Safety of ipsilesional anodal transcranial direct current stimulation in acute photothrombotic stroke: implications for early neurorehabilitation.

Early rehabilitation in the acute phase of stroke, that bears unique neuroplastic properties, is the current standard to reduce disability. Anodal transcranial direct current stimulation can augment neurorehabilitation in chronic stroke. Studies in the acute phase are sparse and held back by inconclusive preclinical data pointing towards potential negative interaction of the excitability increasing tDCS modality with stroke-induced glutamate toxicity. In this present study, we aimed to evaluate structural and behavioral safety of anodal tDCS applied in the acute phase of stroke. Photothrombotic stroke including the right primary motor cortex was induced in rats. 24 h after stroke anodal tDCS was applied for 20 min ipsilesionally at one of four different current densities in freely moving animals. Effects on the infarct volume and on stroke induced neuroinflammation were assessed. Behavioral consequences were monitored. Infarct volume and the modified Neurological Severity Score were not affected by anodal tDCS. Pasta handling, a more sensitive task for sensorimotor deficits, and microglia reactivity indicated potentially harmful effects at the highest tDCS current density tested (47.8 A/m2), which is more than 60 times higher than intensities commonly used in humans. Compared to published safety limits of anodal tDCS in healthy rats, recent stroke does not increase the sensitivity of the brain to anodal tDCS, as assessed by lesion size and neuroinflammatory response. Behavioral deficits only occurred at the highest intensity, which was associated with increased neuroinflammation. When safety limits of commonly used clinical tDCS are met, augmentation of early neurorehabilitation after stroke by anodal tDCS appears to be feasible.

Cx3cr1 deficiency interferes with learning- and direct current stimulation-mediated neuroplasticity of the motor cortex.

Microglia are essential contributors to synaptic transmission and stability and communicate with neurons via the fractalkine pathway. Transcranial direct current stimulation [(t)DCS], a form of non-invasive electrical brain stimulation, modulates cortical excitability and promotes neuroplasticity, which has been extensively demonstrated in the motor cortex and for motor learning. The role of microglia and their fractalkine receptor CX3CR1 in motor cortical neuroplasticity mediated by DCS or motor learning requires further elucidation. We demonstrate the effects of pharmacological microglial depletion and genetic Cx3cr1 deficiency on the induction of DCS-induced long-term potentiation (DCS-LTP) ex vivo. The relevance of microglia-neuron communication for DCS response and structural neuroplasticity underlying motor learning are assessed via 2-photon in vivo imaging. The behavioural consequences of impaired CX3CR1 signalling are investigated for both gross and fine motor learning. We show that DCS-mediated neuroplasticity in the motor cortex depends on the presence of microglia and is driven in part by CX3CR1 signalling ex vivo and provide the first evidence of microglia interacting with neurons during DCS in vivo. Furthermore, CX3CR1 signalling is required for motor learning and underlying structural neuroplasticity in concert with microglia interaction. Although we have recently demonstrated the microglial response to DCS in vivo, we now provide a link between microglial integrity and neuronal activity for the expression of DCS-dependent neuroplasticity. In addition, we extend the knowledge on the relevance of CX3CR1 signalling for motor learning and structural neuroplasticity. The underlying molecular mechanisms and the potential impact of DCS in rescuing CX3CR1 deficits remain to be addressed in the future.

Direct current stimulation increases blood flow and permeability of cortical microvasculature in vivo.

BACKGROUND AND PURPOSE: Transcranial direct current stimulation (DCS) structurally and functionally modulates neuronal networks and microglia dynamics. Neurovascular coupling adapts regional cerebral blood flow to neuronal activity and metabolic demands. METHODS: In this study, we examined effects of anodal DCS on vessel morphology, blood flow parameters, permeability of cortical microvasculature, and perivascular microglia motility by time-lapse two-photon microscopy in anaesthetized mice. RESULTS: Low-intensity DCS significantly increased vessel diameter and blood flow parameters. These effects were transient and dependent on the spontaneous vasomotion characteristics of the individual vessel. Vessel leakage increased significantly after DCS at 1.1 and was more pronounced at 2.2 A/m2 , indicating a dose-dependent increase in vascular permeability. Perivascular microglia exhibited increased soma motility post-DCS at both intensities, potentially triggered by the extravasation of intravascular substrates. CONCLUSIONS: Our findings demonstrate that DCS affected only vessels with spontaneous vasomotion. This rapid vascular response may occur as an adaptation of regional blood supply to neuronal excitability altered by DCS or as a direct effect on the vessel wall. In contrast to these immediate effects during stimulation, increases in cortical vessel permeability and perivascular microglia motility appeared after the stimulation had ended.

