. In particular, PEMF exposure induces a significant upregulation of A2A and A3 adenosine receptors in different neuronal cell types. Several lines of evidence suggest that PEMF exposure might play a neuroprotective role after ischemic damage. MATERIALS AND METHODS: This review summarizes the current knowledge on the mechanism of action of PEMFs and their biological effects on neuronal damage both in preclinical and clinical studies. RESULTS: PEMFs counteract hypoxia-induced apoptosis and ROS production in neuronal-like cells and exert a strong anti-inflammatory effect on microglial cells. Data from stroke animal models showed that PEMFs exposure is able to reduce the size of the infarct area and decrease the levels of pro-inflammatory mediators. In clinical studies, PEMFs stimulation proved to be safe and well tolerated. Preliminary results on acute ischemic stroke patients showed a dose-dependent reduction in the lesion size. CONCLUSIONS: Altogether, these data demonstrate the efficacy of PEMFs against several mechanisms underlying ischemic damage and suggest that PEMFs might represent a novel noninvasive adjunctive treatment for acute ischemic stroke, providing neuroprotection and reducing functional deficits following ischemia.

Electric-driven membrane poration: A rationale for water role in the kinetics of pore formation.

Electroporation is a well-established technique used to stimulate cells, enhancing membrane permeability by inducing reversible membrane pores. In the absence of experimental observation of the dynamics of pore creation, molecular dynamics studies provide the molecular-level evidence that the electric field promotes pore formation. Although single steps in the pore formation process are well assessed, a kinetic model representing the mathematical description of the electroporation process, is lacking. In the present work we studied the basis of the pore formation process, providing a rationale for the definition of a first-order kinetic scheme. Here, authors propose a three-state kinetic model for the process based on the assessed mechanism of water defects intruding at the water/lipid interface, when applying electric field intensities at the edge of the linear regime. The methodology proposed is based on the use of two robust biophysical quantities analyzed for the water molecules intruding at the water/lipid interface: (i) number of hydrogen bonds; (ii) number of contacts. The final model, sustained by a robust statistical sampling, provides kinetic constants for the transitions from the intact bilayer state to the hydrophobic pore state.

The Impact of Bilayer Rigidity on the Release from Magnetoliposomes Vesicles Controlled by PEMFs.

Stimuli-sensitive nanocarriers have recently been developed as a powerful tool in biomedical applications such as drug delivery, detection, and gene transfer techniques. Among the external triggers investigated, low intensity magnetic fields represent a non-invasive way to remotely control the release of compounds from a magneto-sensitive carrier. Magnetoliposomes (MLs), i.e., liposomes entrapping magnetic nanoparticles (MNPs), are studied due to their capacity to transport hydrophobic and hydrophilic agents, their easy production, and due to the ability of MNPs to respond to a magnetic actuation determining the triggered release of the encapsulated compounds. Here we investigated the design and optimization of the MLs to obtain an efficient on-demand release of the transported compounds, due to the magneto-mechanical actuation induced by applying low-intensity pulsed electromagnetic fields (PEMFs). In particular we studied the effect of the bilayer packing on the ability of MLs, with oleic acid-coated MNPs encapsulated in the bilayer, to respond to PEMFs application. Three kinds of MLs are produced with an increasing rigidity of the bilayer, defined as Liquid Disorder, Liquid Order, and Gel MLs and the delivery of a hydrophilic dye (as a model drug) is investigated. Results demonstrate the efficacy of the magnetic trigger on high-ordered bilayers, which are unable to dampen the perturbation produced by MNPs motion.

Effect of skin conductivity on the electric field induced by transcranial stimulation techniques in different head models.

This study aims at quantifying the effect that using different skin conductivity values has on the estimation of the electric (E)-field distribution induced by transcranial magnetic stimulation (TMS) and transcranial direct current stimulation (tDCS) in the brain of two anatomical models. The induced E-field was calculated with numerical simulations inside MIDA and Duke models, assigning to the skin a conductivity value estimated from a multi-layered skin model and three values taken from literature. The effect of skin conductivity variations on the local E-field induced by tDCS in the brain was up to 70%. In TMS, minor local differences, in the order of 20%, were obtained in regions of interest for the onset of possible side effects. Results suggested that an accurate model of the skin is necessary in all numerical studies that aim at precisely estimating the E-field induced during TMS and tDCS applications. This also highlights the importance of further experimental studies on human skin characterization, especially at low frequencies.

