Reconstruction of Functional Connectivity from Multielectrode Recordings and Calcium Imaging.

In the last two decades, increasing research efforts in neuroscience have been focused on determining both structural and functional connectivity of brain circuits, with the main goal of relating the wiring diagram of neuronal systems to their emerging properties, from the microscale to the macroscale. While combining multisite parallel recordings with structural circuits' reconstruction in vivo is still very challenging, the reductionist in vitro approach based on neuronal cultures offers lower technical difficulties and is much more stable under control conditions. In this chapter, we present different approaches to infer the connectivity of cultured neuronal networks using multielectrode array or calcium imaging recordings. We first formally introduce the used methods, and then we will describe into details how those methods were applied in case studies. Since multielectrode array and calcium imaging recordings provide distinct and complementary spatiotemporal features of neuronal activity, in this chapter we present the strategies implemented with the two different methodologies in distinct sections.

A novel simplistic fabrication technique for cranial epidural electrodes for chronic recording and stimulation in rats.

BACKGROUND: The demand for neuromodulatory and recording tools has resulted in a surge of publications describing techniques for fabricating devices and accessories in-house suitable for neurological recordings. However, many of these fabrication protocols use equipment which are not common to biological laboratories, thus limiting researchers to the use of commercial alternatives. New method:We have developed a simple yet robust implantable stimulating surface electrode which can be fabricated in all wet-bench laboratories. RESULTS: Female Sprague-Dawley rats received epidural implantation of the electrodes over the fore and hind limb areas of their motor cortex. Stimulation of the motor cortex successfully evoked fore- and hind limb motor outputs. The device was also able to record surface potentials of the motor cortex following epidural stimulation of the spinal cord. Comparisons with existing methods:For stimulation of the motor cortex, often stiff stainless or copper wires are roughly tucked underneath the skull, with little accuracy of localization. While, commercially available devices utilize burr holes and screw electrodes. Our new electrode design provides us stereotaxic accuracy that was not previously available. CONCLUSION: We developed a chronic implantable electrode capable of being fabricated in all wet-labs, are robust, versatile and electrically sensitive enough for long-term chronic use. The simple and versatile electrode design provides scientific, economical and ethical benefits.

Noninvasive Magnetogastrography Detects Erythromycin-Induced Effects on the Gastric Slow Wave.

OBJECTIVE: The prokinetic action of erythromycin is clinically useful under conditions associated with gastrointestinal hypomotility. Although erythromycin is known to affect the electrogastrogram, no studies have examined the effects that erythromycin has on gastric slow wave magnetic fields. METHODS: In this study, gastric slow wave activity was assessed simultaneously using noninvasive magnetogastrogram (MGG), electrogastrogram, and mucosal electromyogram recordings. Recordings were obtained for 30 min prior to and 60 min after intravenous administration of erythromycin at dosages of 3 and 6 mg/kg. RESULTS: MGG recordings showed significant changes in the percentage power distribution of gastric signal after infusion of both 3 and 6 mg/kg erythromycin at t = 1-5 min that persisted for t = 30-40 min after infusion. These changes agree with the changes observed in the electromyogram. We did not observe any statistically significant difference in MGG amplitude before or after injection of either 3 or 6 mg/kg erythromycin. Both 3 and 6 mg/kg erythromycin infusion showed retrograde propagation with a statistically significant decrease in slow wave propagation velocity 11-20 min after infusion. Propagation velocity started returning toward baseline values after approximately 21-30 min for the 3 mg/kg dosage and after 31-40 min for a dosage of 6 mg/kg. CONCLUSION: Our results showed that the magnetic signatures were sensitive to disruptions in normal slow wave activity induced by pharmacological and prokinetic agents such as erythromycin. SIGNIFICANCE: This study shows that repeatable noninvasive bio-electro-magnetic techniques can objectively characterize gastric dysrhythmias and may quantify treatment efficacy in patients with functional gastric disorders.

Cracking the Function of Layers in the Sensory Cortex.

Understanding how cortical activity generates sensory perceptions requires a detailed dissection of the function of cortical layers. Despite our relatively extensive knowledge of their anatomy and wiring, we have a limited grasp of what each layer contributes to cortical computation. We need to develop a theory of cortical function that is rooted solidly in each layer's component cell types and fine circuit architecture and produces predictions that can be validated by specific perturbations. Here we briefly review the progress toward such a theory and suggest an experimental road map toward this goal. We discuss new methods for the all-optical interrogation of cortical layers, for correlating in vivo function with precise identification of transcriptional cell type, and for mapping local and long-range activity in vivo with synaptic resolution. The new technologies that can crack the function of cortical layers are finally on the immediate horizon.