Electrified microglia: Impact of direct current stimulation on diverse properties of the most versatile brain cell.

BACKGROUND: Transcranial direct current stimulation [(t)DCS], modulates cortical excitability and promotes neuroplasticity. Microglia has been identified to respond to electrical currents as well as neuronal activity, but its response to DCS is mostly unknown. OBJECTIVE: This study addresses effects of DCS applied in vivo to the sensorimotor cortex on physiological microglia properties and neuron-microglia communication. METHODS: Time lapse in vivo 2-photon microscopy in anaesthetized mice was timely coupled with DCS of the sensorimotor cortex to observe microglia dynamics on a population-based and single cell level. Neuron-microglia communication during DCS was investigated in mice with a functional knock out of the fractalkine receptor CX3CR1. Moreover, the role of voltage gated microglial channels and DCS effects on phagocytosis were studied. RESULTS: DCS promoted several physiological microglia properties, depending on the glial activation state and stimulation intensity. On a single cell level, process motility was predominantly enhanced in ramified cells whereas horizontal soma movement and galvanotaxis was pronounced in reactive microglia. Blockage of voltage sensitive microglial channels suppressed DCS effects in vivo and in vitro. Microglial motility changes were partially driven by the fractalkine signaling pathway. Moreover, phagocytosis increased after DCS in vitro. CONCLUSION: Microglia dynamics are rapidly influenced by DCS. This is the first in vivo demonstration of a direct effect of electrical currents on microglia and indirect effects potentially driven by neuronal activity via the fractalkine pathway.

Direct current stimulation-induced synaptic plasticity in the sensorimotor cortex: structure follows function.

BACKGROUND: Non-invasive direct current stimulation (DCS) of the brain induces functional plasticity in vitro and facilitates motor learning across species. The effect of DCS on structural synaptic plasticity is currently unknown. OBJECTIVE: This study addresses the effects and the underlying mechanisms of anodal DCS on structural plasticity and morphology of dendritic spines in the sensorimotor cortex (M1/S1). METHODS: A DCS electrode setup was combined with a chronic cranial window over M1/S1 in transgenic Thy1-GFP mice, to allow for in vivo 2-photon microscopy and simultaneous DCS. Contralateral electrical forepaw stimulation (eFS) was used to mimic the second synapse specific input, a previously shown requirement to induce functional plasticity by DCS. Changes in spine density and spine morphology were compared between DCS/eFS and sham, as well as two control conditions (sham-DCS/eFS, DCS/sham-eFS). Furthermore, the role of BDNF for stimulation-induced changes in spine density was assessed in heterozygous Thy1-GFP x BDNF+/- mice. RESULTS: Combined DCS/eFS rapidly increased spine density during stimulation and changes outlasted the intervention for 24 h. This effect was due to increased survival of original spines and a preferential formation of new spines after intervention. The latter were morphologically characterized by larger head sizes. The DCS-induced spine density increase was absent in mice with reduced BDNF expression. CONCLUSION: Previous findings of DCS-induced functional synaptic plasticity can be extended to structural plasticity in M1/S1 that similarly depends on a second synaptic input (eFS) and requires physiological BDNF expression. These findings show considerable parallels to motor learning-induced M1 spine dynamics.

Prevention of schizophrenia deficits via non-invasive adolescent frontal cortex stimulation in rats.

Schizophrenia is a severe neurodevelopmental psychiatric affliction manifested behaviorally at late adolescence/early adulthood. Current treatments comprise antipsychotics which act solely symptomatic, are limited in their effectiveness and often associated with side-effects. We here report that application of non-invasive transcranial direct current stimulation (tDCS) during adolescence, prior to schizophrenia-relevant behavioral manifestation, prevents the development of positive symptoms and related neurobiological alterations in the maternal immune stimulation (MIS) model of schizophrenia.