Possible molecular and cellular mechanisms at the basis of atmospheric electromagnetic field bioeffects.

Mechanisms of how electromagnetic (EM) field acts on biological systems are governed by the same physics regardless of the origin of the EM field (technological, atmospheric...), given that EM parameters are the same. We draw from a large body of literature of bioeffects of a man-made electromagnetic field. In this paper, we performed a focused review on selected possible mechanisms of how atmospheric electromagnetic phenomena can act at the molecular and cellular level. We first briefly review the range of frequencies and field strengths for both electric and magnetic fields in the atmosphere. Then, we focused on a concise description of the current knowledge on weak electric and magnetic field bioeffects with possible molecular mechanisms at the basis of possible EM field bioeffects combined with modeling strategies to estimate reliable outcomes and speculate about the biological effects linked to lightning or pyroelectricity. Indeed, we bring pyroelectricity as a natural source of voltage gradients previously unexplored. While very different from lightning, it can result in similar bioeffects based on similar mechanisms, which can lead to close speculations on the importance of these atmospheric electric fields in the evolution.

Numerical Evaluation of Human Body Near Field Exposure to a Vehicular Antenna for Military Applications.

BACKGROUND: The use of electromagnetic (EM) technologies for military applications is gaining increasing interest to satisfy different operational needs, such as improving battlefield communications or jamming counterpart's signals. This is achieved by the use of high-power EM waves in several frequency bands (e.g., HF, VHF, and UHF). When considering military vehicles, several antennas are present in close proximity to the crew personnel, which are thus potentially exposed to high EM fields. METHODS: A typical exposure scenario was reproduced numerically to evaluate the EM exposure of the human body in the presence of an HF vehicular antenna (2-30 MHz). The antenna was modeled as a monopole connected to a 3D polygonal structure representing the vehicle. Both the EM field levels in the absence and in the presence of the human body and also the specific absorption rate (SAR) values were calculated. The presence of the operator, partially standing outside the vehicle, was simulated with the virtual human body model Duke (Virtual Population, V.3). Several exposure scenarios were considered. The presence of a protective helmet was modeled as well. RESULTS: In the area usually occupied by the personnel, E-field intensity radiated by the antenna can reach values above the limits settled by international safety guidelines. Nevertheless, local SAR values induced inside the human body reached a maximum value of 14 mW/kg, leading to whole-body averaged and 10-g averaged SAR values well below the corresponding limits. CONCLUSION: A complex and realistic near-field exposure scenario of the crew of a military vehicle was simulated. The obtained E-field values radiated in the free space by a HF vehicular antenna may reach values above the safety guidelines reference levels. Such values are not necessarily meaningful for the exposed subject. Indeed, SAR and E-field values induced inside the body remain well below safety limits.

Galvanotactic Phenomenon Induced by Non-Contact Electrostatic Field: Investigation in a Scratch Assay.

Non-contact galvanotaxis as a way to drive the cells migration could be a promising tool for a variety of biomedical applications, such as wound healing control, avoiding the interaction between electrodes and cell cultures. To this regard, the efficacy of this electrical stimulus application has to be deeper studied to control physiological migratory phenomena in a remote way.Aim of this work is to provide an experimental investigation on the mobility of cells exposed to a static electric field in a "noncontact" mode, supported by a suitable modeling of the electric field distribution inside the experimental setup. In particular, scratch assays have been carried out placing the electrodes outside the cells medium support and changing the cells holder to study more than one configuration.Clinical Relevance- In this study the in vitro experiments on the non-contact galvanotaxis, together with the numerical simulations of the exposure setup, provide a way to investigate the effects that could affect an electrically drive cell migration.

Confocal Microscopy Improves 3D Microdosimetry Applied to Nanoporation Experiments Targeting Endoplasmic Reticulum.

