Microelectrode Array Measurements Research Articles

Development of an evaluation method for addictive compounds based on electrical activity of human iPS cell-derived dopaminergic neurons using microelectrode array.

Addiction is known to occur through the consumption of substances such as pharmaceuticals, illicit drugs, food, alcohol and tobacco. These addictions can be viewed as drug addiction, resulting from the ingestion of chemical substances contained in them. Multiple neural networks, including the reward system, anti-reward/stress system and central immune system in the brain, are believed to be involved in the onset of drug addiction. Although various compound evaluations using microelectrode array (MEA) as an in vitro testing methods to evaluate neural activities have been conducted, methods for assessing addiction have not been established. In this study, we aimed to develop an in vitro method for assessing the addiction of compounds, as an alternative to animal experiments, using human iPS cell-derived dopaminergic neurons with MEA measurements. MEA data before and after chronic exposure revealed specific changes in addictive compounds compared to non-addictive compounds, demonstrating the ability to estimate addiction of compound. Additionally, conducting gene expression analysis on cultured samples after the tests revealed changes in the expression levels of various receptors (nicotine, dopamine and GABA) due to chronic administration of addictive compounds, suggesting the potential interpretation of these expression changes as addiction-like responses in MEA measurements. The addiction assessment method using MEA measurements in human iPS cell-derived dopaminergic neurons conducted in this study proves effective in evaluating addiction of compounds on human neural networks.

Contraindicated Drug Responses in Dravet Syndrome Brain Organoids Utilizing Micro Electrode Array Assessment Methods

Ensuring drug safety for patients with specific neurological disorders is of paramount importance. For instance, certain antiepileptic drugs (AEDs) are contraindicated in Dravet Syndrome (DS), which is characterized by a deficiency in Na+ channel function. Constructing in vitro assessment methods capable of detecting contraindicated drug responses and medication effects on neurons derived from DS patients is highly anticipated for drug safety assessment and therapeutic innovation. This study used micro electrode array (MEA) measurements with low-frequency analysis on human iPSC-derived DS organoids to investigate AED responses. When exposed to the contraindicated drugs carbamazepine and phenytoin, the number of network oscillations increased in DS organoids while maintaining oscillation intensity. Furthermore, carbamazepine administration appeared to enhance activities beyond oscillations which is partially consistent with findings in the DS mouse model. Conversely, treatment with the therapeutic drug sodium valproate resulted in a similar decrease in activity both in healthy and DS organoids. The frequency characteristics of spontaneous firings and AEDs responsiveness in DS organoids demonstrated partial correlation with typical electroencephalography patterns observed in vivo. In conclusion, this study, employing MEA measurements with low-frequency analysis, revealed contraindicated drug responses and disease-specific functional characteristics in DS organoids, effective for DS patient safety assessment, precision medicine, and antiepileptic drug screening.

Evaluation of neurotoxicity for pesticide-related compounds in human iPS cell-derived neurons using microelectrode array

In vivo evaluations of chemicals in neurotoxicity have certain limitations due to the considerable time and cost required, necessity of extrapolation from rodents to humans, and limited information on toxicity mechanisms. To address this issue, the development of in vitro test methods using new approach methodologies (NAMs) is important to evaluate the chemicals in neurotoxicity. Microelectrode array (MEA) allows the assessment of changes in neural network activity caused by compound administration. However, studies on compound evaluation criteria are scarce. In this study, we evaluated the impact of pesticides on neural activity using MEA measurements of human iPSC-derived neurons. A principal component analysis was performed on the electrical physiological parameters obtained by MEA measurements, and the influence of excessive neural activity due to compound addition was defined using the standard deviation of neural activity with solvent addition as the reference. By using known seizurogenic compounds as positive controls for neurotoxicity in MEA and evaluating pesticides with insufficient verification of their neurotoxicity in humans, we demonstrated that these pesticides exhibit neurotoxicity in humans. In conclusion, our data suggest that the neurotoxicity evaluation method in human iPSC neurons using MEA measurements could be one of the in vitro neurotoxicity test methods that could replace animal experiments.