Cerebral granulomatosis as a manifestation of Crohn's disease.

BACKGROUND: Crohn's disease (CD) is associated with a variety of extra-intestinal manifestations. Most commonly these involve the eye, skin, joints, coagulation system and liver. Cerebral manifestations of CD have been reported to a far lesser extent. The extensive detrimental impact of neurological symptoms on a patient's quality of life makes an early diagnosis and treatment particularly important. In previous case-reports, diagnosis of cerebral manifestations in CD often relied upon magnetic resonance imaging (MRI) and computed tomography (CT) alone. To our knowledge, only one case-report has documented a histologically confirmed case of cerebral lesions associated with CD so far. CASE PRESENTATION: A 39-year-old right-handed woman with a history of CD was referred to our hospital with etiologically unexplained Gadolinium (Gd)-enhancing cortical lesions, triggering epileptic seizures. A CT-scan of the thorax and bronchoalveolar lavage found no signs of sarcoidosis. Lumbar punctures and laboratory testing found no underlying infection or coincidental autoimmune disorders and MRI-scans showed progression of lesion load. Consequently, the patient underwent stereotactic biopsy of a cortical lesion. Histological examination revealed a mixed lympho-histiocytic and tuberculoid granulomatous inflammation surrounding small vessels and no signs for infection. After exclusion of other granulomatous diseases and the typical histological findings we diagnosed a cerebral granulomatosis as a manifestation of CD. The patient was initially started on azathioprine, which had to be switched to corticosteroids and methotrexate because of an azathioprine related pancreatitis. The patient has not suffered any further epileptic seizures to date. CONCLUSION: Cerebral manifestation of CD is a possibly underreported entity that may respond well to immunosuppressive treatment. In contrast to earlier reports of cerebral manifestations in CD, our patient showed no coincident gastrointestinal symptoms indicating an activity of CD during the progression of cortical lesion load, suggesting that similar to other extra-intestinal manifestations in CD, the activity of gastrointestinal symptoms does not necessarily reflect the activity of CD associated cerebral vasculitis. Therefore, diagnosis and therapy of cerebral manifestation may be delayed when focusing on gastrointestinal symptoms alone.

Transcranial Direct Current Stimulation Enhances Motor Skill Learning but Not Generalization in Chronic Stroke.

BACKGROUND: Motor training alone or combined with transcranial direct current stimulation (tDCS) positioned over the motor cortex (M1) improves motor function in chronic stroke. Currently, understanding of how tDCS influences the process of motor skill learning after stroke is lacking. OBJECTIVE: To assess the effects of tDCS on the stages of motor skill learning and on generalization to untrained motor function. METHODS: In this randomized, sham-controlled, blinded study of 56 mildly impaired chronic stroke patients, tDCS (anode over the ipsilesional M1 and cathode on the contralesional forehead) was applied during 5 days of training on an unfamiliar, challenging fine motor skill task (sequential visual isometric pinch force task). We assessed online and offline learning during the training period and retention over the following 4 months. We additionally assessed the generalization to untrained tasks. RESULTS: With training alone (sham tDCS group), patients acquired a novel motor skill. This skill improved online, remained stable during the offline periods and was largely retained at follow-up. When tDCS was added to training (real tDCS group), motor skill significantly increased relative to sham, mostly in the online stage. Long-term retention was not affected by tDCS. Training effects generalized to untrained tasks, but those performance gains were not enhanced further by tDCS. CONCLUSIONS: Training of an unfamiliar skill task represents a strategy to improve fine motor function in chronic stroke. tDCS augments motor skill learning, but its additive effect is restricted to the trained skill.

Transcranial Electrical Brain Stimulation in Alert Rodents.

