BACKGROUND: The local pH change mediated by the pathogenic bacterial species Streptococcus mutans plays a significant role in the corrosion of hydroxyapatite (HA) present in the tooth in the dynamic oral cavity. The acid produced by the bacteria decreases the local pH and releases Ca2+ ions from the HA. We studied the bacteria-mediated demineralization of HA by scanning electrochemical microscopy (SECM) after growing S. mutans biofilm on HA for 7 days.
RESULTS: We notably developed a triple-function SECM-compatible tip that could be positioned above the biofilm. It can also measure the pH and [Ca2+] change simultaneously above the biofilm-HA substrate. The triple-function SECM tip is a combination of a potentiometric pH sensor deposited with iridium oxide and a dual-function carbon-based Ca2+ ion-selective membrane electrode with a slope of 67 mV/pH and 34.3 mV/log [Ca2+], respectively. The distance-controlled triple-function SECM tip monitored real-time pH and [Ca2+] changes 30 μm above the S. mutans biofilm. The high temporal resolution pH data demonstrated that after approximately 20 min of sucrose addition, S. mutans started to produce acid to titrate the solution buffer, causing a pH change from 7.2 to 6.5 for HA and from 7.2 to 5 for the glass substrate. We observed that, after 30 min of acid production, ∼300 μM of Ca2+ ions were increased at pH 6.5 above the biofilm surface as a result of the pH change in the local microenvironment. After the release of Ca2+ from HA, the pH environment again shifted toward the neutral side, from 6.5 to 7.2. Therefore, precipitation of Ca2+ happens at the top of the biofilm, thus corroding the HA from underneath. For a glass substrate, in contrast, no Ca2+ ions were released, and the pH did not change back to 7.2. We were able to observe the dynamics of the HA demineralization-remineralization process simultaneously with our newly developed triple-function SECM tip or microprobe.
CONCLUSIONS: This technique could notably advance the study of similar complex processes, such as bacteria-mediated corrosion in biomedical and environmental contexts.

Exocytosis of a single cell has been extensively researched in recent years due to its close association with numerous diseases. However, current methods only investigate exocytosis at either the single-cell or multiple-cell level, and a method for simultaneously studying exocytosis at both levels has yet to be established. In this study, a combined device incorporating ultramicroelectrode (UME) electrochemistry and surface plasmon resonance (SPR) was developed, enabling the simultaneous monitoring of single-cell and multiple-cell exocytosis. PC12 cells were cultured directly on the SPR sensing Au film, with a carboxylated carbon nanopipette (c-CNP) electrode employed for electrochemical detection in the SPR reaction cell. Upon exocytosis, the released dopamine diffuses onto the inner wall of c-CNP, undergoing an electrochemical reaction to generate a current peak. Concurrently, exocytosis can also induce changes in the refractive index of the Au film surface, leading to the SPR signal. Consequently, the device enables real-time monitoring of exocytosis from both single and multiple cells with a high spatiotemporal resolution. The c-CNP electrode exhibited excellent resistance to protein contamination, high sensitivity for dopamine detection, and the capability to continuously monitor dopamine exocytosis over an extended period. Analysis of both SPR and electrochemical signals revealed a positive correlation between changes in the SPR signal and the frequency of exocytosis. This study introduces a novel method and platform for the simultaneous investigation of single-cell and multiple-cell exocytosis.

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are frequently used for preclinical cardiotoxicity testing and remain an important tool for confirming model-based predictions of drug effects in accordance with the comprehensive in vitro proarrhythmia assay (CiPA). Despite the considerable benefits hiPSC-CMs provide, concerns surrounding experimental reproducibility have emerged. We investigated the effects of temporal changes and experimental parameters on hiPSC-CM electrophysiology. iCell cardiomyocytes2 were cultured and biosignals were acquired using a microelectrode array (MEA) system (2-14 days). Continuous recordings revealed a 22.6% increase in the beating rate and 7.7% decrease in the field potential duration (FPD) during a 20-min equilibration period. Location-specific differences across a multiwell plate were also observed, with iCell cardiomyocytes2 in the outer rows beating 8.8 beats/min faster than the inner rows. Cardiac endpoints were also impacted by cell culture duration; from 2 to 14 days, the beating rate decreased (-12.7 beats/min), FPD lengthened (+257 ms), and spike amplitude increased (+3.3 mV). Cell culture duration (4-10 days) also impacted cardiomyocyte drug responsiveness (E-4031, nifedipine, isoproterenol). qRT-PCR results suggest that daily variations in cardiac metrics may be linked to the continued maturation of hiPSC-CMs in culture (2-30 days). Daily experiments were also repeated using a second cell line (Cor.4U). Collectively, our study highlights multiple sources of variability to consider and address when performing hiPSC-CM MEA studies. To improve reproducibility and data interpretation, MEA-based studies should establish a standardized protocol and report key experimental conditions (e.g., cell line, culture time, equilibration time, electrical stimulation settings, and raw data values).NEW & NOTEWORTHY We demonstrate that iCell cardiomyocytes2 electrophysiology measurements are impacted by deviations in experimental techniques including electrical stimulation protocols, equilibration time, well-to-well variability, and length of hiPSC-CM culture. Furthermore, our results indicate that hiPSC-CM drug responsiveness changes within the first 2 wk following defrost.