In the last years, microdosimetric numerical models of cells including intracellular compartments have been proposed, aiming to investigate the poration induced by the application of nanosecond pulsed electric fields (nsPEFs). A limitation of such models was the extremely approximate cell and organelle shapes, leading to an incorrect estimation of the electric field or transmembrane potential distribution in the studied domain. In order to obtain a reliable model of in vitro experiments and a one-to-one comparison between experimental and simulated results, here, a realistic model of 12 human mesenchymal stem cells was built starting from their optical microscopy images where different cell compartments were highlighted. The microdosimetric analysis of the cells group was quantified in terms of electric field and transmembrane potentials (TMPs) induced by an externally applied 10-ns trapezoidal pulse with rise and fall times of 2 ns, with amplitudes ranging from 2 to 30 MV/m. The obtained results showed that the plasma and endoplasmic reticulum (ER) membrane of each cell respond in a different way to the same electric field amplitude, depending on differences in shape, size, and position of the single cell with respect to the applied electric field direction. Therefore, also the threshold for an efficient electroporation is highly different from cell to cell. This difference was quantitatively estimated through the cumulative distribution function of the pore density for the plasma and ER membrane of each cell, representing the probability that a certain percentage of membrane has reached a specific value of pore density. By comparing the dose-response curves resulted from the simulations and those from the experimental study of De Menorval et al. (2016), we found a very good matching of results for plasma and ER membrane when 2% of the porated area is considered sufficient for permeabilizing the membrane. This result is worth of noting as it highlights the possibility to effectively predict the behavior of a cell (or of a population of cells) exposed to nsPEFs. Therefore, the microdosimetric realistic model described here could represent a valid tool in setting up more efficient and controlled electroporation protocols.

Proof-of-Concept of Electrical Activation of Liposome Nanocarriers: From Dry to Wet Experiments.

The increasing interest toward biocompatible nanotechnologies in medicine, combined with electric fields stimulation, is leading to the development of electro-sensitive smart systems for drug delivery applications. To this regard, recently the use of pulsed electric fields to trigger release across phospholipid membranes of liposomes has been numerically studied, for a deeper understanding of the phenomena at the molecular scale. Aim of this work is to give an experimental validation of the feasibility to control the release from liposome vesicles, using nanosecond pulsed electric fields characterized by a 10 ns duration and intensity in the order of MV/m. The results are supported by multiphysics simulations which consider the coupling of three physics (electromagnetics, thermal and pore kinetics) in order to explain the occurring physical interactions at the microscopic level and provide useful information on the characteristics of the train of pulses needed to obtain quantitative results in terms of liposome electropermeabilization. Finally, a complete characterization of the exposure system is also provided to support the reliability and validity of the study.

Patient Semi-specific Computational Modeling of Electromagnetic Stimulation Applied to Neuroprotective Treatments in Acute Ischemic Stroke.

Neuroprotective effects of pulsed electromagnetic fields (PEMFs) have been demonstrated both in vivo and in vitro. Moreover, preliminary clinical studies have been conducted and suggested PEMFs as a possible alternative therapy to treat acute ischemic stroke. In this work, we show that it's possible to build-up a patient semi-specific head model, where the 3D reconstruction of the ischemic lesion of the patient under treatment is inserted in the head of the human body model "Duke" (v.1.0, Zurich MedTech AG). The semi-specific model will be used in the randomized, placebo-controlled, double-blind study currently ongoing. Three patients were modelled and simulated, and results showed that each ischemic lesion experiences a magnetic flux density field comparable to the one for which biological effects have been attested. Such a kind of dosimetric analysis reveals a reliable tool to assess the correlation between levels of exposure and the beneficial effect. Thus, once the on-going double blind study is complete it will prove if PEMFs treatment triggers a clinical effect, and we will then be able to characterize a dose-response curve with the methodology arranged in this study.

Nanosecond pulsed electric signals can affect electrostatic environment of proteins below the threshold of conformational effects: The case study of SOD1 with a molecular simulation study.

Electric fields can be a powerful tool to interact with enzymes or proteins, with an intriguing perspective to allow protein manipulation. Recently, researchers have focused the interest on intracellular enzyme modifications triggered by the application of nanosecond pulsed electric fields. These findings were also supported by theoretical predictions from molecular dynamics simulations focussing on significant variations in protein secondary structures. In this work, a theoretical study utilizing molecular dynamics simulations is proposed to explore effects of electric fields of high intensity and very short nanosecond duration applied to the superoxide dismutase (Cu/Zn-SOD or SOD-1), an important enzyme involved in the cellular antioxidant defence mechanism. The effects of 100-nanosecond pulsed electric fields, with intensities ranging from 108 to 7x108 V/m, on a single SOD1 enzyme are presented. We demonstrated that the lowest intensity of 108 V/m, although not inducing structural changes, can produce electrostatic modifications on the reaction centre of the enzyme, as apparent from the dipolar response and the electric field distribution of the protein active site. Electric pulses above 5x108 V/m produced a fast transition between the folded and a partially denatured state, as inferred by the secondary structures analysis. Finally, for the highest field intensity used (7x108 V/m), a not reversible transition toward an unfolded state was observed.