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in humans is transcranial direct current stimulation (tDCS). Less frequently, transcranial alternating current stimulation (tACS) or transcranial random noise stimulation (tRNS), a specific form of tACS using an electrical current applied randomly within a pre-defined frequency range, is used. The increase of noninvasive electrical brain stimulation research in humans, both for experimental and clinical purposes, has yielded an increased need for basic, mechanistic, safety studies in animals. This article describes a model for transcranial electrical brain stimulation (tES) through the intact skull targeting the motor system in alert rodents. The protocol provides step-by-step instructions for the surgical set-up of a permanent epicranial electrode socket combined with an implanted counter electrode on the chest. By placing a stimulation electrode into the epicranial socket, different electrical stimulation types, comparable to tDCS, tACS, and tRNS in humans, can be delivered. Moreover, the practical steps for tES in alert rodents are introduced. The applied current density, stimulation duration, and stimulation type may be chosen depending on the experimental needs. The caveats, advantages, and disadvantages of this set-up are discussed, as well as safety and tolerability aspects.

Anodal Transcranial Direct Current Stimulation Enhances Survival and Integration of Dopaminergic Cell Transplants in a Rat Parkinson Model.

Restorative therapy concepts, such as cell based therapies aim to restitute impaired neurotransmission in neurodegenerative diseases. New strategies to enhance grafted cell survival and integration are still needed to improve functional recovery. Anodal direct current stimulation (DCS) promotes neuronal activity and secretion of the trophic factor BDNF in the motor cortex. Transcranial DCS applied to the motor cortex transiently improves motor symptoms in Parkinson's disease (PD) patients. In this proof-of-concept study, we combine cell based therapy and noninvasive neuromodulation to assess whether neurotrophic support via transcranial DCS would enhance the restitution of striatal neurotransmission by fetal dopaminergic transplants in a rat Parkinson model. Transcranial DCS was applied daily for 20 min on 14 consecutive days following striatal transplantation of fetal ventral mesencephalic (fVM) cells derived from transgenic rat embryos ubiquitously expressing GFP. Anodal but not cathodal transcranial DCS significantly enhanced graft survival and dopaminergic reinnervation of the surrounding striatal tissue relative to sham stimulation. Behavioral recovery was more pronounced following anodal transcranial DCS, and behavioral effects correlated with the degree of striatal innervation. Our results suggest anodal transcranial DCS may help advance cell-based restorative therapies in neurodegenerative diseases. In particular, such an assistive approach may be beneficial for the already established cell transplantation therapy in PD.

Transcriptional profile of hippocampal dentate granule cells in four rat epilepsy models.

Global expression profiling of neurologic or psychiatric disorders has been confounded by variability among laboratories, animal models, tissues sampled, and experimental platforms, with the result being that few genes demonstrate consistent expression changes. We attempted to minimize these confounds by pooling dentate granule cell transcriptional profiles from 164 rats in seven laboratories, using three status epilepticus (SE) epilepsy models (pilocarpine, kainate, self-sustained SE), plus amygdala kindling. In each epilepsy model, RNA was harvested from laser-captured dentate granule cells from six rats at four time points early in the process of developing epilepsy, and data were collected from two independent laboratories in each rodent model except SSSE. Hierarchical clustering of differentially-expressed transcripts in the three SE models revealed complete separation between controls and SE rats isolated 1 day after SE. However, concordance of gene expression changes in the SE models was only 26-38% between laboratories, and 4.5% among models, validating the consortium approach. Transcripts with unusually highly variable control expression across laboratories provide a 'red herring' list for low-powered studies.

Epileptic Activity Increases Cerebral Amino Acid Transport Assessed by 18F-Fluoroethyl-l-Tyrosine Amino Acid PET: A Potential Brain Tumor Mimic.