High‐Aspect‐Ratio Nanoelectrodes Enable Long‐Term Recordings of Neuronal Signals with Subthreshold Resolution

The further development of neurochips requires high-density and high-resolution recordings that also allow neuronal signals to be observed over a long period of time. Expanding fields of network neuroscience and neuromorphic engineering demand the multiparallel and direct estimations of synaptic weights, and the key objective is to construct a device that also records subthreshold events. Recently, 3D nanostructures with a high aspect ratio have become a particularly suitable interface between neurons and electronic devices, since the excellent mechanical coupling to the neuronal cell membrane allows very high signal-to-noise ratio recordings. In the light of an increasing demand for a stable, noninvasive and long-term recording at subthreshold resolution, a combination of vertical nanostraws with nanocavities is presented. These structures provide a spontaneous tight coupling with rat cortical neurons, resulting in high amplitude sensitivity and postsynaptic resolution capability, as directly confirmed by combined patch-clamp and microelectrode array measurements.

Principal Component Analysis to Distinguish Seizure Liability of Drugs in Human iPS Cell-Derived Neurons

Abstract Screening for drug discovery targeting the central nervous system requires the establishment of efficient and highly accurate toxicity test methods that can reduce costs and time while maintaining high throughput using the function of an in vitro neural network. In particular, an evaluation system using a human-derived neural network is desirable in terms of species difference. Despite the attention, the microelectrode array (MEA) is attracting among the evaluation systems that can measure in vitro neural activity, an effective analysis method for evaluation of toxicity and mechanism of action has not yet been established. Here we established analytical parameters and multivariate analysis method capable of detecting seizure liability of drugs using MEA measurement of human iPS cell-derived neurons. Using the spike time series data of all drugs, we established periodicity as a new analytical parameter. Periodicity has facilitated the detection of responses to seizurogenic drugs, previously difficult to detect with conventional analytical parameters. By constructing a multivariate analytical method that identifies a parameter set that achieves an arbitrary condition, we found that the parameter set comprising total spikes, maximum frequency (MF), inter- MF interval (IMFI), coefficient of variance of IMFI, and periodicity can uniformly detect the seizure liability of seizurogenic drugs with different mechanisms of action. Seizurogenic drugs were suggested to increase the regularity of the network burst in MEA measurements in human iPS cell-derived neurons.

Culture of Rat Primary Cortical Cells for Microelectrode Array (MEA) Recordings to Screen for Acute and Developmental Neurotoxicity.

Neurotoxicity testing of chemicals, drug candidates, and environmental pollutants still relies on extensive in vivo studies that are very costly, time-consuming, and ethically debated due to the large number of animals typically used. Currently, rat primary cortical cultures are widely used for in vitro neurotoxicity studies, as they closely resemble the in vitro brain with respect to the diversity of cell types, their physiological functions, and the pathological processes that they undergo. Common in vitro assays for neurotoxicity screening often focus on very target-specific endpoints such as morphological, biochemical, or electrophysiological changes, and such narrow focus can hamper translation and interpretation. Microelectrode array (MEA) recordings provide a non-invasive platform for extracellular recording of electrical activity of cultured neuronal cells, thereby enabling the evaluation of changes in neuronal (network) function as a sensitive and integrated endpoint for neurotoxicity screening. Here, we describe an in vitro approach for assessing changes in neuronal network function as a measure for neurotoxicity, using rat primary cortical cultures grown on MEAs. We provide a detailed protocol for the culture of rat primary cortical cells, and describe several experimental procedures to address acute, subchronic, and chronic exposure scenarios. We additionally describe the steps for processing and analyzing MEA and cell viability data. © 2021 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Isolation and culture of rat primary cortical cells on 48-well MEA plates Support Protocol 1: Pretreatment and washing of 48-well MEA plates before first use or for re-use Support Protocol 2: Coating of 48-well MEA plates with 0.1% PEI solution Basic Protocol 2: MEA measurements during acute exposure Alternate Protocol 1: MEA measurements during subchronic exposure Alternate Protocol 2: MEA measurements during chronic exposure Support Protocol 3: Determination of cell viability after MEA experiments Basic Protocol 3: MEA data processing Basic Protocol 4: Analyzing MEA experiments after acute and subchronic exposure Alternate Protocol 3: Analyzing MEA experiments after chronic exposure.

