We have collected a comprehensive set of multi-unit data on dissociated cortical cultures. Previous studies of the development of the electrical activity of dissociated cultures of cortical neurons each focused on limited aspects of its dynamics, and were often based on small numbers of observed cultures. We followed 58 cultures of different densities – 3000 to 50,000 neurons on areas of 30 to 75 mm2 – growing on multi-electrode arrays (MEAs) during the first five weeks of their development. Plating density had a profound effect on development. While the aggregate spike detection rate scaled linearly with density, as expected from the number of cells in proximity to electrodes, dense cultures started to exhibit bursting behavior earlier in development than sparser cultures. Analysis of responses to electrical stimulation suggests that axonal outgrowth likewise occurred faster in dense cultures. After two weeks, the network activity was dominated by population bursts in most cultures. In contrast to previous reports, development continued with changing burst patterns throughout the observation period. Burst patterns were extremely varied, with inter-burst intervals between 1 and 300 s, different amounts of temporal clustering of bursts, and different firing rate profiles during bursts. During certain stages of development bursts were organized into tight clusters with highly conserved internal structure. Dissociated cultures of cortical cells exhibited a much richer repertoire of activity patterns than previously reported. Except for the very sparsest cultures, all cultures exhibited globally synchronized bursts, but bursting patterns changed over the course of development, and varied considerably between preparations. This emphasizes the importance of using multiple preparations – not just multiple cultures from one preparation – in any study involving neuronal cultures. These results are based on 963 half-hour-long recordings. To encourage further investigation of the rich range of behaviors exhibited by cortical cells in vitro, we are making the data available to other researchers, together with Matlab code to facilitate access.

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An extremely rich repertoire of bursting patterns during the development of cortical cultures

Maternal licking and grooming (LG) in infancy influences stress responsiveness and cognitive performance in the offspring. We examined the effects of variation in the frequency of pup LG on morphological, electrophysiological, and behavioral aspects of hippocampal synaptic plasticity under basal and stress-like conditions. We found shorter dendritic branch length and lower spine density in CA1 cells from the adult offspring of low compared with high LG offspring. We also observed dramatic effects on long-term potentiation (LTP) depending on corticosterone treatment. Low LG offspring, in contrast to those of high LG mothers, displayed significantly impaired LTP under basal conditions but surprisingly a significantly enhanced LTP in response to high corticosterone in vitro. This enhanced plasticity under conditions that mimic those of a stressful event was apparent in vivo. Adult low LG offspring displayed enhanced memory relative to high LG offspring when tested in a hippocampal-dependent, contextual fear-conditioning paradigm. Hippocampal levels of glucocorticoid and mineralocorticoid receptors were reduced in low compared with high LG offspring. Such effects, as well as the differences in dendritic morphology, likely contribute to LTP differences under resting conditions, as well as to the maternal effects on synaptic plasticity and behavior in response to elevated corticosterone levels. These results suggest that maternal effects may modulate optimal cognitive functioning in environments varying in demand in later life, with offspring of high and low LG mothers showing enhanced learning under contexts of low and high stress, respectively.

https://www.jneurosci.org/content/jneuro/28/23/6037.full.pdf

Maternal care and hippocampal plasticity: evidence for experience-dependent structural plasticity, altered synaptic functioning, and differential responsiveness to glucocorticoids and stress

Ion channels represent highly attractive targets for drug discovery and are implicated in a diverse range of disorders, in particular in the central nervous and cardiovascular systems. Moreover, assessment of cardiac ion-channel activity of new chemical entities is now an integral component of drug discovery programmes to assess potential for cardiovascular side effects. Despite their attractiveness as drug discovery targets ion channels remain an under-exploited target class, which is in large part due to the labour-intensive and low-throughput nature of patch-clamp electrophysiology. This Review provides an update on the current state-of-the-art for the various automated electrophysiology platforms that are now available and critically evaluates their impact in terms of ion-channel screening, lead optimization and the assessment of cardiac ion-channel safety liability.

High-throughput electrophysiology: an emerging paradigm for ion-channel screening and physiology