A very large-scale microelectrode array for cellular-resolution electrophysiology.

In traditional electrophysiology, spatially inefficient electronics and the need for tissue-to-electrode proximity defy non-invasive interfaces at scales of more than a thousand low noise, simultaneously recording channels. Using compressed sensing concepts and silicon complementary metal-oxide-semiconductors (CMOS), we demonstrate a platform with 65,536 simultaneously recording and stimulating electrodes in which the per-electrode electronics consume an area of 25.5 µm by 25.5 µm. Application of this platform to mouse retinal studies is achieved with a high-performance processing pipeline with a 1 GB/s data rate. The platform records from 65,536 electrodes concurrently with a ~10 µV r.m.s. noise; senses spikes from more than 34,000 electrodes when recording across the entire retina; automatically sorts and classifies greater than 1700 neurons following visual stimulation; and stimulates individual neurons using any number of the 65,536 electrodes while observing spikes over the entire retina. The approaches developed here are applicable to other electrophysiological systems and electrode configurations.

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats.

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the pathophysiological basis of these diseases, it is necessary to precisely characterize in which way the processing of information is disturbed between the different neuronal parts of the circuit. Using extracellular in vivo electrophysiological recordings, it is possible to accurately delineate neuronal activity within a neuronal network. The application of this method has several advantages over alternative techniques, e.g., functional magnetic resonance imaging and calcium imaging, as it allows a unique temporal and spatial resolution and does not rely on genetically engineered organisms. However, the use of extracellular in vivo recordings is limited since it is an invasive technique that cannot be universally applied. In this article, a simple and easy to use method is presented with which it is possible to simultaneously record extracellular potentials such as local field potentials and multiunit activity at multiple sites of a network. It is detailed how a precise targeting of subcortical nuclei can be achieved using a combination of stereotactic surgery and online analysis of multi-unit recordings. Thus, it is demonstrated, how a complete network such as the hyperdirect cortico-basal ganglia loop can be studied in anesthetized animals in vivo.

Effect of synthetic cannabinoids on spontaneous neuronal activity: Evaluation using Ca(2+) spiking and multi-electrode arrays.

Activation of cannabinoid receptor 1 (CB1) inhibits synaptic transmission in hippocampal neurons. The goal of this study was to evaluate the ability of benchmark and emerging synthetic cannabinoids to suppress neuronal activity in vitro using two complementary techniques, Ca(2+) spiking and multi-electrode arrays (MEAs). Neuron culture and fluorescence imaging conditions were extensively optimized to provide maximum sensitivity for detection of suppression of neural activity by cannabinoids. The neuronal Ca(2+) spiking frequency was significantly suppressed within 10min by the prototypic aminoalkylindole cannabinoid, WIN 55,212-2 (10µM). Suppression by WIN 55,212-2 was not improved by pharmacological intervention with signaling pathways known to interfere with CB1 signaling. The naphthoylindole CB1 agonist, JWH-018 suppressed Ca(2+) spiking at a lower concentration (2.5µM), and the CB1 antagonist rimonabant (5µM), reversed this suppression. In the MEA assay, the ability of synthetic CB1 agonists to suppress spontaneous electrical activity of hippocampal neurons was evaluated over 80min sessions. All benchmark (WIN 55,212-2, HU-210, CP 55,940 and JWH-018) and emerging synthetic cannabinoids (XLR-11, JWH-250, 5F-PB-22, AB-PINACA and MAM-2201) suppressed neural activity at a concentration of 10µM; furthermore, several of these compounds also significantly suppressed activity at 1µM concentrations. Rimonabant partially reversed spiking suppression of 5F-PB-22 and, to a lesser extent, of MAM-2201, supporting CB1-mediated involvement, although the inactive WIN 55,212-3 also partially suppressed activity. Taken together, synthetic cannabinoid CB1-mediated suppression of neuronal activity was detected using Ca(2+) spiking and MEAs.

Optogenetic activation of neocortical neurons in vivo with a sapphire-based micro-scale LED probe.

