scholarly journals A simple Ca2+-imaging approach to neural network analysis in cultured neurons

2020 ◽  
Author(s):  
Zijun Sun ◽  
Thomas C. Südhof
Keyword(s):  
Single Neuron ◽  
Rapid Assessment ◽  
Scale Up ◽  
Network Activity ◽  
Cultured Neurons ◽  
Electrode Arrays ◽  
Network Analyses ◽  
Conditional Deletion ◽  
Human Neurons ◽  
Human And Mouse

AbstractBackgroundCa2+-imaging is a powerful tool to measure neuronal dynamics and network activity. To monitor network-level changes in cultured neurons, neuronal activity is often evoked by electrical or optogenetic stimulation and assessed using multi-electrode arrays or sophisticated imaging. Although such approaches allow detailed network analyses, multi-electrode arrays lack single-cell precision, whereas optical physiology generally requires advanced instrumentation.New MethodHere we developed a simple, stimulation-free protocol with associated Matlab algorithms that enables scalable analyses of network activity in cultured human and mouse neurons. The approach allows analysis of overall networks and single-neuron dynamics, and is amenable to scale-up for screening purposes.ResultsWe validated the protocol by assessing human neurons with a heterozygous conditional deletion of Munc18-1, and mouse neurons with a homozygous conditional deletion of neurexins. The approach described here enabled identification of differential changes in these mutant neurons at the network level and of the amplitude and frequency of calcium peaks at the single-neuron level. These results demonstrate the utility of the approach.Comparison with existing methodCompared with current imaging platforms, our method is simple, scalable, and easy to implement. It enables quantification of more detailed parameters than multi-electrode arrays, but does not have the resolution and depth of more sophisticated yet labour-intensive analysis methods, such as electrophysiology.ConclusionThis method is scalable for a rapid assessment of neuronal function in culture, and can be applied to both human and mouse neurons. Thus, the method can serve as a basis for phenotypical analysis of mutations and for drug discovery efforts.

scholarly journals Resolving rates of mutation in the brain using single-neuron genomics

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Gilad D Evrony ◽  
Eunjung Lee ◽  
Peter J Park ◽  
Christopher A Walsh
Keyword(s):  
Single Cell ◽  
Somatic Mutation ◽  
Brain Function ◽  
Single Neuron ◽  
Bioinformatic Analysis ◽  
Normal Brain ◽  
Neuronal Function ◽  
Human Neurons ◽  
Normal Brain Function ◽  
The Brain

Whether somatic mutations contribute functional diversity to brain cells is a long-standing question. Single-neuron genomics enables direct measurement of somatic mutation rates in human brain and promises to answer this question. A recent study (<xref ref-type="bibr" rid="bib65">Upton et al., 2015</xref>) reported high rates of somatic LINE-1 element (L1) retrotransposition in the hippocampus and cerebral cortex that would have major implications for normal brain function, and suggested that these events preferentially impact genes important for neuronal function. We identify aspects of the single-cell sequencing approach, bioinformatic analysis, and validation methods that led to thousands of artifacts being interpreted as somatic mutation events. Our reanalysis supports a mutation frequency of approximately 0.2 events per cell, which is about fifty-fold lower than reported, confirming that L1 elements mobilize in some human neurons but indicating that L1 mosaicism is not ubiquitous. Through consideration of the challenges identified, we provide a foundation and framework for designing single-cell genomics studies.

scholarly journals Coherent and intermittent ensemble oscillations emerge from networks of irregular spiking neurons

2016 ◽  
Vol 115 (1) ◽  
pp. 457-469 ◽  
Author(s):  
Mahmood S. Hoseini ◽  
Ralf Wessel
Keyword(s):  
Single Neuron ◽  
Experimental Studies ◽  
Field Potential ◽  
Gamma Oscillations ◽  
Peak Frequency ◽  
Spiking Neurons ◽  
Network Activity ◽  
Cortical Circuits ◽  
Model Network ◽  
Low Rate