O-(2-18F-fluoroethyl)-l-tyrosine (18F-FET) PET is a well-established method increasingly used for diagnosis, treatment planning, and monitoring in gliomas. Epileptic activity, frequently occurring in glioma patients, can influence MRI findings. Whether seizures also affect 18F-FET PET imaging is currently unknown. The aim of this retrospective analysis was to investigate the brain amino acid metabolism during epileptic seizures by 18F-FET PET and to elucidate the pathophysiologic background. METHODS: Ten patients with 11 episodes of serial seizures or status epilepticus, who underwent MRI and 18F-FET PET, were studied. The main diagnosis was glioma World Health Organization grade II-IV (n = 8); 2 patients suffered from nonneoplastic diseases. Immunohistochemical assessment of LAT1/LAT2/CD98 amino acid transporters was performed in seizure-affected cortex (n = 2) and compared with glioma tissues (n = 3). RESULTS: All patients exhibited increased seizure-associated strict gyral 18F-FET uptake, which was reversible in follow-up studies or negative shortly before and without any histologic or clinical signs of tumor recurrence. 18F-FET uptake corresponded to structural MRI changes, compatible with cortical vasogenic and cytotoxic edema, partial contrast enhancement, and hyperperfusion. Patients with prolonged postictal symptoms lasting up to 8 wk displayed intensive and widespread (≥ 1 lobe) cortical 18F-FET uptake. LAT1/LAT2/CD98 was strongly expressed in neurons and endothelium of seizure-affected brains and less in reactive astrocytosis. CONCLUSION: Seizure activity, in particular status epilepticus, increases cerebral amino acid transport with a strict gyral 18F-FET uptake pattern. Such periictal pseudoprogression represents a potential pitfall of 18F-FET PET and may mimic brain tumor. Our data also indicate a seizure-induced upregulation of neuronal, endothelial, and less astroglial LAT1/LAT2/CD98 amino acid transporter expression.

Glia: A Neglected Player in Non-invasive Direct Current Brain Stimulation.

Non-invasive electrical brain stimulation by application of direct current (DCS) promotes plasticity in neuronal networks in vitro and in in vivo. This effect has been mainly attributed to the direct modulation of neurons. Glia represents approximately 50% of cells in the brain. Glial cells are electrically active and participate in synaptic plasticity. Despite of that, effects of DCS on glial structures and on interaction with neurons are only sparsely investigated. In this perspectives article we review the current literature, present own dose response data and provide a framework for future research from two points of view: first, the direct effects of DCS on glia and second, the contribution of glia to DCS related neuronal plasticity.

Safety of Transcranial Direct Current Stimulation: Evidence Based Update 2016.

This review updates and consolidates evidence on the safety of transcranial Direct Current Stimulation (tDCS). Safety is here operationally defined by, and limited to, the absence of evidence for a Serious Adverse Effect, the criteria for which are rigorously defined. This review adopts an evidence-based approach, based on an aggregation of experience from human trials, taking care not to confuse speculation on potential hazards or lack of data to refute such speculation with evidence for risk. Safety data from animal tests for tissue damage are reviewed with systematic consideration of translation to humans. Arbitrary safety considerations are avoided. Computational models are used to relate dose to brain exposure in humans and animals. We review relevant dose-response curves and dose metrics (e.g. current, duration, current density, charge, charge density) for meaningful safety standards. Special consideration is given to theoretically vulnerable populations including children and the elderly, subjects with mood disorders, epilepsy, stroke, implants, and home users. Evidence from relevant animal models indicates that brain injury by Direct Current Stimulation (DCS) occurs at predicted brain current densities (6.3-13 A/m(2)) that are over an order of magnitude above those produced by conventional tDCS. To date, the use of conventional tDCS protocols in human trials (≤40 min, ≤4 milliamperes, ≤7.2 Coulombs) has not produced any reports of a Serious Adverse Effect or irreversible injury across over 33,200 sessions and 1000 subjects with repeated sessions. This includes a wide variety of subjects, including persons from potentially vulnerable populations.

Non-Invasive Electrical Brain Stimulation Montages for Modulation of Human Motor Function.

Non-invasive electrical brain stimulation (NEBS) is used to modulate brain function and behavior, both for research and clinical purposes. In particular, NEBS can be applied transcranially either as direct current stimulation (tDCS) or alternating current stimulation (tACS). These stimulation types exert time-, dose- and in the case of tDCS polarity-specific effects on motor function and skill learning in healthy subjects. Lately, tDCS has been used to augment the therapy of motor disabilities in patients with stroke or movement disorders. This article provides a step-by-step protocol for targeting the primary motor cortex with tDCS and transcranial random noise stimulation (tRNS), a specific form of tACS using an electrical current applied randomly within a pre-defined frequency range. The setup of two different stimulation montages is explained. In both montages the emitting electrode (the anode for tDCS) is placed on the primary motor cortex of interest. For unilateral motor cortex stimulation the receiving electrode is placed on the contralateral forehead while for bilateral motor cortex stimulation the receiving electrode is placed on the opposite primary motor cortex. The advantages and disadvantages of each montage for the modulation of cortical excitability and motor function including learning are discussed, as well as safety, tolerability and blinding aspects.

