A multiplexed assay for determination of neurotoxicant effects on spontaneous network activity and viability from microelectrode arrays

Microelectrode array (MEA) recordings are increasingly being used as an in vitro method to detect and characterize the ability of drugs, chemicals and particles to cause neurotoxicity. While compound effects on spontaneous network activity are easily determined by MEA recordings, compound cytotoxicity is not routinely assessed, particularly within the same network from which recordings are collected. With the advent of higher-throughput 48 and 96 well MEA systems, rapid and simple methods to measure compound effects on cell health are required to facilitate efficient compound screening using MEAs. The present experiments sought to develop a multiplexed approach that allows measurement of network activity and cell health in the same MEA well. Primary cultures from rat cortex were exposed to six different compounds (glyphosate, β-cyfluthrin, domoic acid, tributyltin, lindane and fipronil). Effects of these compounds (0.03–100 μM) on spontaneous network activity (mean firing rate; MFR), cellular metabolic activity (Cell Titer Blue™ (CTB) assay) and lactate dehydrogenase (LDH) release were determined in the same well following a 60-min exposure. Glyphosate elicited no effect on MFR, LDH release or CTB reduction. Tributyltin caused concomitant decreases in MFR and CTB reduction and increases LDH release, while domoic acid and β-cyfluthrin decreased MFR in a concentration-dependent manner without altering either LDH release or CTB reduction. By contrast, lindane and fipronil did not alter LDH release or CTB reduction, but caused biphasic alterations in MFR, with increases in MFR at lower concentrations followed by decreases at higher concentrations. These results demonstrate a simple and rapid method for the simultaneous determination of test compound effects on spontaneous electrical activity and cell health from the same network, and will facilitate rapid screening of compounds for potential neurotoxicity.     

Effects of an environmentally-relevant mixture of pyrethroid insecticides on spontaneous activity in primary cortical networks on microelectrode arrays

Pyrethroid insecticides exert their insecticidal and toxicological effects primarily by disrupting voltage-gated sodium channel (VGSC) function, resulting in altered neuronal excitability. Numerous studies of individual pyrethroids have characterized effects on mammalian VGSC function and neuronal excitability, yet studies examining effects of complex pyrethroid mixtures in mammalian neurons, especially in environmentally relevant mixture ratios, are limited. In the present study, concentration-response functions were characterized for five pyrethroids (permethrin, deltamethrin, cypermethrin, β-cyfluthrin and esfenvalerate) in an in vitro preparation containing cortical neurons and glia. As a metric of neuronal network activity, spontaneous mean network firing rates (MFR) were measured using microelectorde arrays (MEAs). In addition, the effect of a complex and exposure relevant mixture of the five pyrethroids (containing 52% permethrin, 28.8% cypermethrin, 12.9% β-cyfluthrin, 3.4% deltamethrin and 2.7% esfenvalerate) was also measured. Data were modeled to determine whether effects of the pyrethroid mixture were predicted by dose-addition. At concentrations up to 10 μM, all compounds except permethrin reduced MFR. Deltamethrin and β-cyfluthrin were the most potent and reduced MFR by as much as 60 and 50%, respectively, while cypermethrin and esfenvalerate were of approximately equal potency and reduced MFR by only ∼20% at the highest concentration. Permethrin caused small (∼24% maximum), concentration-dependent increases in MFR. Effects of the environmentally relevant mixture did not depart from the prediction of dose-addition. These data demonstrate that an environmentally relevant mixture caused dose-additive effects on spontaneous neuronal network activity in vitro, and is consistent with other in vitro and in vivo assessments of pyrethroid mixtures.     

Effect of acute organophosphorus exposure on 2', 3',-cyclic nucleotide 3'-phosphohydrolase activity in hen neural tissue

The effect of various organophosphorus (OP) compounds on 2', 3'-cyclic nucleotide 3'-phosphohydrolase (CNP) activity and the relationship to neurotoxic response was studied. Adult White Leghorn hens were dosed with either diisopropyl fluorophosphate (DFP), tri-o-cresyl phosphate (TOCP), leptophos, or paraoxon. CNP activity was determined utilizing crude homogenates or purified myelin preparations from whole brain, spinal cord, and sciatic nerves at maximal neurologic dysfunction, evaluated as locomotor impairment. Enzyme assays of CNP revealed a decrease in brain CNP activity at the time of maximal locomotor impairment after a single, oral dose of leptophos. Decreased CNP activity was also noted in spinal cord preparations from DFP-treated hens at the time of maximal locomotor impairment. Sciatic nerve CNP activity was not altered following treatment with OP-compounds. Paraoxon, a non-neurotoxic OP-compound, had little effect on neural tissue CNP activity. Neurotoxic OP-compounds, that cause secondary degeneration of myelin (Wallerian), may be associated with decreased brain and spinal cord CNP activity at the time of maximal locomotor impairment.     