Local field potential (LFP) recordings from spatially distant cortical circuits reveal episodes of coherent gamma oscillations that are intermittent, and of variable peak frequency and duration. Concurrently, single neuron spiking remains largely irregular and of low rate. The underlying potential mechanisms of this emergent network activity have long been debated. Here we reproduce such intermittent ensemble oscillations in a model network, consisting of excitatory and inhibitory model neurons with the characteristics of regular-spiking (RS) pyramidal neurons, and fast-spiking (FS) and low-threshold spiking (LTS) interneurons. We find that fluctuations in the external inputs trigger reciprocally connected and irregularly spiking RS and FS neurons in episodes of ensemble oscillations, which are terminated by the recruitment of the LTS population with concurrent accumulation of inhibitory conductance in both RS and FS neurons. The model qualitatively reproduces experimentally observed phase drift, oscillation episode duration distributions, variation in the peak frequency, and the concurrent irregular single-neuron spiking at low rate. Furthermore, consistent with previous experimental studies using optogenetic manipulation, periodic activation of FS, but not RS, model neurons causes enhancement of gamma oscillations. In addition, increasing the coupling between two model networks from low to high reveals a transition from independent intermittent oscillations to coherent intermittent oscillations. In conclusion, the model network suggests biologically plausible mechanisms for the generation of episodes of coherent intermittent ensemble oscillations with irregular spiking neurons in cortical circuits.

scholarly journals L-type voltage-gated calcium channel regulation of in vitro human cortical neuronal networks

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
William Plumbly ◽  
Nick Brandon ◽  
Tarek Z. Deeb ◽  
Jeremy Hall ◽  
Adrian J. Harwood
Keyword(s):  
Calcium Channel ◽  
Neuronal Networks ◽  
Gene Mutations ◽  
Synaptic Function ◽  
Neuronal Firing ◽  
Network Activity ◽  
Electrode Arrays ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channel

Abstract The combination of in vitro multi-electrode arrays (MEAs) and the neuronal differentiation of stem cells offers the capability to study human neuronal networks from patient or engineered human cell lines. Here, we use MEA-based assays to probe synaptic function and network interactions of hiPSC-derived neurons. Neuronal network behaviour first emerges at approximately 30 days of culture and is driven by glutamate neurotransmission. Over a further 30 days, inhibitory GABAergic signalling shapes network behaviour into a synchronous regular pattern of burst firing activity and low activity periods. Gene mutations in L-type voltage gated calcium channel subunit genes are strongly implicated as genetic risk factors for the development of schizophrenia and bipolar disorder. We find that, although basal neuronal firing rate is unaffected, there is a dose-dependent effect of L-type voltage gated calcium channel inhibitors on synchronous firing patterns of our hiPSC-derived neural networks. This demonstrates that MEA assays have sufficient sensitivity to detect changes in patterns of neuronal interaction that may arise from hypo-function of psychiatric risk genes. Our study highlights the utility of in vitro MEA based platforms for the study of hiPSC neural network activity and their potential use in novel compound screening.

scholarly journals GABA Withdrawal Modifies Network Activity in Cultured Hippocampal Neurons

2000 ◽  
Vol 7 (1-2) ◽  
pp. 31-42 ◽  
Author(s):  
H. Golan ◽  
K. Mikenberg ◽  
V. Greenberger ◽  
M. Segal
Keyword(s):  
Withdrawal Syndrome ◽  
Hippocampal Neurons ◽  
High Rate ◽  
Network Activity ◽  
Cultured Neurons ◽  
Synaptic Currents ◽  
Firing Rates ◽  
Cultured Hippocampal Neurons

Dissociated hippocampal neurons, grown in culture for 2 to 3 weeks, tended to fire bursts of synaptic currents at fairly regular intervals, representing network activity. A brief exposure of cultured neurons to GABA caused a total suppression of the spontaneous network activity. Following a washout of GABA, the activity was no longer clustered in bursts and instead, the cells fired at a high rate tonic manner. The effect of removing GABA could be seen as long as 1 to 2 days after GABA withdrawal and is expressed as an increase in the number of active cells in a network, as well as in their firing rates. Such striking effects of GABA removal may underlie part of the GABA withdrawal syndrome seen elsewhere.

scholarly journals Chronic recording and electrochemical performance of Utah microelectrode arrays implanted in rat motor cortex