Time- but not sleep-dependent consolidation of tDCS-enhanced visuomotor skills.

Consolidation of motor skills after training can occur in a time- or sleep-dependent fashion. Recent studies revealed time-dependent consolidation as a common feature of visuomotor tasks. We have previously shown that anodal transcranial direct current stimulation (tDCS) in combination with repeated motor training benefits consolidation by the induction of offline skill gains in a complex visuomotor task, preventing the regular occurrence of skill loss between days. Here, we asked 2 questions: What is the time course of consolidation between days for this task and do exogenously induced offline gains develop as a function of time or overnight sleep? We found that both the development of offline skill loss in sham-stimulated subjects and offline skill gains induced by anodal tDCS critically depend on the passage of time after training, but not on overnight sleep. These findings support the view that tDCS interacts directly with the physiological consolidation process. However, in a control experiment, anodal tDCS applied after the training did not induce skill gains, implying that coapplication of tDCS and training is required to induce offline skill gains, pointing to the initiation of consolidation already during training.

Transcranial direct current stimulation over left and right DLPFC: Lateralized effects on planning performance and related eye movements.

Left and right dorsolateral prefrontal cortex (dlPFC) were recently found to be differentially affected by unilateral continuous theta-burst stimulation, reflected in an oppositional alteration of initial thinking time (ITT) in the Tower of London planning task. Here, we further explored this finding using bilateral transcranial direct current stimulation (tDCS) and simultaneous tracking of eye movements. Results revealed a decrease in ITT during concurrent cathodal tDCS of left dlPFC and anodal tDCS of right dlPFC. Eye-movement analyses showed that this facilitating tDCS effect was associated with the actual planning phase, thus reflecting a planning-specific impact of stimulation. For the reverse stimulation pattern of cathodal tDCS of right dlPFC and anodal tDCS of left dlPFC, an increase in gaze shifts was observed, without a significant impact on ITT. Taken together, these findings corroborate that enhanced planning performance can be obtained by boosting right dlPFC and dismantling the inhibitory impact of left dlPFC.

Polymer-based, flexible glutamate and lactate microsensors for in vivo applications.

We present a flexible microsensor, based on a polymer substrate, for multiparametric, electrochemical in vivo monitoring. The sensor strip with a microelectrode array at the tip was designed for insertion into tissue, for fast and localized online monitoring of physiological parameters. The microsystem fabrication on a wafer-level is based on a polyimide substrate and includes the patterning of platinum microelectrodes as well as epoxy and dry-film-resist insulation in a cost-effective thin-film and laminate process. A stable, electrodeposited silver/silver chloride reference electrode on-chip and a perm-selective membrane as an efficient interference rejection scheme are integrated on a wafer-level. Amperometric, electrochemical, enzyme-based biosensors for the neurotransmitter L-glutamate and the energy metabolite L-lactate have been developed. Hydrogel membranes or direct cross-linking as stable concepts for the enzyme immobilization are shown. Sensor performance including high selectivity, tailoring of sensitivity and long-term stability is discussed. For glutamate, a high sensitivity of 2.16 nAmm(-2) µM(-1) was found. For lactate, a variation in sensitivity between 2.6 and 32 nAmm(-2)mM(-1) was achieved by different membrane compositions. The in vivo application in an animal model is demonstrated by glutamate measurements in the brain of rats. Local glutamate alterations in the micromolar range and in nanoliter-range volumes can be detected and quantified with high reproducibility and temporal resolution. A novel, versatile platform for the integration of various electrochemical sensors on a small, flexible sensor strip for a variety of in vivo applications is presented.