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Frontiers in Endocrinology 2024Pancreatic islets are important in nutrient homeostasis and improved cellular models of clonal origin may very useful especially in view of relatively scarce primary...
BACKGROUND
Pancreatic islets are important in nutrient homeostasis and improved cellular models of clonal origin may very useful especially in view of relatively scarce primary material. Close 3D contact and coupling between β-cells are a hallmark of physiological function improving signal/noise ratios. Extracellular electrophysiology using micro-electrode arrays (MEA) is technically far more accessible than single cell patch clamp, enables dynamic monitoring of electrical activity in 3D organoids and recorded multicellular slow potentials (SP) provide unbiased insight in cell-cell coupling.
OBJECTIVE
We have therefore asked whether 3D spheroids enhance clonal β-cell function such as electrical activity and hormone secretion using human EndoC-βH1, EndoC-βH5 and rodent INS-1 832/13 cells.
METHODS
Spheroids were formed either by hanging drop or proprietary devices. Extracellular electrophysiology was conducted using multi-electrode arrays with appropriate signal extraction and hormone secretion measured by ELISA.
RESULTS
EndoC-βH1 spheroids exhibited increased signals in terms of SP frequency and especially amplitude as compared to monolayers and even single cell action potentials (AP) were quantifiable. Enhanced electrical signature in spheroids was accompanied by an increase in the glucose stimulated insulin secretion index. EndoC-βH5 monolayers and spheroids gave electrophysiological profiles similar to EndoC-βH1, except for a higher electrical activity at 3 mM glucose, and exhibited moreover a biphasic profile. Again, physiological concentrations of GLP-1 increased AP frequency. Spheroids also exhibited a higher secretion index. INS-1 cells did not form stable spheroids, but overexpression of connexin 36, required for cell-cell coupling, increased glucose responsiveness, dampened basal activity and consequently augmented the stimulation index.
CONCLUSION
In conclusion, spheroid formation enhances physiological function of the human clonal β-cell lines and these models may provide surrogates for primary islets in extracellular electrophysiology.
Topics: Humans; Insulin-Secreting Cells; Spheroids, Cellular; Electrophysiological Phenomena; Insulin Secretion; Glucose; Insulin; Action Potentials; Animals
PubMed: 38883608
DOI: 10.3389/fendo.2024.1402880 -
Electrochimica Acta Aug 2024Laser-induced graphene (LIG) electrodes have become popular for electrochemical sensor fabrication due to their simplicity for batch production without the use of...
Laser-induced graphene (LIG) electrodes have become popular for electrochemical sensor fabrication due to their simplicity for batch production without the use of reagents. The high surface area and favorable electrocatalytic properties also enable the design of small electrochemical devices while retaining the desired electrochemical performance. In this work, we systematically investigated the effect of LIG working electrode size, from 0.8 mm to 4.0 mm diameter, on their electrochemical properties, since it has been widely assumed that the electrochemistry of LIG electrodes is independent of size above the microelectrode size regime. The background and faradaic current from cyclic voltammetry (CV) of an outer-sphere redox probe [Ru(NH3)6] showed that smaller LIG electrodes had a higher electrode roughness factor and electroactive surface ratio than those of the larger electrodes. Moreover, CV of the surface-sensitive redox probes [Fe(CN)6] and dopamine revealed that smaller electrodes exhibited better electrocatalytic properties, with enhanced electron transfer kinetics. Scanning electron microscopy, Raman spectroscopy, and X-ray photoelectron spectroscopy showed that the physical and chemical surface structure were different at the electrode center versus the edges, so the electrochemical properties of the smaller electrodes were improved by having rougher surface and more density of the graphitic edge planes, and more oxide-containing groups, leading to better electrochemistry. The difference could be explained by the different photothermal reaction time from the laser scribing process that causes different stable carbon morphology to form on the polymer surface. Our results give a new insight on relationships between surface structure and electrochemistry of LIG electrodes and are useful for designing miniaturized electrochemical devices.
PubMed: 38881690
DOI: 10.1016/j.electacta.2024.144452 -
The European Journal of Neuroscience Jun 2024Age is a primary risk factor for Parkinson's disease (PD); however, the effects of aging on the Parkinsonian brain remain poorly understood, particularly for deep brain...