Optogenetics has proven to be a revolutionary technology in neuroscience and has advanced continuously over the past decade. However, optical stimulation technologies for in vivo need to be developed to match the advances in genetics and biochemistry that have driven this field. In particular, conventional approaches for in vivo optical illumination have a limitation on the achievable spatio-temporal resolution. Here we utilize a sapphire-based microscale gallium nitride light-emitting diode (µLED) probe to activate neocortical neurons in vivo. The probes were designed to contain independently controllable multiple µLEDs, emitting at 450 nm wavelength with an irradiance of up to 2 W/mm(2). Monte-Carlo stimulations predicted that optical stimulation using a µLED can modulate neural activity within a localized region. To validate this prediction, we tested this probe in the mouse neocortex that expressed channelrhodopsin-2 (ChR2) and compared the results with optical stimulation through a fiber at the cortical surface. We confirmed that both approaches reliably induced action potentials in cortical neurons and that the µLED probe evoked strong responses in deep neurons. Due to the possibility to integrate many optical stimulation sites onto a single shank, the µLED probe is thus a promising approach to control neurons locally in vivo.

Beat synchronization predicts neural speech encoding and reading readiness in preschoolers.

Temporal cues are important for discerning word boundaries and syllable segments in speech; their perception facilitates language acquisition and development. Beat synchronization and neural encoding of speech reflect precision in processing temporal cues and have been linked to reading skills. In poor readers, diminished neural precision may contribute to rhythmic and phonological deficits. Here we establish links between beat synchronization and speech processing in children who have not yet begun to read: preschoolers who can entrain to an external beat have more faithful neural encoding of temporal modulations in speech and score higher on tests of early language skills. In summary, we propose precise neural encoding of temporal modulations as a key mechanism underlying reading acquisition. Because beat synchronization abilities emerge at an early age, these findings may inform strategies for early detection of and intervention for language-based learning disabilities.

Localising and classifying neurons from high density MEA recordings.

Neuronal microcircuits are formed of a myriad of spatially and functionally specific cell classes. Despite the importance of the spatial component in the characterisation of neural circuits, it has not received the attention it deserves. While multi-electrodes are widely used in the study of microcircuits, the spatial information available from them remains largely unexploited for analysis beyond spike sorting. Here we show how the spatial pattern of the extracellular signal is determined by both the electrophysiology and morphology of neurons. Starting from known current source models for the generation of the extracellular potential, we use the spatial pattern observed across a multi-electrode array to localise and classify neurons into putative morphological classes. We evaluated the localisation and classification models with low fitting errors in simulated data. When applying them to recorded data we found correspondence between localisation statistics and expected recording radius and found evidence to support the separation into putative morphological classes. While existing localisation methods do not hold for the recording distances expected on multi-electrode recordings (under 60µm), classification methods have been limited to the temporal component by either characterising spike shape or firing patterns. We show here how the information available from extracellular recordings can be used to localise and classify neurons based on the spatial pattern seen by multi-electrode arrays. Together they can improve current characterisation and classification of neurons based on complementary criteria such us firing pattern and functional characterisation.

The effect of site placement within silicon microelectrodes on the long-term electrophysiological recordings.

Intracortical microelectrodes can be used to treat various neurological disorders given their capabilities to interface with single or multiple populations of neurons. However, most of these penetrating devices have been reported to fail over time, within weeks to months, putatively due to the foreign body response (FBR) which persistently aggravates the surrounding brain tissues. A number of studies have confirmed that various electrode properties, such as size, shape, and surface area, may play a role in the biological responses to the microelectrode. Further experimental data is needed to determine the effect of these properties on the FBR and the recording performance. In this paper, we evaluate the effect of site placement using Michigan arrays with sites on the center, edge, and tip of the shank. The results show that there is significant performance variance between the center, edge, and tip sites.

Simultaneous stimulation and recording of cardiac depolarization enabled by high-frequency stimulation.

High-frequency stimulation techniques have been recently proposed for the pacing and control of excitability of cardiac tissues. This paper introduces a system designed specifically for such stimulation, and demonstrates the unique ability to record depolarization events on the same electrode used for stimulation, during the stimulus. Experimental results with HL-1 cardiomyocytes are presented, highlighting key concepts enabled by this system, such as direct strength-duration relationship measurement and beat-to-beat stimulation threshold monitoring following pacing onset or pharmacological modulation.

Diagnóstico diferencial entre doble respuesta ventricular y extrasistolia hisiana bigeminada. Respuesta / Differential diagnosis between dual ventricular response and bigeminy arising from the bundle of his. Response

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Recording evoked potentials during deep brain stimulation: development and validation of instrumentation to suppress the stimulus artefact.