Ictal Inhibition: Sync Globally, Slack Locally.

Role of Paroxysmal Depolarization in Focal Seizure Activity Tryba AK, Merricks EM, Lee S, et al. J Neuroph. 2019;122(5):1861-1873. doi.org/10.1152/jn.00392.2019We analyze the role of inhibition in sustaining focal epileptic seizure activity. We review ongoing seizure activity at the mesoscopic scale that can be observed with microelectrode arrays as well as at the macroscale of standard clinical electroencephalogram. We provide clinical, experimental, and modeling data to support the hypothesis that paroxysmal depolarization (PD) is a critical component of the ictal machinery. We present dual-patch recordings in cortical cultures showing reduced synaptic transmission associated with presynaptic occurrence of PD, and we find that the PD threshold is cell size related. We further find evidence that optically evoked PD activity in parvalbumin neurons can promote propagation of neuronal excitation in neocortical networks in vitro. Spike sorting results from microelectrode array measurements around ictal wave propagation in human focal seizures demonstrate a strong increase in putative inhibitory firing with an approaching excitatory wave, followed by a sudden reduction of firing at passage. At the macroscopic level, we summarize evidence that this excitatory ictal wave activity is strongly correlated with oscillatory activity across a centimeter-sized cortical network. We summarize Wilson–Cowan-type modeling showing how inhibitory function is crucial for this behavior. Our findings motivated us to develop a network motif of neurons in silico, governed by a reduced version of the Hodgkin–Huxley formalism, to show how feedforward, feedback, PD, and local failure of inhibition contribute to observed dynamics across network scales. The presented multidisciplinary evidence suggests that the PD not only is a cellular marker or epiphenomenon but actively contributes to seizure activity. NEW & NOTEWORTHY: We present mechanisms of ongoing focal seizures across meso- and macroscales of microelectrode array and standard clinical recordings, respectively. We find modeling, experimental, and clinical evidence for a dual role of inhibition across these scales: local failure of inhibition allows propagation of a mesoscopic ictal wave, whereas inhibition elsewhere remains intact and sustains macroscopic oscillatory activity. We present evidence for PD as a mechanism behind this dual role of inhibition in shaping ictal activity.

Development of 3D neuromuscular bioactuators.

Neuronal control of skeletal muscle bioactuators represents a critical milestone toward the realization of future biohybrid machines that may generate complex motor patterns and autonomously navigate through their environment. Animals achieve these feats using neural networks that generate robust firing patterns and coordinate muscle activity through neuromuscular units. Here, we designed a versatile 3D neuron-muscle co-culture platform to serve as a test-bed for neuromuscular bioactuators. We used our platform in conjunction with microelectrode array electrophysiology to study the roles of synergistic interactions in the co-development of neural networks and muscle tissues. Our platform design enables co-culture of a neuronal cluster with up to four target muscle actuators, as well as quantification of muscle contraction forces. Using engineered muscle tissue targets, we first demonstrated the formation of functional neuromuscular bioactuators. We then investigated possible roles of long-range interactions in neuronal outgrowth patterns and observed preferential outgrowth toward muscles compared to the acellular matrix or fibroblasts, indicating muscle-specific chemotactic cues acting on motor neurons. Next, we showed that co-cultured muscle strips exhibited significantly higher spontaneous contractility as well as improved sarcomere assembly compared to muscles cultured alone. Finally, we performed microelectrode array measurements on neuronal cultures, which revealed that muscle-conditioned medium enhances overall neural firing rates and the emergence of synchronous bursting patterns. Overall, our study illustrates the significance of neuron-muscle cross talk for the in vitro development of neuromuscular bioactuators.