This paper discusses the electrochemical properties of thin-film, planar, titanium-platinum (Ti-Pt) microelectrodes fabricated using glass or silicon substrates and compares their performance to the classic platinum (Pt) microelectrodes embedded in glass. To analyze the possible differences coming both from the size of the tested electrodes as well as from the substrate, short- and long-term electrochemical tests were performed on selected water electrolytes (KCl, HCl, KOH). To study the electrochemical response of the electrodes, the cyclic voltammetry (CV) measurements were carried out at different scanning rates (from 5 to 200 mV/s). Long-term tests were also conducted, including one thousand cycles with a 100 mV/s scan rate to investigate the stability of the tested electrodes. Before and after electrochemical measurements, the film morphology was analyzed using a scanning electron microscope (SEM). The good quality of the thin-film Pt electrodes and the high repeatability in electrochemical response have been shown. There are minor differences in standard deviation values taken from electrochemical measurements, comparing thin-film and wire-based electrodes. Damages or any changes on the electrodes\' surfaces were revealed by SEM observations after long-term electrochemical tests.

Microelectrodes serve as a fundamental tool in electrophysiology research throughout the nervous system, providing a means of exploring neural function with a high resolution of neural firing information. We constructed a hybrid computational model using the finite element method and multicompartment cable models to explore factors that contribute to extracellular voltage waveforms that are produced by sensory pseudounipolar neurons, specifically smaller A-type neurons, and that are recorded by microelectrodes in dorsal root ganglia. The finite element method model included a dorsal root ganglion, surrounding tissues, and a planar microelectrode array. We built a multicompartment neuron model with multiple trajectories of the glomerular initial segment found in many A-type sensory neurons. Our model replicated both the somatic intracellular voltage profile of Aδ low-threshold mechanoreceptor neurons and the unique extracellular voltage waveform shapes that are observed in experimental settings. Results from this model indicated that tortuous glomerular initial segment geometries can introduce distinct multiphasic properties into a neuron\'s recorded waveform. Our model also demonstrated how recording location relative to specific microanatomical components of these neurons, and recording distance from these components, can contribute to additional changes in the multiphasic characteristics and peak-to-peak voltage amplitude of the waveform. This knowledge may provide context for research employing microelectrode recordings of pseudounipolar neurons in sensory ganglia, including functional mapping and closed-loop neuromodulation. Furthermore, our simulations gave insight into the neurophysiology of pseudounipolar neurons by demonstrating how the glomerular initial segment aids in increasing the resistance of the stem axon and mitigating rebounding somatic action potentials.NEW & NOTEWORTHY We built a computational model of sensory neurons in the dorsal root ganglia to investigate factors that influence the extracellular waveforms recorded by microelectrodes. Our model demonstrates how the unique structure of these neurons can lead to diverse and often multiphasic waveform profiles depending on the location of the recording contact relative to microanatomical neural components. Our model also provides insight into the neurophysiological function of axon glomeruli that are often present in these neurons.

MicroGraphited-Diamond-Multi Electrode Arrays (μG-D-MEAs) can be successfully used to reveal, in real time, quantal exocytotic events occurring from many individual neurosecretory cells and/or from many neurons within a network. As μG-D-MEAs arrays are patterned with up to 16 sensing microelectrodes, each of them recording large amounts of data revealing the exocytotic activity, the aim of this work was to support an adequate analysis code to speed up the signal detection. The cutting-edge technology of microGraphited-Diamond-Multi Electrode Arrays (μG-D-MEAs) has been implemented with an automated analysis code (APE, Amperometric Peak Analysis) developed using Matlab R2022a software to provide easy and accurate detection of amperometric spike parameters, including the analysis of the pre-spike foot that sometimes precedes the complete fusion pore dilatation. Data have been acquired from cultured PC12 cells, either collecting events during spontaneous exocytosis or after L-DOPA incubation. Validation of the APE code was performed by comparing the acquired spike parameters with those obtained using Quanta Analysis (Igor macro) by Mosharov et al.