The clinical efficacy of deep brain stimulation (DBS) for the treatment of movement disorders depends on the identification of appropriate stimulation parameters. Since the mechanisms of action of DBS remain unclear, programming sessions can be time consuming, costly and result in sub-optimal outcomes. Measurement of electrically evoked compound action potentials (ECAPs) during DBS, generated by activated neurons in the vicinity of the stimulating electrode, could offer insight into the type and spatial extent of neural element activation and provide a potential feedback signal for the rational selection of stimulation parameters and closed-loop DBS. However, recording ECAPs presents a significant technical challenge due to the large stimulus artefact, which can saturate recording amplifiers and distort short latency ECAP signals. We developed DBS-ECAP recording instrumentation combining commercial amplifiers and circuit elements in a serial configuration to reduce the stimulus artefact and enable high fidelity recording. We used an electrical circuit equivalent model of the instrumentation to understand better the sources of the stimulus artefact and the mechanisms of artefact reduction by the circuit elements. In vitro testing validated the capability of the instrumentation to suppress the stimulus artefact and increase gain by a factor of 1000 to 5000 compared to a conventional biopotential amplifier. The distortion of mock ECAP (mECAP) signals was measured across stimulation parameters, and the instrumentation enabled high fidelity recording of mECAPs with latencies of only 0.5 ms for DBS pulse widths of 50 to 100 µs/phase. Subsequently, the instrumentation was used to record in vivo ECAPs, without contamination by the stimulus artefact, during thalamic DBS in an anesthetized cat. The characteristics of the physiological ECAP were dependent on stimulation parameters. The novel instrumentation enables high fidelity ECAP recording and advances the potential use of the ECAP as a feedback signal for the tuning of DBS parameters.

[Study on signal transmission characteristics of meridian based on electrical network theory and experiments].

Study on features of acupoints with resistance test in the past half century is reviewed in this article. Mechanism and technology of the method are introduced as well as its shortcomings. The determination method of signal transmission along meridians with the combination of electrical network theories and practice is advanced. And the result of a series experiments on one meridian at the superficial part of the body are given as well. Thus, it is concluded that the signals of the point-in/point-out and the signals along a non-meridian path with the same distance are significantly different, which gives a verification of the feasibility of the method by using electrical network theories to set out characteristics of signal transmission along meridians dynamically.

A bundled microwire array for long-term chronic single-unit recording in deep brain regions of behaving rats.

Chronic single-unit recording in subcortical brain regions is increasingly important in neurophysiological studies. However, methods for long-term, stable recording of multiple single-units in deep brain regions and in dura-surrounded ganglion have not yet been established. In the present study, we propose a bundled microwire array design which is capable of long-term recording of the trigeminal ganglion and deep-brain units. This electrode set is easy to construct from common materials and tools found in an electrophysiological laboratory. The salient features of our design include: (1) short and separated tungsten microwires for stable chronic recording; (2) the use of a 30-guage stainless steel guide tube for facilitating penetration and aiming for deep targets as well as electrical grounding; (3) the inclusion of a reference of the same microwire material inside the bundle to enhance common mode rejection of far field noises; and (4) an adjustable connector. In our case, we used a 90° backward bending connector so that implanted rats could perform the same hole-seeking behavior and their faces and the whiskers could be stimulated in the behaving state. It was demonstrated that this multi-channel electrode caused minimal tissue damage at the recording site and we were able to obtain good, stable single-unit recordings from the trigeminal ganglion and ventroposterior medial thalamus areas of freely moving rats for up to 80 days. This methodology is useful for the studies that require long term and high quality unit recording in the deep brain or in the trigeminal system.

Reliability of AcuGraph system for measuring skin conductance at acupoints.