Age is a primary risk factor for Parkinson's disease (PD); however, the effects of aging on the Parkinsonian brain remain poorly understood, particularly for deep brain structures. We investigated intraoperative micro-electrode recordings from the subthalamic nucleus (STN) of PD patients aged between 42 and 76 years. Age was associated with decreased oscillatory beta power and non-oscillatory high-frequency power, independent of PD-related variables. Single unit firing and burst rates were also reduced, whereas the coefficient of variation and the structure of burst activity were unchanged. Phase synchronization (debiased weighed phase lag index [dWPLI]) between sites was pronounced in the beta band between electrodes in the superficial STN but was unaffected by age. Our results show that aging is associated with reduced neuronal activity without changes to its temporal structure. We speculate that the loss of activity in the STN may mediate the relationship between PD and age.
PubMed: 38880896
DOI: 10.1111/ejn.16435 -
NPJ Parkinson's Disease Jun 2024The subthalamic nucleus (STN) is pivotal in basal ganglia function in health and disease. Micro-electrode recordings of >25,000 recording sites from 146 Parkinson's...
The subthalamic nucleus (STN) is pivotal in basal ganglia function in health and disease. Micro-electrode recordings of >25,000 recording sites from 146 Parkinson's patients undergoing deep brain stimulation (DBS) allowed differentiation between subthalamic input, represented by local field potential (LFP), and output, reflected in spike discharge rate (SPK). As with many natural systems, STN neuronal activity exhibits power-law dynamics characterized by the exponent α. We, therefore, dissected STN data into aperiodic and periodic components using the Fitting Oscillations & One Over F (FOOOF) tool. STN LFP showed significantly higher aperiodic exponents than SPK. Additionally, SPK beta oscillations demonstrated a downward frequency shift compared to LFP. Finally, the STN aperiodic and spiking parameters explained a significant fraction of the variance of the burden and treatment efficacy of Parkinson's disease. The unique STN input-output dynamics may clarify its role in Parkinson's physiology and can be utilized in closed-loop DBS therapy.
PubMed: 38879564
DOI: 10.1038/s41531-024-00737-8 -
Analytical and Bioanalytical Chemistry Jun 2024Organic-inorganic hybrid nanocomposites (OIHN), with tailored surface chemistry, offer ultra-sensitive architecture capable of detecting ultra-low concentrations of...
Organic-inorganic hybrid nanocomposites (OIHN), with tailored surface chemistry, offer ultra-sensitive architecture capable of detecting ultra-low concentrations of target analytes with precision. In the present work, a novel nano-biosensor was fabricated, acquainting dynamic synergy of reduced graphene oxide (rGO) decorated hexagonal boron nitride nanosheets (hBNNS) for detection of carcinoembryonic antigen (CEA). Extensive spectroscopic and microscopic analyses confirmed the successful hydrothermal synthesis of cross-linked rGO-hBNNS nanocomposite. Uniform micro-electrodes of rGO-hBNNS onto pre-hydrolyzed ITO were obtained via electrophoretic deposition (EPD) technique at low DC potential (15 V). Optimization of antibody incubation time, pH of supporting electrolyte, and immunoelectrode preparation was thoroughly investigated to enhance nano-biosensing efficacy. rGO-modified hBNNS demonstrated 29% boost in electrochemical performance over bare hBNNS, signifying remarkable electro-catalytic activity of nano-biosensor. The presence of multifunctional groups on the interface facilitated stable crosslinking chemistry, increased immobilization density, and enabled site-specific anchoring of Anti-CEA, resulting in improved binding affinity. The nano-biosensor demonstrated a remarkably low limit of detection of 5.47 pg/mL (R = 0.99963), indicating exceptional sensitivity and accuracy in detecting CEA concentrations from 0 to 50 ng/mL. The clinical evaluation confirmed its exceptional shelf life, minimal cross-reactivity, and robust recovery rates in human serum samples, thereby unraveling the potential for early, highly sensitive, and reliable CEA detection.
PubMed: 38878180
DOI: 10.1007/s00216-024-05379-6 -
The Journal of Physiology Jun 2024Large trans-sarcolemmal ionic shifts occur with fatiguing exercise or stimulation of isolated muscles. However, it is unknown how resting membrane potential (E) and...