Tubulin response to intense nanosecond-scale electric field in molecular dynamics simulation.

Intense pulsed electric fields are known to act at the cell membrane level and are already being exploited in biomedical and biotechnological applications. However, it is not clear if electric pulses within biomedically-attainable parameters could directly influence intra-cellular components such as cytoskeletal proteins. If so, a molecular mechanism of action could be uncovered for therapeutic applications of such electric fields. To help clarify this question, we first identified that a tubulin heterodimer is a natural biological target for intense electric fields due to its exceptional electric properties and crucial roles played in cell division. Using molecular dynamics simulations, we then demonstrated that an intense - yet experimentally attainable - electric field of nanosecond duration can affect the bß-tubulin's C-terminus conformations and also influence local electrostatic properties at the GTPase as well as the binding sites of major tubulin drugs site. Our results suggest that intense nanosecond electric pulses could be used for physical modulation of microtubule dynamics. Since a nanosecond pulsed electric field can penetrate the tissues and cellular membranes due to its broadband spectrum, our results are also potentially significant for the development of new therapeutic protocols.

Evidences of plasma membrane-mediated ROS generation upon ELF exposure in neuroblastoma cells supported by a computational multiscale approach.

BACKGROUND: Molecular mechanisms of interaction between cells and extremely low frequency magnetic fields (ELF-MFs) still represent a matter of scientific debate. In this paper, to identify the possible primary source of oxidative stress induced by ELF-MF in SH-SY5Y human neuroblastoma cells, we estimated the induced electric field and current density at the cell level. METHODS: We followed a computational multiscale approach, estimating the local electric field and current density from the whole sample down to the single cell level. The procedure takes into account morphological modeling of SH-SY5Y cells, arranged in different topologies. Experimental validation has been carried out: neuroblastoma cells have been treated with Diphenyleneiodonium (DPI) -an inhibitor of the plasma membrane enzyme NADPH oxidase (Nox)- administered 24 h before exposure to 50 Hz (1 mT) MF. RESULTS: Macroscopic and microscopic dosimetric evaluations suggest that increased current densities are induced at the plasma membrane/extra-cellular medium interface; identifying the plasma membrane as the main site of the ELF-neuroblastoma cell interaction. The in vitro results provide an experimental proof that plasma membrane Nox exerts a key role in the redox imbalance elicited by ELF, as DPI treatment reverts the generation of reactive oxygen species induced by ELF exposure. GENERAL SIGNIFICANCE: Microscopic current densities induced at the plasma membrane are likely to play an active physical role in eliciting ELF effects related to redox imbalance. Multiscale computational dosimetry, supported by an in vitro approach for validation, is proposed as the innovative and rigorous paradigm to unveil mechanisms underlying the complex ELF-MF interactions.

Cancellation of nerve excitation by the reversal of nanosecond stimulus polarity and its relevance to the gating time of sodium channels.

The initiation of action potentials (APs) by membrane depolarization occurs after a brief vulnerability period, during which excitation can be abolished by the reversal of the stimulus polarity. This vulnerability period is determined by the time needed for gating of voltage-gated sodium channels (VGSC). We compared nerve excitation by ultra-short uni- and bipolar stimuli to define the time frame of bipolar cancellation and of AP initiation. Propagating APs in isolated frog sciatic nerve were elicited by cathodic pulses (200 ns-300 µs), followed by an anodic (canceling) pulse of the same duration after a 0-200-µs delay. We found that the earliest and the latest boundaries for opening the critical number of VGSC needed to initiate AP are, respectively, between 11 and 20 µs and between 100 and 200 µs after the onset of depolarization. Stronger depolarization accelerated AP initiation, apparently due to faster VGSC opening, but not beyond the 11-µs limit. Bipolar cancellation was augmented by reducing pulse duration, shortening the delay between pulses, decreasing the amplitude of the cathodic pulse, and increasing the amplitude of the anodic one. Some of these characteristics contrasted the bipolar cancellation of cell membrane electroporation (Pakhomov et al. in Bioelectrochemistry 122:123-133, 2018; Gianulis et al. in Bioelectrochemistry 119:10-19, 2017), suggesting different mechanisms. The ratio of nerve excitation thresholds for a unipolar cathodic pulse and a symmetrical bipolar pulse increased as a power function as the pulse duration decreased, in remarkable agreement with the predictions of SENN model of nerve excitation (Reilly and Diamant in Health Phys 83(3):356-365, 2002).