HiPSCs Derived Cardiac Cells for Drug and Toxicity Screening and Disease Modeling: What Micro- Electrode-Array Analyses Can Tell Us.

Human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CM) have been intensively used in drug development and disease modeling. Since iPSC-cardiomyocyte (CM) was first generated, their characterization has become a major focus of research. Multi-/micro-electrode array (MEA) systems provide a non-invasive user-friendly platform for detailed electrophysiological analysis of iPSC cardiomyocytes including drug testing to identify potential targets and the assessment of proarrhythmic risk. Here, we provide a systematical overview about the physiological and technical background of micro-electrode array measurements of iPSC-CM. We introduce the similarities and differences between action- and field potential and the advantages and drawbacks of MEA technology. In addition, we present current studies focusing on proarrhythmic side effects of novel and established compounds combining MEA systems and iPSC-CM. MEA technology will help to open a new gateway for novel therapies in cardiovascular diseases while reducing animal experiments at the same time.

Role of paroxysmal depolarization in focal seizure activity.

We analyze the role of inhibition in sustaining focal epileptic seizure activity. We review ongoing seizure activity at the mesoscopic scale that can be observed with microelectrode arrays as well as at the macroscale of standard clinical EEG. We provide clinical, experimental, and modeling data to support the hypothesis that paroxysmal depolarization (PD) is a critical component of the ictal machinery. We present dual-patch recordings in cortical cultures showing reduced synaptic transmission associated with presynaptic occurrence of PD, and we find that the PD threshold is cell size related. We further find evidence that optically evoked PD activity in parvalbumin neurons can promote propagation of neuronal excitation in neocortical networks in vitro. Spike sorting results from microelectrode array measurements around ictal wave propagation in human focal seizures demonstrate a strong increase in putative inhibitory firing with an approaching excitatory wave, followed by a sudden reduction of firing at passage. At the macroscopic level, we summarize evidence that this excitatory ictal wave activity is strongly correlated with oscillatory activity across a centimeter-sized cortical network. We summarize Wilson-Cowan-type modeling showing how inhibitory function is crucial for this behavior. Our findings motivated us to develop a network motif of neurons in silico, governed by a reduced version of the Hodgkin-Huxley formalism, to show how feedforward, feedback, PD, and local failure of inhibition contribute to observed dynamics across network scales. The presented multidisciplinary evidence suggests that the PD not only is a cellular marker or epiphenomenon but actively contributes to seizure activity.NEW & NOTEWORTHY We present mechanisms of ongoing focal seizures across meso- and macroscales of microelectrode array and standard clinical recordings, respectively. We find modeling, experimental, and clinical evidence for a dual role of inhibition across these scales: local failure of inhibition allows propagation of a mesoscopic ictal wave, whereas inhibition elsewhere remains intact and sustains macroscopic oscillatory activity. We present evidence for paroxysmal depolarization as a mechanism behind this dual role of inhibition in shaping ictal activity.

Composite Biomarkers Derived from Micro-Electrode Array Measurements and Computer Simulations Improve the Classification of Drug-Induced Channel Block.

The Micro-Electrode Array (MEA) device enables high-throughput electrophysiology measurements that are less labor-intensive than patch-clamp based techniques. Combined with human-induced pluripotent stem cells cardiomyocytes (hiPSC-CM), it represents a new and promising paradigm for automated and accurate in vitro drug safety evaluation. In this article, the following question is addressed: which features of the MEA signals should be measured to better classify the effects of drugs? A framework for the classification of drugs using MEA measurements is proposed. The classification is based on the ion channels blockades induced by the drugs. It relies on an in silico electrophysiology model of the MEA, a feature selection algorithm and automatic classification tools. An in silico model of the MEA is developed and is used to generate synthetic measurements. An algorithm that extracts MEA measurements features designed to perform well in a classification context is described. These features are called composite biomarkers. A state-of-the-art machine learning program is used to carry out the classification of drugs using experimental MEA measurements. The experiments are carried out using five different drugs: mexiletine, flecainide, diltiazem, moxifloxacin, and dofetilide. We show that the composite biomarkers outperform the classical ones in different classification scenarios. We show that using both synthetic and experimental MEA measurements improves the robustness of the composite biomarkers and that the classification scores are increased.