Large trans-sarcolemmal ionic shifts occur with fatiguing exercise or stimulation of isolated muscles. However, it is unknown how resting membrane potential (E) and intracellular sodium concentration ([Na]) change with repeated contractions in living mammals. We investigated (i) whether [Na] (peak, kinetics) can reveal changes of Na-K pump activity during brief or fatiguing stimulation and (ii) how resting E and [Na] change during fatigue and recovery of rat soleus muscle in situ. Muscles of anaesthetised rats were stimulated with brief (10 s) or repeated tetani (60 Hz for 200 ms, every 2 s, for 30 s or 300 s) with isometric force measured. Double-barrelled ion-sensitive microelectrodes were used to quantify resting E and [Na]. Post-stimulation data were fitted using polynomials and back-extrapolated to time zero recovery. Mean pre-stimulation resting E (layer 2-7 fibres) was -71 mV (surface fibres were more depolarised), and [Na] was 14 mM. With deeper fibres, 10 s stimulation (2-150 Hz) increased [Na] to 38-46 mM whilst simultaneously causing hyperpolarisations (7.3 mV for 2-90 Hz). Fatiguing stimulation for 30 s or 300 s led to end-stimulation resting E of -61 to -53 mV, which recovered rapidly (T, 8-22 s). Mean end-stimulation [Na] increased to 86-101 mM with both fatigue protocols and the [Na] recovery time-course (T, 21-35 s) showed no difference between protocols. These combined findings suggest that brief stimulation hyperpolarises the resting E, likely via maximum Na-induced stimulation of the Na-K pump. Repeated tetani caused massive depolarisation and elevations of [Na] that together lower force, although they likely interact with other factors to cause fatigue. [Na] recovery kinetics provided no evidence of impaired Na-K pump activity with fatigue. KEY POINTS: It is uncertain how resting membrane potential, intracellular sodium concentration ([Na]), and sodium-potassium (Na-K) pump activity change during repeated muscle contractions in living mammals. For rat soleus muscle fibres in situ, brief tetanic stimulation for 10 s led to raised [Na], anticipated to evoke maximal Na-induced stimulation of the Na-K pump causing an immediate hyperpolarisation of the sarcolemma. More prolonged stimulation with repeated tetanic contractions causes massive elevations of [Na], which together with large depolarisations (via K disturbances) likely reduce force production. These effects occurred without impairment of Na-K pump function. Together these findings suggest that rapid activation of the Na-K pump occurs with brief stimulation to maintain excitability, whereas more prolonged stimulation causes rundown of the trans-sarcolemmal K gradient (hence depolarisation) and Na gradient, which in combination can impair contraction to contribute to fatigue in living mammals.
PubMed: 38877870
DOI: 10.1113/JP285870 -
Scientific Reports Jun 2024Cortico-cortical evoked potentials (CCEPs) elicited by single-pulse electric stimulation (SPES) are widely used to assess effective connectivity between cortical areas...
Cortico-cortical evoked potentials (CCEPs) elicited by single-pulse electric stimulation (SPES) are widely used to assess effective connectivity between cortical areas and are also implemented in the presurgical evaluation of epileptic patients. Nevertheless, the cortical generators underlying the various components of CCEPs in humans have not yet been elucidated. Our aim was to describe the laminar pattern arising under SPES evoked CCEP components (P1, N1, P2, N2, P3) and to evaluate the similarities between N2 and the downstate of sleep slow waves. We used intra-cortical laminar microelectrodes (LMEs) to record CCEPs evoked by 10 mA bipolar 0.5 Hz electric pulses in seven patients with medically intractable epilepsy implanted with subdural grids. Based on the laminar profile of CCEPs, the latency of components is not layer-dependent, however their rate of appearance varies across cortical depth and stimulation distance, while the seizure onset zone does not seem to affect the emergence of components. Early neural excitation primarily engages middle and deep layers, propagating to the superficial layers, followed by mainly superficial inhibition, concluding in a sleep slow wave-like inhibition and excitation sequence.
Topics: Humans; Male; Female; Adult; Electric Stimulation; Evoked Potentials; Cerebral Cortex; Drug Resistant Epilepsy; Electroencephalography; Young Adult; Middle Aged; Epilepsy
PubMed: 38877093
DOI: 10.1038/s41598-024-62433-0 -
Biosensors & Bioelectronics Oct 2024Electroplating of poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) is important in many neuroelectronic applications but is challenging to achieve...