OBJECTIVE: There are many commercially available instruments for measuring electrical conductance, but there is little information about their reliability. The aim of this study was to quantify measurement variability and assess reliability of the AcuGraph system-a commonly used electrodermal screening device. METHODS: Four experiments were conducted to measure variability in electrical conductance readings obtained by the AcuGraph system. The first involved measuring known resistors. The second measured non-human organic matter. The third was a test-retest assessment of the Yuan-Source and Jing-Well points in 30 healthy volunteers who were measured by a single operator. The fourth was an interoperator reliability evaluation of seven acupuncturists at the Yuan-Source and Jing-Well acupoints on four individuals at two time points. RESULTS: Against known resistors, the AcuGraph had an average coefficient of variability (CV) of 1.8% between operators and test-retests. On non-human organic material the AcuGraph had an average CV of 0.9% and 2.8%. When a single operator tested 30 participants, the average reliability for the Yuan-Source points was 0.86 and 0.76 for Jing-Well points with a CV of 23.2% and 25.9% respectively. The average CV for the seven acupuncturists was 24.5% on Yuan-Source points and 23.7% on Jing-Well points. CONCLUSIONS: The AcuGraph measures known resistors and organic matter accurately and reliably. Skin conductance at acupoints recorded by one operator was also reliable. There was less consistency in electrodermal recordings obtained by seven different operators. Operator training and technical improvements to the AcuGraph may improve consistency among operators.

Feedback controlled piezo-motor microdrive for accurate electrode positioning in chronic single unit recording in behaving mice.

The microdrive is one of the most essential tools for extracellular, single-unit recordings in freely behaving animals to detect and isolate the single-unit activities from brain regions of interest. Due to the increasing number of neuroscience research projects using genetically engineered mice, the demand for effective recording devices in freely moving mice is also increasing. Although manually and automatically operated microdrive devices are available, they are limited in terms of size, weight, accuracy, manipulability, and convenience for single-unit recording in mice. The present study proposed a novel microdrive that employs a small, lightweight piezo-motor and a magnetoresistive (MR) sensor with a closed-loop position feedback control system. The total weight of the device is 1.82 g, which is perfectly suitable for application to mice. Most importantly, the proposed microdrive is capable of monitoring and adjusting electrode movement on-line by integrating a closed-loop feedback control system, which enhances the accuracy of micro-advancement of the electrode by utilizing position feedback. The performance of this newly developed microdrive was extensively evaluated for both mechanical and physiological concerns at both free-loading and various-loading conditions, including agarose gel matrix and then the hippocampus and thalamus of mice. In summary, this proposed microdrive can enhance the quality of recording single unit activities in freely moving mice in terms of the size and weight of the device, the convenience and accuracy of manipulation, and, most of all, in isolating single neurons and recording stability by providing accurate positioning of an electrode.

Microelectrode arrays: a physiologically based neurotoxicity testing platform for the 21st century.

Microelectrode arrays (MEAs) have been in use over the past decade and a half to study multiple aspects of electrically excitable cells. In particular, MEAs have been applied to explore the pharmacological and toxicological effects of numerous compounds on spontaneous activity of neuronal and cardiac cell networks. The MEA system enables simultaneous extracellular recordings from multiple sites in the network in real time, increasing spatial resolution and thereby providing a robust measure of network activity. The simultaneous gathering of action potential and field potential data over long periods of time allows the monitoring of network functions that arise from the interaction of all cellular mechanisms responsible for spatio-temporal pattern generation. In these functional, dynamic systems, physical, chemical, and pharmacological perturbations are holistically reflected by the tissue responses. Such features make MEA technology well suited for the screening of compounds of interest, and also allow scaling to high throughput systems that can record from multiple, separate cell networks simultaneously in multi-well chips or plates. This article is designed to be useful to newcomers to this technology as well as those who are currently using MEAs in their research. It explains how MEA systems operate, summarizes what systems are available, and provides a discussion of emerging mathematical schemes that can be used for a rapid classification of drug or chemical effects. Current efforts that will expand this technology to an influential, high throughput, electrophysiological approach for reliable determinations of compound toxicity are also described and a comprehensive review of toxicological publications using MEAs is provided as an appendix to this publication. Overall, this article highlights the benefits and promise of MEA technology as a high throughput, rapid screening method for toxicity testing.

Electrophysiological and neurochemical techniques to investigate sensory neurons in analgesia research.

The primary afferent nociceptive neuron has recently attracted major research interest because of the cloning of very selectively expressed and well-conserved ion channel genes. All parts of the neuron, sensory terminals, axon and cell body, are accessible to validated research techniques in vitro using various isolated tissues or cells taken from laboratory animals. Single-unit recording and measuring stimulated calcitonin gene-related peptide (CGRP) release as well as patch-clamping and calcium imaging of cultured sensory neurons provide different kinds of information, and no model alone answers all questions. In combination, however, consistent results and complementary evidence form a solid basis for translational research to follow.