2018 ◽  
Vol 120 (4) ◽  
pp. 2083-2090 ◽  
Author(s):  
Bryan J. Black ◽  
Aswini Kanneganti ◽  
Alexandra Joshi-Imre ◽  
Rashed Rihani ◽  
Bitan Chakraborty ◽  
...  
Keyword(s):  
Electrical Stimulation ◽  
Motor Cortex ◽  
Electrochemical Performance ◽  
Failure Modes ◽  
Cortical Network ◽  
Network Activity ◽  
Rodent Models ◽  
Electrode Arrays ◽  
Chronic Recording ◽  
Wire Breakage

Multisite implantable electrode arrays serve as a tool to understand cortical network connectivity and plasticity. Furthermore, they enable electrical stimulation to drive plasticity, study motor/sensory mapping, or provide network input for controlling brain-computer interfaces. Neurobehavioral rodent models are prevalent in studies of motor cortex injury and recovery as well as restoration of auditory/visual cues due to their relatively low cost and ease of training. Therefore, it is important to understand the chronic performance of relevant electrode arrays in rodent models. In this report, we evaluate the chronic recording and electrochemical performance of 16-channel Utah electrode arrays, the current state-of-the-art in pre-/clinical cortical recording and stimulation, in rat motor cortex over a period of 6 mo. The single-unit active electrode yield decreased from 52.8 ± 10.0 ( week 1) to 13.4 ± 5.1% ( week 24). Similarly, the total number of single units recorded on all electrodes across all arrays decreased from 106 to 15 over the same time period. Parallel measurements of electrochemical impedance spectra and cathodic charge storage capacity exhibited significant changes in electrochemical characteristics consistent with development of electrolyte leakage pathways over time. Additionally, measurements of maximum cathodal potential excursion indicated that only a relatively small fraction of electrodes (10–35% at 1 and 24 wk postimplantation) were capable of delivering relevant currents (20 µA at 4 nC/ph) without exceeding negative or positive electrochemical potential limits. In total, our findings suggest mainly abiotic failure modes, including mechanical wire breakage as well as degradation of conducting and insulating substrates. NEW & NOTEWORTHY Multisite implantable electrode arrays serve as a tool to record cortical network activity and enable electrical stimulation to drive plasticity or provide network feedback. The use of rodent models in these fields is prevalent. We evaluated chronic recording and electrochemical performance of 16-channel Utah electrode arrays in rat motor cortex over a period of 6 mo. We primarily observed abiotic failure modes suggestive of mechanical wire breakage and/or degradation of insulation.

scholarly journals Motor cortex activity across movement speeds is predicted by network-level strategies for generating muscle activity

2021 ◽  
Author(s):  
Shreya Saxena ◽  
Abigail A. Russo ◽  
John P. Cunningham ◽  
Mark M. Churchland
Keyword(s):  
Motor Cortex ◽  
Muscle Activity ◽  
Single Neuron ◽  
Activity Patterns ◽  
Network Activity ◽  
Movement Speed ◽  
Neural Population ◽  
Recurrent Networks ◽  
Neural Signals ◽  
Population Activity

AbstractLearned movements can be skillfully performed at different paces. What neural strategies produce this flexibility? Can they be predicted and understood by network modeling? We trained monkeys to perform a cycling task at different speeds, and trained artificial recurrent networks to generate the empirical muscle-activity patterns. Network solutions reflected the principle that smooth well-behaved dynamics require low trajectory tangling, and yielded quantitative and qualitative predictions. To evaluate predictions, we recorded motor cortex population activity during the same task. Responses supported the hypothesis that the dominant neural signals reflect not muscle activity, but network-level strategies for generating muscle activity. Single-neuron responses were better accounted for by network activity than by muscle activity. Similarly, neural population trajectories shared their organization not with muscle trajectories, but with network solutions. Thus, cortical activity could be understood based on the need to generate muscle activity via dynamics that allow smooth, robust control over movement speed.

scholarly journals Stress related network activity in the intact adrenal medulla

2021 ◽  
Author(s):  
Jose R. Lopez Ruiz ◽  
Stephen A. Ernst ◽  
Ronald W. Holz ◽  
Edward L. Stuenkel
Keyword(s):  
Stress Response ◽  
Adrenal Medulla ◽  
Chromaffin Cells ◽  
Physiological Stress ◽  
Critical Role ◽  
Splanchnic Nerve ◽  
Network Activity ◽  
Electrode Arrays ◽  
Functional Architecture ◽  
Local Networks