Electroplating of poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) is important in many neuroelectronic applications but is challenging to achieve uniformity on large-scale microelectrode arrays (MEA) using conventional galvanostatic methods. In this study, we address this challenge through a potentiostatic method and demonstrate highly uniform electroplating of PEDOT:PSS on MEA with more than one hundred electrodes, all at cellular sizes. The validation of this approach involves comparisons with galvanostatic deposition methods, showcasing unparalleled deposition yield and uniformity. Systematic electrochemical characterizations reveal similarities in structure and stability from potentiostatic deposited coatings. The advances developed here establish the potentiostatic method and detailed process to achieve a uniform coating of PEDOT:PSS on large-scale MEA, with broad utility in neuroelectronics.
Topics: Polystyrenes; Microelectrodes; Electroplating; Biosensing Techniques; Bridged Bicyclo Compounds, Heterocyclic; Polymers; Animals; Electrochemical Techniques; Thiophenes
PubMed: 38875864
DOI: 10.1016/j.bios.2024.116418 -
Frontiers in Pharmacology 2024We investigated drug-induced acute neuronal electrophysiological changes using Micro-Electrode arrays (MEA) to rat primary neuronal cell cultures. Data based on 6-key...
We investigated drug-induced acute neuronal electrophysiological changes using Micro-Electrode arrays (MEA) to rat primary neuronal cell cultures. Data based on 6-key MEA parameters were analyzed for plate-to-plate vehicle variability, effects of positive and negative controls, as well as data from over 100 reference drugs, mostly known to have pharmacological phenotypic and clinical outcomes. A Least Absolute Shrinkage and Selection Operator (LASSO) regression, coupled with expert evaluation helped to identify the 6-key parameters from many other MEA parameters to evaluate the drug-induced acute neuronal changes. Calculating the statistical tolerance intervals for negative-positive control effects on those 4-key parameters helped us to develop a new weighted hazard scoring system on drug-induced potential central nervous system (CNS) adverse effects (AEs). The weighted total score, integrating the effects of a drug candidate on the identified six-pivotal parameters, simply determines if the testing compound/concentration induces potential CNS AEs. Hereto, it uses four different categories of hazard scores: non-neuroactive, neuroactive, hazard, or high hazard categories. This new scoring system was successfully applied to differentiate the new compounds with or without CNS AEs, and the results were correlated with the outcome of studies in mice for one internal program. Furthermore, the Random Forest classification method was used to obtain the probability that the effect of a compound is either inhibitory or excitatory. In conclusion, this new neuronal scoring system on the cell assay is actively applied in the early de-risking of drug development and reduces the use of animals and associated costs.
PubMed: 38873414
DOI: 10.3389/fphar.2024.1308547 -
Biosensors & Bioelectronics Oct 2024Multichannel arrays capable of real-time sensing of neuromodulators in the brain are crucial for gaining insights into new aspects of neural communication. However,...
Multichannel arrays capable of real-time sensing of neuromodulators in the brain are crucial for gaining insights into new aspects of neural communication. However, measuring neurochemicals, such as dopamine, at low concentrations over large areas has proven challenging. In this research, we demonstrate a novel approach that leverages the scalability and processing power offered by microelectrode array devices integrated with a functionalized, high-density microwire bundle, enabling electrochemical sensing at an unprecedented scale and spatial resolution. The sensors demonstrate outstanding selective molecular recognition by incorporating a selective polymeric membrane. By combining cutting-edge commercial multiplexing, digitization, and data acquisition hardware with a bio-compatible and highly sensitive neurochemical interface array, we establish a powerful platform for neurochemical analysis. This multichannel array has been successfully utilized in vitro and ex vivo systems. Notably, our results show a sensing area of 2.25 mm with an impressive detection limit of 820 pM for dopamine. This new approach paves the way for investigating complex neurochemical processes and holds promise for advancing our understanding of brain function and neurological disorders.
Topics: Biosensing Techniques; Dopamine; Animals; Electrochemical Techniques; Microelectrodes; Limit of Detection; Equipment Design; Brain; Humans; Neurotransmitter Agents
PubMed: 38870827
DOI: 10.1016/j.bios.2024.116474