State-dependent, bidirectional modulation of neural network activity by endocannabinoids

The endocannabinoid (eCB) system and the cannabinoid CB1 receptor (CB1R) play key roles in the modulation of brain functions. Although actions of eCBs and CB1Rs are well described at the synaptic level, little is known of their modulation of neural activity at the network level. Using microelectrode arrays, we have examined the role of CB1R activation in the modulation of the electrical activity of rat and mice cortical neural networks in vitro. We find that exogenous activation of CB1Rs expressed on glutamatergic neurons decreases the spontaneous activity of cortical neural networks. Moreover, we observe that the net effect of the CB1R antagonist AM251 inversely correlates with the initial level of activity in the network: blocking CB1Rs increases network activity when basal network activity is low, whereas it depresses spontaneous activity when its initial level is high. Our results reveal a complex role of CB1Rs in shaping spontaneous network activity, and suggest that the outcome of endogenous neuromodulation on network function might be state dependent.     

Toxic neurofilamentous axonopathies and fast anterograde axonal transport. II. The effects of single doses of neurotoxic and non-neurotoxic diketones and beta, beta'-iminodipropionitrile (IDPN) on the rate and capacity of transport

The site and mode of action of neurotoxic chemicals producing neurofilamentous axonopathies has been speculated to be the axonal transport system. The current study determined the effects of neurotoxic and non-neurotoxic gamma-diketones as well as beta, beta'-iminodipropionitrile (IDPN) upon both the rate and quantity of protein transported in the fast anterograde component of the rat sciatic nerve. 2,5-Hexanedione (2,5-HD), given as 4, 6 and 8 mmoles/kg single ip injections reduced the rate of transport by 18.4-24.7% but more significantly reduced the quantity of protein transported 50-63%. 3,4-Dimethyl-2,5-HD (3,4-DMHD) at single doses of 0.25, 0.50 and 1.0 mmoles/kg similarly reduced the rate and capacity of protein transport. The toxicants did not alter the uptake of leucine and synthesis of protein during the three hour time frame used to measure transport. Although high doses of IDPN reduced the rate of anterograde transport, this toxicant, as well as the non-neurotoxic diketones studied, had no effect upon the quantity of protein transported. Therefore, neurotoxic gamma-diketones which produce distal nerve degeneration had a common effect in decreasing the quantity of protein delivered to the nerve after just a single exposure.     

Development of spontaneous neuronal activity in the caudate nucleus, globus pallidus-entopeduncular nucleus, and substantia nigra of the cat

Spontaneous single unit activity was obtained from caudate (Cd), globus pallidus-entopeduncular nucleus (GP-Ento), and substantia nigra (SN) neurons in kittens of 1-60 days of age and adult cats. Five developmental trends were found in the spontaneous firing patterns of these neurons: (1) overall mean interspike intervals (ISIs) decreased with age; (2) the occurrence of neurons with shorter mean ISIs (less than 400 ms) increased with age; (3) the occurrence of neurons with burst activity increased with age; (4) burst activity became more complex with age; and (5) the rate of burst occurrence in neurons with burst activity increased with age. Neurons within each region of the basal ganglia had characteristic patterns of spontaneous activity. Furthermore, the developmental patterns of spontaneous neuronal activity were different in each structure. The spontaneous activity of GP-Ento and SN neurons matured before the spontaneous activity of Cd neurons. Thus, spontaneous firing may mature in the output nuclei of the basal ganglia prior to its maturation in the Cd.     