AbstractThe adrenal medulla has long been recognized as playing a critical role in mammalian homeostasis and the stress response. The adrenal medulla is populated by clustered chromaffin cells that secrete epinephrine or norepinephrine along with other peptides into the general bloodstream affecting multiple distant target organs. Although the sympatho-adrenal pathway has been heavily studied, detailed knowledge on the central control and in-situ spatiotemporal responsiveness remains poorly understood. For this work we implemented electrophysiological techniques originally developed to elucidate CNS circuitry to characterize the functional micro-architecture of the adrenal medulla. To achieve this, we continuously monitored the electrical activity inside the adrenal medulla in the living anesthetized rat under basal conditions and under physiological stress. Under basal conditions, chromaffin cells fired action potentials with frequencies between ∼0.2 and 4 Hz. Activity was exclusively driven by sympathetic inputs coming through the splanchnic nerve. Furthermore, chromaffin cells were organized into arrays of independent local networks in which cells fire in a specific order, with latencies from hundreds of microseconds to few milliseconds. Electrical stimulation of the splanchnic nerve evoked the exact same spatiotemporal firing patterns that occurred spontaneously. Induction of hypoglycemic stress by administration of insulin resulted in an increase in the activity of a subset of the chromaffin cell networks. In contrast, respiratory arrest induced by anesthesia overdose resulted in an increase in the activity of the entire adrenal medulla before cessation of all activity when the animal died. The results suggest the differential activation of specific networks inside the adrenal gland depending on the stressor. These results revealed a surprisingly complex electrical organization and circuitry of the adrenal medulla that likely reflects the dynamic nature of its neuroendocrine output during basal conditions and during different types of physiological stress. To our knowledge, these experiments are the first to use multi-electrode arrays in vivo to examine the electrical and functional architecture of any endocrine gland.Significance StatementStress from extrinsic (environmental, psychological) and intrinsic (biological) challenges plays a critical role in disturbing the homeostatic balance. While the body’s responses to stress are designed to ameliorate these imbalances, prolonged and dysregulated stress often drives adverse health consequences in many chronic illnesses. The better understanding of the sympatho-adrenal stress response, will potentially impact and improve the treatment of several stress related illnesses. This work focusses on the study of the functional architecture of the adrenal medulla, a key component in neuronal stress response.

scholarly journals L-type voltage-gated calcium channel regulation of in vitro human cortical neuronal networks

2018 ◽  
Author(s):  
William Plumbly ◽  
Nicholas J. Brandon ◽  
Tarek Z. Deeb ◽  
Jeremy Hall ◽  
Adrian J. Harwood
Keyword(s):  
Calcium Channel ◽  
Neuronal Networks ◽  
Gene Mutations ◽  
Synaptic Function ◽  
Neuronal Firing ◽  
Network Activity ◽  
Electrode Arrays ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channel

The combination of in vitro multi-electrode arrays (MEAs) and the neuronal differentiation of stem cells offers the capability to study human neuronal networks from patient or engineered human cell lines. Here, we use MEA-based assays to probe synaptic function and network interactions of hiPSC-derived neurons. Neuronal network behaviour first emerges at approximately 30 days of culture and is driven by glutamate neurotransmission. Over a further 30 days, inhibitory GABergic signalling shapes network behaviour into a synchronous regular pattern of burst firing activity and low activity periods. Gene mutations in L-type voltage gated calcium channel subunit genes are strongly implicated as genetic risk factors for the development of schizophrenia and bipolar disorder. We find that, although basal neuronal firing rate is unaffected, there is a dose-dependent effect of L-type voltage gated calcium channel inhibitors on synchronous firing patterns of our hiPSC-derived neural networks. This demonstrates that MEA assays have sufficient sensitivity to detect changes in patterns of neuronal interaction that may arise from hypo-function of psychiatric risk genes. Our study highlights the utility of in vitro MEA based platforms for the study of hiPSC neural network activity and their potential use in novel compound screening.

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