The effect of ɑ-synuclein preformed fibrils on network activity development in cultured cortical neurons

The effect of ɑ-synuclein preformed fibrils on network activity development in cultured cortical neurons

Linking kindling to increased glutamate release in the dentate gyrus of the hippocampus through the STXBP5/tomosyn-1 gene.

IntroductionIn kindling, repeated electrical stimulation of certain brain areas causes progressive and permanent intensification of epileptiform activity resulting in generalized seizures. We focused on the role(s) of glutamate and a negative regulator of glutamate release, STXBP5/tomosyn‐1, in kindling.MethodsStimulating electrodes were implanted in the amygdala and progression to two successive Racine stage 5 seizures was measured in wild‐type and STXBP5/tomosyn‐1−/− (Tom−/−) animals. Glutamate release measurements were performed in distinct brain regions using a glutamate‐selective microelectrode array (MEA).ResultsNaïve Tom−/− mice had significant increases in KCl‐evoked glutamate release compared to naïve wild type as measured by MEA of presynaptic release in the hippocampal dentate gyrus (DG). Kindling progression was considerably accelerated in Tom−/− mice, requiring fewer stimuli to reach a fully kindled state. Following full kindling, MEA measurements of both kindled Tom+/+ and Tom−/− mice showed significant increases in KCl‐evoked and spontaneous glutamate release in the DG, indicating a correlation with the fully kindled state independent of genotype. Resting glutamate levels in all hippocampal subregions were significantly lower in the kindled Tom−/− mice, suggesting possible changes in basal control of glutamate circuitry in the kindled Tom−/− mice.ConclusionsOur studies demonstrate that increased glutamate release in the hippocampal DG correlates with acceleration of the kindling process. Although STXBP5/tomosyn‐1 loss increased evoked glutamate release in naïve animals contributing to their prokindling phenotype, the kindling process can override any attenuating effect of STXBP5/tomosyn‐1. Loss of this “braking” effect of STXBP5/tomosyn‐1 on kindling progression may set in motion an alternative but ultimately equally ineffective compensatory response, detected here as reduced basal glutamate release.

Joint analysis of extracellular spike waveforms and neuronal network bursts

BackgroundNeuronal networks are routinely assessed based on extracellular electrophysiological microelectrode array (MEA) measurements by spike sorting, and spike and burst statistics. We propose to jointly analyze sorted spikes and detected bursts, and hypothesize that the obtained spike type compositions of the bursts can provide new information on the functional networks. New methodSpikes are detected and sorted to obtain spike types and bursts are detected. In the proposed joint analysis, each burst spike is associated with a spike type, and the spike type compositions of the bursts are assessed. ResultsThe proposed method was tested with simulations and MEA measurements of in vitro human stem cell derived neuronal networks under different pharmacological treatments. The results show that the treatments altered the spike type compositions of the bursts. For example, 6-cyano-7-nitroquinoxaline-2,3-dione almost completely abolished two types of spikes which had composed the bursts in the baseline, while bursts of spikes of two other types appeared more frequently. This phenomenon was not observable by spike sorting or burst analysis alone, but was revealed by the proposed joint analysis. Comparison with existing methodsThe existing methods do not provide the information obtainable with the proposed method: for the first time, the spike type compositions of bursts are analyzed. ConclusionsWe showed that the proposed method provides useful and novel information, including the possible changes in the spike type compositions of the bursts due to external factors. Our method can be employed on any data exhibiting sortable action potential waveforms and detectable bursts.