Irregular activation of individual sweat glands in human sole observed by a videomicroscopy

Sweat secretion from individual sweat glands on the human sole was observed in four male subjects by using a videomicroscope and correlated with sudomotor neural activity recorded from the tibial nerve by means of microneurography. Individual sweat glands could be distinguished as active, less active and inactive according to the incidence of sweat secretion during spontaneous sweating. The threshold amplitude of the sudomotor burst necessary for sweat secretion varied from gland to gland. The number of sweat secretion was significantly related to the threshold amplitude. Sweat glands often failed to produce sweat secretion even when a suprathreshold burst occurred: only 46.1+/-3.8% (mean +/- S.E.M.) of the suprathreshold bursts elicited sweat secretion. Failure of the sweat secretion tended to appear after several bursts occurred consecutively with short intervals. In spite of the variability in sweat gland activity, the number of sweat glands recruited was linearly related to the amplitude of the sudomotor burst (P < 0.001). Thus, although sweat secretion from each sweat gland depends primarily on the intensity of sudomotor neural activity. the activity of each sweat gland may fluctuate temporally as the result of irregular activation of sudomotor fibers and possibly some intrinsic factors of the gland.     

Alteration of the electrophysiological activity in symphathetic ganglia infected with a neurotropic virus. I. Presynaptic origin of the spontaneous bioelectric activity

The bioelectric activity of the rat superior cervical ganglion (SCG) infected with pseudorabies virus (PRV) was examined in vitro 30–38 h after inoculation. Simultaneous intra- and extracellular recordings on the internal (ICN) and external carotid nerves (ECN) revealed a synchronized spontaneous activity. This synchronization can be ascribed either to the functional organization of the ganglion or to the mechanism of initiation itself.In the infected ganglia two categories of cells were observed: cells displaying abnormal spontaneous discharges, and silent cells whose electrophysiological behavior was similar to control cells.Spontaneously active cells showed intermittent spiking and bursting activity. The discharge pattern was associated with the firing rate of the emitting cell: sporadically active cells emitted single spikes whereas highly active cells fired bursts of action potentials (APs). Long lasting intracellular recordings demonstrated that the cells undergo gradual changes evolving from sporadic on to high activity.Spontaneous APs usually rode on prepotentials similar to the excitatory postsynaptic potentials (EPSPs). A comparative study of spontaneous prepotentials and orthodromically evoked EPSPs in the same cell demonstrated that the spontaneous prepotentials are real synaptic potentials. No pace-maker potentials were observed.The passive and active electrical membrane properties of spontaneously active neurons were not different from those of silent cells or control cells impaled in uninfected ganglia.d-Tubocurarine abolished the spontaneous activity in the whole ganglion.Ortho- and antidromic electrical stimulations of suprathreshold intensity elicited an evoked response in neurons displaying spontaneous activity, followed by a delayed burst whose shape was similar to the spontaneous burst of the cell. Stimuli of subthreshold intensities induced this delayed burst independently from the evoked response.We conclude that the spontaneous bioelectrical activity is of presynaptic, but not necessarily of preganglionic origin. The possible existence of a cholinergic intraganglionic pathway revealed by the viral infection is discussed.     

Chronic electrical stimulation of cultured hippocampal networks increases spontaneous spike rates

We chronically stimulated hippocampal networks in culture for either 0, 1 or 3 h/day between 7 and 22 days in culture in an effort to increase spontaneous spike rates and to give these networks some portion of external stimuli that brain networks receive during their formation. Chronic electrical stimulation of hippocampal networks on multi-electrode arrays (MEAs) increased spike rates 2-fold after 3 weeks of culture compared to cultures that received no external stimulation prior to recording. More than 90% of the spikes for all experimental conditions occurred within bursts. The frequency of spikes within a burst increased with time of stimulation during culture up to 2-fold higher (90 Hz) compared to networks without chronic stimulation. However, spontaneous overall spike rates did not correlate well with the amount of stimulation either as h/day or proximity to the limited number of stimulation sites due to shorter burst duration with 3 h/day stimulation. The results suggest that chronic stimulation applied during network development recruits activity at 50% more electrodes and enables higher rates of spontaneous activity within bursts in cultured hippocampal networks.     

Spontaneous synchronous synaptic calcium transients in cultured cortical neurons.