BACKGROUND: In the framework of the IMI2-NeuroDeRisk consortium, three in vitro electrophysiology assays were compared to improve preclinical prediction of seizure-inducing liabilities.
METHODS: Two cell models, primary rat cortical neurons and human induced pluripotent stem cell (hiPSC)-derived glutamatergic neurons co-cultured with hiPSC-derived astrocytes were tested on two different microelectrode array (MEA) platforms, Maestro Pro (Axion Biosystems) and Multiwell-MEA-System (Multi Channel Systems), in three separate laboratories. Pentylenetetrazole (PTZ) and/or picrotoxin (PTX) were included in each plate as positive (n = 3-6 wells) and ≤0.2% DMSO was used as negative controls (n = 3-12 wells). In general, concentrations in a range of 0.1-30 μM were tested, anchored, when possible, on clinically relevant exposures (unbound Cmax) were tested. Activity thresholds for drug-induced changes were set at 20%. To evaluate sensitivity, specificity and predictivity of the cell models, seizurogenic responses were defined as changes in 4 or more endpoints. Concentration dependence trends were also considered.
RESULTS: Neuronal activity of 33 compounds categorized as positive tool drugs, seizure-positive or seizure-negative compounds was evaluated. Acute drug effects (<60 min) were compared to baseline recordings. Time points < 15 min exhibited stronger, less variable responses to many of the test agents. For many compounds a reduction and cessation of neuronal activity was detected at higher test concentrations. There was not a single pattern of seizurogenic activity detected, even among tool compounds, likely due to different mechanisms of actions and/or off-target profiles. A post-hoc analysis focusing on changes indicative of neuronal excitation is presented.
CONCLUSIONS: All cell models showed good sensitivity, ranging from 70 to 86%. Specificity ranged from 40 to 70%. Compared to more conventional measurements of evoked activity in hippocampal slices, these plate-based models provide higher throughput and the potential to study subacute responses. Yet, they may be limited by the random, spontaneous nature of their network activity.

Closed-loop neural implants based on continuous brain activity recording and intracortical microstimulation are extremely effective and promising devices to monitor and address many neurodegenerative diseases. The efficiency of these devices depends on the robustness of the designed circuits which rely on precise electrical equivalent models of the electrode/brain interface. This is true in the case of amplifiers for differential recording, voltage or current drivers for neurostimulation, and potentiostats for electrochemical bio-sensing. This is of paramount importance, especially for the next generation of wireless and ultra-miniaturised CMOS neural implants. Circuits are usually designed and optimized considering the electrode/brain impedance with a simple electrical equivalent model whose parameters are stationary over time. However, the electrode/brain interfacial impedance varies simultaneously in frequency and in time after implantation. The aim of this study is to monitor the impedance changes occurring on microelectrodes inserted in ex-vivo porcine brains to derive an opportune electrode/brain model describing the system and its evolution in time. In particular, impedance spectroscopy measurements have been performed for 144 hours to characterise the evolution of the electrochemical behaviour in two different setups analysing both the neural recording and the chronic stimulation scenarios. Then, different equivalent electrical circuit models have been proposed to describe the system. Results showed a decrease in the resistance to charge transfer, attributed to the interaction between biological material and the electrode surface. These findings are crucial to support circuit designers in the field of neural implants.

Deep brain stimulation (DBS) implantation under general anaesthesia (GA) has been applied to patients with Parkinson\'s disease (PD) with severe comorbidities or disabling off-medication symptoms. However, general anaesthetics may affect intraoperative microelectrode recording (MER) to varying degrees. At present, there are few studies on the effects of sedatives or general anaesthetics on multiunit activity characteristics performed by MER in patients with PD during DBS. Therefore, the effect of the choice of anaesthesia on MER remains unclear.
This is a prospective randomised controlled, non-inferiority study that will be carried out at Beijing Tiantan Hospital, Capital Medical University. Patients undergoing elective bilateral subthalamic nucleus (STN)-DBS will be enrolled after careful screening for eligibility. One hundred and eighty-eight patients will be randomised to receive either conscious sedation (CS) or GA at a 1:1 ratio. The primary outcome is the proportion of high normalised root mean square (NRMS) recorded by the MER signal.
The study was approved by the Ethics Committee of Beijing Tiantan Hospital of Capital Medical University (KY2022-147-02). Negative study results will indicate that GA using desflurane has a non-inferior effect on MER during STN-DBS compared with CS. The results of this clinical trial will be presented at national or international conferences and submitted to a peer-reviewed journal.
NCT05550714.

Both the cellular- and population-level properties of involved neurons are essential for unveiling the learning and memory functions of the brain. To give equal attention to these two aspects, neural sensors based on microelectrode arrays (MEAs) have been in the limelight due to their noninvasive detection and regulation capabilities. Here, we fabricated a neural sensor using carboxylated graphene/3,4-ethylenedioxythiophene:polystyrenesulfonate (cGO/PEDOT:PSS), which is effective in sensing and monitoring neuronal electrophysiological activity in vitro for a long time. The cGO/PEDOT:PSS-modified microelectrodes exhibited a lower electrochemical impedance (7.26 ± 0.29 kΩ), higher charge storage capacity (7.53 ± 0.34 mC/cm2), and improved charge injection (3.11 ± 0.25 mC/cm2). In addition, their performance was maintained after 2 to 4 weeks of long-term cell culture and 50,000 stimulation pulses. During neural network training, the sensors were able to induce learning function in hippocampal neurons through precise electrical stimulation and simultaneously detect changes in neural activity at multiple levels. At the cellular level, not only were three kinds of transient responses to electrical stimulation sensed, but electrical stimulation was also found to affect inhibitory neurons more than excitatory neurons. As for the population level, changes in connectivity and firing synchrony were identified. The cGO/PEDOT:PSS-based neural sensor offers an excellent tool in brain function development and neurological disease treatment.