Generation of Bioartificial Heart Tissue by Combining a Three-Dimensional Gel-Based Cardiac Construct with Decellularized Small Intestinal Submucosa

The in vitro generation of a bioartificial cardiac construct (CC) represents a promising tool for the repair of ischemic heart tissue. Several approaches to engineer cardiac tissue in vitro have been conducted. The main drawback of these studies is the insufficient size of the resulting construct for clinical applications. The focus of this study was the generation of an artificial three-dimensional (3D), contractile, and suturable myocardial patch by combining a gel-based CC with decellularized porcine small intestinal submucosa (SIS), thereby engineering an artificial tissue of 11 cm² in size. The alignment and morphology of rat neonatal cardiomyocytes (rCMs) in SIS-CC complexes were investigated as well as the re-organization of primary endothelial cells which were co-isolated in the rCM preparation. The ability of a rat heart endothelial cell line (RHE-A) to re-cellularize pre-existing vessel structures within the SIS or a biological vascularized matrix (BioVaM) was determined. SIS-CC contracted spontaneously, uniformly, and rhythmically with an average rate of 200 beats/min in contrast to undirected contractions observed in CC without SIS support. rCM exhibited an elongated morphology with well-defined sarcomeric structures oriented along the longitudinal axis in the SIS-CC, whereas round-shaped and random-arranged rCM were observed in CC. Electric coupling of rCM was demonstrated by microelectrode array measurements. A dense network of CD31⁺/eNOS⁺ cells was detected as permeating the whole construct. Superficial supplementation of RHE-A cells to SIS-CC led to the migration of these cells through the CC, resulting in the re-population of pre-existing vessel structures within the decelluarized SIS. By infusion of RHE-A cells into the BioVaM venous and arterial pedicles, a re-population of the BioVaM vessel bed as well as distribution of RHE-A cells throughout the CC was achieved. Rat endothelial cells within the CC were in contact with RHE-A cells. Ingrowth and formation of a network by endothelial cells infused through the BioVaM represent a promising step toward engineering a functional perfusion system, enabling the engineering of vascularized and well-nourished 3D CC of dimensions relevant for therapeutic heart repair.

Serotonin Augments Gut Pacemaker Activity via 5-HT3 Receptors

Serotonin (5-hydroxytryptamine: 5-HT) affects numerous functions in the gut, such as secretion, muscle contraction, and enteric nervous activity, and therefore to clarify details of 5-HT's actions leads to good therapeutic strategies for gut functional disorders. The role of interstitial cells of Cajal (ICC), as pacemaker cells, has been recognised relatively recently. We thus investigated 5-HT actions on ICC pacemaker activity. Muscle preparations with myenteric plexus were isolated from the murine ileum. Spatio-temporal measurements of intracellular Ca2+ and electric activities in ICC were performed by employing fluorescent Ca2+ imaging and microelectrode array (MEA) systems, respectively. Dihydropyridine (DHP) Ca2+ antagonists and tetrodotoxin (TTX) were applied to suppress smooth muscle and nerve activities, respectively. 5-HT significantly enhanced spontaneous Ca2+ oscillations that are considered to underlie electric pacemaker activity in ICC. LY-278584, a 5-HT3 receptor antagonist suppressed spontaneous Ca2+ activity in ICC, while 2-methylserotonin (2-Me-5-HT), a 5-HT3 receptor agonist, restored it. GR113808, a selective antagonist for 5-HT4, and O-methyl-5-HT (O-Me-5-HT), a non-selective 5-HT receptor agonist lacking affinity for 5-HT3 receptors, had little effect on ICC Ca2+ activity. In MEA measurements of ICC electric activity, 5-HT and 2-Me-5-HT caused excitatory effects. RT-PCR and immunostaining confirmed expression of 5-HT3 receptors in ICC. The results indicate that 5-HT augments ICC pacemaker activity via 5-HT3 receptors. ICC appear to be a promising target for treatment of functional motility disorders of the gut, for example, irritable bowel syndrome.