The firing pattern displayed by neuronal aggregates is thought to play a key role in cortical development and physiology. In this study, we have employed optical recording of intracellular calcium to monitor activity of multiple neurons simultaneously in primary cortical cultures. With this approach, we have observed spontaneous synchronous calcium transients among adjacent cortical neurons. These transients appear to be mediated by prominent spontaneous synaptic excitation, as they are enhanced by picrotoxin, a blocker of inhibitory GABAergic transmission, and reduced by antagonism of glutamate receptors or addition of TTX. After picrotoxin treatment, the calcium transients exhibit regular frequency and amplitude, and occur in synchrony with bursts of excitatory synaptic potentials every 10-20 sec. Using electrical stimulation, we have identified a relative refractory period, extending up to 5 sec after a synchronous burst, that may play a role in cell synchronization. NMDA receptor antagonists or reduced extracellular calcium levels lower the amplitude of the calcium transients yet fail to alter their frequency, suggesting that intracellular calcium levels may not be a major determinant of burst frequency. In contrast, mild depolarization with kainic acid (0.5-1 microM) increased burst frequency up to fivefold, suggesting a critical dependence of rhythmic activity on membrane potential. Chronic blockade of electrical activity with TTX beginning a few days after plating of cultures dampens the amplitude and significantly increases the frequency of calcium transients in mature cultures. These studies demonstrate that aggregates of cultured cortical neurons express synchronous firing activity in vitro and that this network activity is dependent in part on neuronal firing during development.     

Sulforhodamine labeling of neural circuits engaged in motor pattern generation in the in vitro turtle brainstem-cerebellum.

A fluorescent molecular probe was used in combination with a novel in vitro preparation to study spatial patterns of neural activity associated with motor pattern generation. The in vitro brainstem-cerebellum preparation takes advantage of the turtle's unusual resistance to anoxia to preserve the entire neural network that connects the cerebellum, red nucleus, and reticular formation. This preparation was bathed in a 0.01% solution of sulforhodamine while it was activated unilaterally by electrical stimulation of the dorsal quadrant of the spinal cord for 1 hr. Sulforhodamine is a small, sulfonated, highly charged fluorescent molecule that is taken up by endocytosis. To examine its distribution in the cerebellum and brainstem, coronal sections were prepared and viewed under epifluorescence illumination. Distinctive spatial patterns of labeling were associated with unilateral electrical stimulation of the in vitro network, suggesting that dye uptake was activity dependent. Blockade of uptake with altered magnesium and calcium concentrations indicated that single spike discharge evoked ortho- or antidromically was insufficient to induce dye uptake. Instead, sulforhodamine staining correlated with the presence of burst discharge that was recorded extracellularly from the red nucleus. Blockade of burst discharge with excitatory amino acid receptor antagonists prevented dye uptake in the red nucleus, the lateral cerebellar nucleus, and other structures that are known to be interconnected by recurrent anatomical pathways. These results suggest that sulforhodamine is internalized by intensely active neurons. The spatial distributions of label support the hypothesis that burst discharges in the turtle red nucleus are mediated by excitatory amino acid neurotransmitters and sustained by recurrent excitation in cerebellorubral synaptic pathways. Positive feedback in these recurrent pathways may provide an important driving force for the generation of motor programs that control limb movements.     

Effects of synaptic depression and recovery on synchronous network activity.

SUMMARY: The output of an artificial neural network of spiking neurons linked by glutamatergic synapses subject to use-dependent depression was compared with physiologic data obtained from rat hippocampal area CA3 in vitro. The authors evaluated how network burst initiation and termination was affected by activity-dependent depression and recovery under a variety of experimental conditions including neuronal membrane depolarization, altered glutamate release probability, the strength of synaptic inhibition, and long-term potentiation and long-term depression of recurrent glutamatergic synapses. The results of computational experiments agreed with the in vitro data and support the idea that synaptic properties, including activity-dependent depression and recovery, play important roles in the timing and duration of spontaneous bursts of network activity. This validated network model is useful for experiments that are not feasible in vitro, and makes possible the investigation of two-dimensional aspects of burst propagation and termination.     

Network burst dynamics under heterogeneous cholinergic modulation of neural firing properties and heterogeneous synaptic connectivity

The characteristics of neural network activity depend on intrinsic neural properties and synaptic connectivity in the network. In brain networks, both of these properties are critically affected by the type and levels of neuromodulators present. The expression of many of the most powerful neuromodulators, including acetylcholine (ACh), varies tonically and phasically with behavioural state, leading to dynamic, heterogeneous changes in intrinsic neural properties and synaptic connectivity properties. Namely, ACh significantly alters neural firing properties as measured by the phase response curve in a manner that has been shown to alter the propensity for network synchronization. The aim of this simulation study was to build an understanding of how heterogeneity in cholinergic modulation of neural firing properties and heterogeneity in synaptic connectivity affect the initiation and maintenance of synchronous network bursting in excitatory networks. We show that cells that display different levels of ACh modulation have differential roles in generating network activity: weakly modulated cells are necessary for burst initiation and provide synchronizing drive to the rest of the network, whereas strongly modulated cells provide the overall activity level necessary to sustain burst firing. By applying several quantitative measures of network activity, we further show that the existence of network bursting and its characteristics, such as burst duration and intraburst synchrony, are dependent on the fraction of cell types providing the synaptic connections in the network. These results suggest mechanisms underlying ACh modulation of brain oscillations and the modulation of seizure activity during sleep states.     

