Characterization of Early Cortical Neural Network Development in Multiwell Microelectrode Array Plates

Cotterill, Ellese; Hall, Diana; Wallace, Kathleen; Mundy, William R.; Eglen, Stephen J.; Shafer, Timothy J.

doi:10.1177/1087057116640520

Cited by 63 publications

(52 citation statements)

References 38 publications

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“…These results suggest the network is likely most active and stable at DIV 14 in our cultures, as we have observed previously [18]. We therefore chose DIV 14, similar to other previous studies [24–27], to perform our recordings. To induce ER stress, we treated the cultures at DIV 14 with DMSO or Tg (1 μM) for one hour.…”

Section: Resultssupporting

confidence: 63%

Novel roles of ER stress in repressing neural activity and seizures through Mdm2- and p53-dependent protein translation

2019

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Seizures can induce endoplasmic reticulum (ER) stress, and sustained ER stress contributes to neuronal death after epileptic seizures. Despite the recent debate on whether inhibiting ER stress can reduce neuronal death after seizures, whether and how ER stress impacts neural activity and seizures remain unclear. In this study, we discovered that the acute ER stress response functions to repress neural activity through a protein translation-dependent mechanism. We found that inducing ER stress promotes the expression and distribution of murine double minute-2 (Mdm2) in the nucleus, leading to ubiquitination and down-regulation of the tumor suppressor p53. Reduction of p53 subsequently maintains protein translation, before the onset of translational repression seen during the latter phase of the ER stress response. Disruption of Mdm2 in an Mdm2 conditional knockdown (cKD) mouse model impairs ER stress-induced p53 down-regulation, protein translation, and reduction of neural activity and seizure severity. Importantly, these defects in Mdm2 cKD mice were restored by both pharmacological and genetic inhibition of p53 to mimic the inactivation of p53 seen during ER stress. Altogether, our study uncovered a novel mechanism by which neurons respond to acute ER stress. Further, this mechanism plays a beneficial role in reducing neural activity and seizure severity. These findings caution against inhibition of ER stress as a neuroprotective strategy for seizures, epilepsies, and other pathological conditions associated with excessive neural activity.

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Section: Resultssupporting

confidence: 63%

Novel roles of ER stress in repressing neural activity and seizures through Mdm2- and p53-dependent protein translation

2019

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“…Consistent with morphological development, electric spiking activity occurs as early as 3 DIV (Siebler et al. 1993), whereas the synchrony of spike activity (Cotterill et al. 2016; Frega et al.…”

Section: Introductionmentioning

confidence: 87%

Influence of Nanomolar Deltamethrin on the Hallmarks of Primary Cultured Cortical Neuronal Network and the Role of Ryanodine Receptors

Zheng

Feng

et al. 2019

Environ Health Perspect

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Background:The pyrethroid deltamethrin (DM) is broadly used for insect control. Although DM hyperexcites neuronal networks by delaying inactivation of axonal voltage-dependent Na+ channels, this mechanism is unlikely to mediate neurotoxicity at lower exposure levels during critical perinatal periods in mammals.Objectives:We aimed to identify mechanisms by which acute and subchronic DM altered axonal and dendritic growth, patterns of synchronous Ca2+ oscillations (SCOs), and electrical spike activity (ESA) functions critical to neuronal network formation.Methods:Measurements of SCOs using Ca2+ imaging, ESA using microelectrode array (MEA) technology, and dendritic complexity using Sholl analysis were performed in primary murine cortical neurons from wild-type (WT) and/or ryanodine receptor 1 (RyR1T4826I/T4826I) mice between 5 and 14 d in vitro (DIV). [H3]ryanodine binding analysis and a single-channel voltage clamp were utilized to measure engagement of RyRs as a direct target of DM.Results:Neuronal networks responded to DM (30–70 nM) as early as 5 DIV, reducing SCO amplitude and depressing ESA and burst frequencies by 60–70%. DM (10–300 nM) enhanced axonal growth in a nonmonotonic manner. DM≥100 nM enhanced dendritic complexity. DM stabilized channel open states of RyR1, RyR2, and cortical preparations expressing all three isoforms. DM (30 nM) altered gating kinetics of RyR1 channels, increasing mean open time, decreasing mean closed time, and thereby enhancing overall open probability. SCO patterns from cortical networks expressing RyR1T4826I/T4826I were more responsive to DM than WT. RyR1T4826I/T4826I neurons showed inherently longer axonal lengths than WT neurons and maintained less length-promoting responses to nanomolar DM.Conclusions:Our findings suggested that RyRs were sensitive molecular targets of DM with functional consequences likely relevant for mediating abnormal neuronal network connectivity in vitro. https://doi.org/10.1289/EHP4583

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“…The developing co-cultures were treated with the mitosis inhibitor 5-fluoro-2'-deoxyuridine (25μM) and uridine (63 μM) for 48 h on day 5 after seeding to prevent further glial proliferation. After 4 weeks in culture, the activity pattern stabilizes and is composed of one coordinated main burst pattern with several coordinated sub-patterns [ 18 , 19 , 25 – 29 ]. In this study cultures between 28 and 35 days in vitro were used.…”

Section: Methodsmentioning

confidence: 99%

Functional characterization of GABAA receptor-mediated modulation of cortical neuron network activity in microelectrode array recordings

et al. 2017

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The numerous γ-aminobutyric acid type A receptor (GABAAR) subtypes are differentially expressed and mediate distinct functions at neuronal level. In this study we have investigated GABAAR-mediated modulation of the spontaneous activity patterns of primary neuronal networks from murine frontal cortex by characterizing the effects induced by a wide selection of pharmacological tools at a plethora of activity parameters in microelectrode array (MEA) recordings. The basic characteristics of the primary cortical neurons used in the recordings were studied in some detail, and the expression levels of various GABAAR subunits were investigated by western blotting and RT-qPCR. In the MEA recordings, the pan-GABAAR agonist muscimol and the GABABR agonist baclofen were observed to mediate phenotypically distinct changes in cortical network activity. Selective augmentation of αβγ GABAAR signaling by diazepam and of δ-containing GABAAR (δ-GABAAR) signaling by DS1 produced pronounced changes in the majority of the activity parameters, both drugs mediating similar patterns of activity changes as muscimol. The apparent importance of δ-GABAAR signaling for network activity was largely corroborated by the effects induced by the functionally selective δ-GABAAR agonists THIP and Thio-THIP, whereas the δ-GABAAR selective potentiator DS2 only mediated modest effects on network activity, even when co-applied with low THIP concentrations. Interestingly, diazepam exhibited dramatically right-shifted concentration-response relationships at many of the activity parameters when co-applied with a trace concentration of DS1 compared to when applied alone. In contrast, the potencies and efficacies displayed by DS1 at the networks were not substantially altered by the concomitant presence of diazepam. In conclusion, the holistic nature of the information extractable from the MEA recordings offers interesting insights into the contributions of various GABAAR subtypes/subgroups to cortical network activity and the putative functional interplay between these receptors in these neurons.

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Characterization of Early Cortical Neural Network Development in Multiwell Microelectrode Array Plates

Cited by 63 publications

References 38 publications

Novel roles of ER stress in repressing neural activity and seizures through Mdm2- and p53-dependent protein translation

Novel roles of ER stress in repressing neural activity and seizures through Mdm2- and p53-dependent protein translation

Influence of Nanomolar Deltamethrin on the Hallmarks of Primary Cultured Cortical Neuronal Network and the Role of Ryanodine Receptors

Functional characterization of GABAA receptor-mediated modulation of cortical neuron network activity in microelectrode array recordings

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