Human cell-based micro electrode array platform for studying neurotoxicity

At present, most of the neurotoxicological analyses are based on in vitro and in vivo models utilizing animal cells or animal models. In addition, the used in vitro models are mostly based on molecular biological end-point analyses. Thus, for neurotoxicological screening, human cell-based analysis platforms in which the functional neuronal networks responses for various neurotoxicants can be also detected real-time are highly needed. Microelectrode array (MEA) is a method which enables the measurement of functional activity of neuronal cell networks in vitro for long periods of time. Here, we utilize MEA to study the neurotoxicity of methyl mercury chloride (MeHgCl, concentrations 0.5–500 nM) to human embryonic stem cell (hESC)-derived neuronal cell networks exhibiting spontaneous electrical activity. The neuronal cell cultures were matured on MEAs into networks expressing spontaneous spike train-like activity before exposing the cells to MeHgCl for 72 h. MEA measurements were performed acutely and 24, 48, and 72 h after the onset of the exposure. Finally, exposed cells were analyzed with traditional molecular biological methods for cell proliferation, cell survival, and gene and protein expression. Our results show that 500 nM MeHgCl decreases the electrical signaling and alters the pharmacologic response of hESC-derived neuronal networks in delayed manner whereas effects can not be detected with qRT-PCR, immunostainings, or proliferation measurements. Thus, we conclude that human cell-based MEA platform is a sensitive online method for neurotoxicological screening.

Microelectrode array evaluation of gut pacemaker activity in wild-type and [formula omitted] mice

Interstitial cells of Cajal in the myenteric plexus region (ICC-MyP) form a network and generate basal pacemaking electrical activity. This morphological feature leads us to believe that these cells may be essential for the coordinating actions of gastrointestinal (GI) motility. We aim to propose a new method for functional assessment of ICC electrical activity and its network. Field potentials in a ∼1 mm 2 region were simultaneously measured using an 8 × 8 microelectrode array (MEA) with a polar distance of 150 μm. The extracellular solution contained nifedipine and tetrodotoxin (TTX) to suppress activities of smooth muscle cells and neurons, respectively. We compared spatial electrical activities between ileal muscle preparations from wild-type (WT) and W / W v mice. In spatio-temporal analyses, basal electrical activities were well synchronized with a propagation delay in WT, while those in W / W v were small in amplitude and irregular in occurrence. The power spectrum in WT had a prominent peak corresponding to the frequency of ICC-MyP pacemaker activity, while that of W / W v lacked it. Consequently, the ratio of the spectral power in 9.4–27.0 cpm was significantly larger in WT than in W / W v . In conclusion, MEA measurements demonstrated that the network-forming ICC-MyP not only generates but also coordinates basal electrical activities. Disorders of GI motility based on morphological and functional impairments of ICC network with the range of several hundreds of micrometers, could be uncovered in future extensive studies.

High resolution gravimetric, optical and electrochemical investigations of microbial biofilm formation in aqueous systems

The formation of microbial biofilms on metallic surfaces was studied by combination of gravimetric, optical and electrochemical methods with high local resolution in dependence on colonization time and biofilm thickness. The use of electrochemical quartz crystal microbalance (EQCM) flowcells allowed simultaneous and continuous in situ monitoring of biofilm formation and resulting open circuit potential U R. Combination of macroscopic and microscopic measurements of U R versus t with confocal scanning laser microscopy (CSLM) offered high resolution, destruction free characterization of the electrochemical influences of growing biofilms and their subsequent changes during disinfection and cleaning processes. Cyclovoltammograms (CVs) on macroscopic Pt electrodes after biofilm colonization proved a strong inhibition of H-adsorption and oxide growth while O 2-reduction was catalyzed. Repeated cycling reduced the biofilm effects until platinum behavior, except for the H-adsorption, could be observed again. Influences of biofilm thickness and age played an inferior role in these effects. Calculation of net charges showed a restriction of electrochemical detection to a sub-μm layer on the electrode surface. Microelectrode array measurements yielded no direct correlation between the localization of colonies and local shift of open circuit potential U R for several film proportions. Disinfection with 70% EtOH killed the organisms, but left U R unchanged. Cleaning with 10% H 2O 2 removed all biomass, resulting in clean microelectrode surfaces and restoration of U R values to those of pure metal.