Modification of parallel activity elicited by propagating bursts in developing networks of rat cortical neurones

Networks of cultured cortical neurones exhibit regular, synchronized, propagating bursts which are synaptically mediated, and which are hypothesized to play a part in activity-dependent formation of connections during development in vivo. The relationship between the strength of synaptic connections and the characteristics of synchronized propagating bursting, however, is unclear. Modification of synchronized activity in cortical cultures in response to electrical stimulation was examined using multisite electrode array recording. By measuring the response of the network to weak, localized, test stimulation (TS), we observed a potentiation of activity following a relatively stronger inducing stimulation (IS). This potentiation was evident as an increased probability of eliciting bursts by TS, an increased frequency of spontaneous bursts and number of spikes per burst, and increased speed of burst propagation, and it lasted for at least 20 min. Changing the parameters of IS revealed that high frequency tetanic stimulation is not necessary to induce potentiation, while it is essential for IS to produce a regeneratively propagating burst. The results provide a direct demonstration of modification of both the spatial and temporal characteristics of synchronized network activity, and suggest an important physiological role for propagating synchronized bursting, as a mechanism for inducing plastic modifications in the developing cortex.     

Human Umbilical Cord Blood-Derived Neural Stem Cell Line as a Screening Model for Toxicity

The aim was to investigate whether a human neural stem cell (NSC) line derived from human umbilical cord blood (hUCB) can be used for toxicity study. Toxicity of both neurotoxic environmental xenobiotics, methyl mercury chloride (CH₃HgCl), lead acetate (CH₃COOPb), and chlorpyrifos (CP), and non-neurotoxic insecticide, dichlorvos, as well as non-neurotoxic drugs, theophylline and acetaminophen were assessed. Additionally, differentiation of neuronal and glial cell lines derived from hUCB was elucidated. It was observed that CH₃HgCl was more toxic to human NSCs in comparison to CH₃COOPb and CP. The minimum inhibitory concentration (MIC) value against NSCs was 3, 10, and 300 mg/L, in each staining process, acridine orange/ethidium bromide (AO/EB) staining, 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (MTT) assay, and Hoechst staining, for CH₃HgCl, CP, and CH₃COOPb, respectively. CH₃HgCl had the LC₂₅ value as 10.0, 14.4, and 12.7 mg/L, by staining method mentioned in succession. CP had the LC₂₅ value as 21.9, 23.7, and 18.4 mg/L; similarly, CH₃COOPb had LC₂₅ values, successively as 616.9, 719.2, and 890.3 mg/L. LC₅₀ values ranged from 18.2 to 21.7 mg/L for CH₃HgCl, 56.4 to 60.2 mg/L for CP, and 1000 to 1460.1 for CH₃COOPb. Theophylline, acetaminophen, and dichlorvos had no impact on the viability of NSCs. This work justified that hUCB-NSC model can be used for toxicity study.     

No phylogeny without ontogeny — a comparative and developmental search for the sources of sleep-like neural and behavioral rhythms

A comprehensive review is presented of reported aspects and putative mechanisms of sleep-like motility rhythms throughout the animal kingdom.It is proposed that ’rapid eye movement(REM)sleep’ be regarded as a special case of a distinct but much broader category of behavior,’rapid body movement(RBM)sleep’,defined by intrinsically-generated and apparently non-purposive movements.Such a classification completes a 2×2 matrix defined by the axes sleep versus waking and active versus quiet.Although ’paradoxical’ arousal of forebrain electrical activity is restricted to warm-blooded vertebrates,we urge that juvenile or even infantile stages of development be investigated in cold-blooded animals,in view of the many reports of REM-like spontaneous motility(RBMs)in a wide range of species during sleep.The neurophysiological bases for motorically active sleep at the brainstem level and for slow-wave sleep in the forebrain appear to be remarkably similar,and to be subserved in both cases by a primitive diffuse mode of neuronal organization.Thus,the spontaneous synchronous burst discharges which are characteristics of the sleeping brain can be readily simulated even by highly unstructured neural network models.Neuromotor discharges during active sleep appear to reflect a hierarchy of simple relaxation oscillation mechanisms,spanning a wide range of spike-dependent relaxation times,whereas the periodic alternation of active and quiet sleep states more likely results from the entrainment of intrinsic cellular rhythms and/or from activity-dependent homeostatic changes in network excitability.     

Parkinsonism Alters Beta Burst Dynamics across the Basal Ganglia–Motor Cortical Network

Elevated synchronized oscillatory activity in the beta band has been hypothesized to be a pathophysiological marker of Parkinson''s disease (PD). Recent studies have suggested that parkinsonism is closely associated with increased amplitude and duration of beta burst activity in the subthalamic nucleus (STN). How beta burst dynamics are altered from the normal to parkinsonian state across the basal ganglia–thalamocortical (BGTC) motor network, however, remains unclear. In this study, we simultaneously recorded local field potential activity from the STN, internal segment of the globus pallidus (GPi), and primary motor cortex (M1) in three female rhesus macaques, and characterized how beta burst activity changed as the animals transitioned from normal to progressively more severe parkinsonian states. Parkinsonism was associated with an increased incidence of beta bursts with longer duration and higher amplitude in the low beta band (8–20 Hz) in both the STN and GPi, but not in M1. We observed greater concurrence of beta burst activity, however, across all recording sites (M1, STN, and GPi) in PD. The simultaneous presence of low beta burst activity across multiple nodes of the BGTC network that increased with severity of PD motor signs provides compelling evidence in support of the hypothesis that low beta synchronized oscillations play a significant role in the underlying pathophysiology of PD. Given its immersion throughout the motor circuit, we hypothesize that this elevated beta-band activity interferes with spatial–temporal processing of information flow in the BGTC network that contributes to the impairment of motor function in PD.SIGNIFICANCE STATEMENT This study fills a knowledge gap regarding the change in temporal dynamics and coupling of beta burst activity across the basal ganglia–thalamocortical (BGTC) network during the evolution from normal to progressively more severe parkinsonian states. We observed that changes in beta oscillatory activity occur throughout BGTC and that increasing severity of parkinsonism was associated with a higher incidence of longer duration, higher amplitude low beta bursts in the basal ganglia, and increased concurrence of beta bursts across the subthalamic nucleus, globus pallidus, and motor cortex. These data provide new insights into the potential role of changes in the temporal dynamics of low beta activity within the BGTC network in the pathogenesis of Parkinson''s disease.     

Application of micro-electrode arrays (MEAs) as an emerging technology for developmental neurotoxicity: evaluation of domoic acid-induced effects in primary cultures of rat cortical neurons

Due to lack of knowledge only a few industrial chemicals have been identified as developmental neurotoxicants. Current developmental neurotoxicity (DNT) guidelines (OECD and EPA) are based entirely on in vivo studies that are both time consuming and costly. Consequently, there is a high demand to develop alternative in vitro methods for initial screening to prioritize chemicals for further DNT testing. One of the most promising tools for neurotoxicity assessment is the measurement of neuronal electrical activity using micro-electrode arrays (MEAs) that provides a functional and neuronal specific endpoint that until now has been used mainly to detect acute neurotoxicity. Here, electrical activity measurements were evaluated to be a suitable endpoint for the detection of potential developmental neurotoxicants. Initially, primary cortical neurons grown on MEA chips were characterized for different cell markers over time, using immunocytochemistry. Our results show that primary cortical neurons could be a promising in vitro model for DNT testing since some of the most critical neurodevelopment processes such as progenitor cell commitment, proliferation and differentiation of astrocytes and maturation of neurons are present. To evaluate if electrical activity could be a suitable endpoint to detect chemicals with DNT effects, our model was exposed to domoic acid (DomA), a potential developmental neurotoxicant for up to 4 weeks. Long-term exposure to a low concentration (50nM) of DomA increased the basal spontaneous electrical activity as measured by spike and burst rates. Moreover, the effect induced by the GABA(A) receptor antagonist bicuculline was significantly lower in the DomA treated cultures than in the untreated ones. The MEA measurements indicate that chronic exposure to DomA changed the spontaneous electrical activity leading to the possible neuronal mal functioning. The obtained results suggest that the MEAs could be a useful tool to identify compounds with DNT potential.