1. Introduction 631.1 Outline 631.2 Universals versus realizations in the study of learning and memory 642. Large random cortical networks developing ex vivo 652.1 Preparation 652.2 Measuring electrical activity 673. Spontaneous development 693.1 Activity 693.2 Connectivity 704. Consequences of spontaneous activity: pharmacological manipulations 724.1 Structural consequences 724.2 Functional consequences 735. Effects of stimulation 745.1 Response to focal stimulation 745.2 Stimulation-induced changes in connectivity 746. Embedding functionality in real neural networks 776.1 Facing the physiological definition of ‘reward’: two classes of theories 786.2 Closing the loop 797. Concluding remarks 848. Acknowledgments 859. References 85The phenomena of learning and memory are inherent to neural systems that differ from each other markedly. The differences, at the molecular, cellular and anatomical levels, reflect the wealth of possible instantiations of two neural learning and memory universals: (i) an extensive functional connectivity that enables a large repertoire of possible responses to stimuli; and (ii) sensitivity of the functional connectivity to activity, allowing for selection of adaptive responses. These universals can now be fully realized in ex-vivo developing neuronal networks due to advances in multi-electrode recording techniques and desktop computing. Applied to the study of ex-vivo networks of neurons, these approaches provide a unique view into learning and memory in networks, over a wide range of spatio-temporal scales. In this review, we summarize experimental data obtained from large random developing ex-vivo cortical networks. We describe how these networks are prepared, their structure, stages of functional development, and the forms of spontaneous activity they exhibit (Sections 2–4). In Section 5 we describe studies that seek to characterize the rules of activity-dependent changes in neural ensembles and their relation to monosynaptic rules. In Section 6, we demonstrate that it is possible to embed functionality into ex-vivo networks, that is, to teach them to perform desired firing patterns in both time and space. This requires ‘closing a loop’ between the network and the environment. Section 7 emphasizes the potential of ex-vivo developing cortical networks in the study of neural learning and memory universals. This may be achieved by combining closed loop experiments and ensemble-defined rules of activity-dependent change.

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Development, learning and memory in large random networks of cortical neurons: lessons beyond anatomy.

Cognitive processes depend on synchronization and propagation of electrical activity within and between neuronal assemblies. In vivo measurements show that the size of individual assemblies depends on their function and varies considerably, but the timescale of assembly activation is in the range of 0.1–0.2 s and is primarily independent of assembly size. Here we use an in vitro experimental model of cortical assemblies to characterize the process underlying the timescale of synchronization, its relationship to the effective topology of connectivity within an assembly, and its impact on propagation of activity within and between assemblies. We show that the basic mode of assembly activation, “network spike,” is a threshold-governed, synchronized population event of 0.1–0.2 s duration and follows the logistics of neuronal recruitment in an effectively scale-free connected network. Accordingly, the sequence of neuronal activation within a network spike is nonrandom and hierarchical; a small subset of neurons is consistently recruited tens of milliseconds before others. Theory predicts that scale-free topology allows for synchronization time that does not increase markedly with network size; our experiments with networks of different densities support this prediction. The activity of early-to-fire neurons reliably forecasts an upcoming network spike and provides means for expedited propagation between assemblies. We demonstrate this capacity by observing the dynamics of two artificially coupled assemblies in vitro , using neuronal activity of one as a trigger for electrical stimulation of the other.

https://www.jneurosci.org/content/jneuro/26/33/8465.full.pdf

Dynamics and Effective Topology Underlying Synchronization in Networks of Cortical Neurons

Extracellular action potentials were recorded from developing dissociated rat neocortical networks continuously for up to 49 days in vitro using planar multielectrode arrays. Spontaneous neuronal activity emerged toward the end of the first week in vitro and from then on exhibited periods of elevated firing rates, lasting for a few days up to weeks, which were largely uncorrelated among different recording sites. On a time scale of seconds to minutes, network activity typically displayed an ongoing repetition of distinctive firing patterns, including short episodes of synchronous firing at many sites ( network bursts). Network bursts were highly variable in their individual spatio-temporal firing patterns but showed a remarkably stable underlying probabilistic structure (obtained by summing consecutive bursts) on a time scale of hours. On still longer time scales, network bursts evolved gradually, with a significant broadening (to about 2 s) in the third week in vitro, followed by a drastic shortening after about one month in vitro. Bursts at this age were characterized by highly synchronized onsets reaching peak firing levels within less than ca. 60 ms. This pattern persisted for the rest of the culture period. Throughout the recording period, active sites showed highly persistent temporal relationships within network bursts. These longitudinal recordings of network firing have, thus, brought to light a reproducible pattern of complex changes in spontaneous firing dynamics of bursts during the development of isolated cortical neurons into synaptically interconnected networks.

Long-term characterization of firing dynamics of spontaneous bursts in cultured neural networks

Longterm stability and developmental changes in spontaneous network burst firing patterns in dissociated rat cerebral cortex cell cultures on multielectrode arrays.

When dissociated cortical tissue is brought into culture, neurons readily grow out by forming axonal and dendritic arborizations and synaptic connections. These developing neuronal networks in vitro display spontaneous firing activity from about the end of the first week in vitro. When cultured on multielectrode arrays firing activity can be recorded from many neurons simultaneously over long periods of time. These experimental approaches provide valuable data for studying firing dynamics in neuronal networks in relation to an ongoing development of neurons and synaptic connectivity in the network. This chapter summarizes recent findings on the characteristics and developmental changes in the spontaneous firing dynamics. These changes include long-lasting transient periods of increased firing at individual sites on a time scale of days to weeks, and an age-specific repetitive pattern of synchronous network firing (network bursts) on a time scale of seconds. Especially the spatio-temporal organization of firing within network bursts showed great stability over many hours. In addition, a progressive day-to-day evolution was observed, with an initial broadening of the burst firing rate profile during the 3rd week in vitro (WIV) and a pattern of abrupt onset and precise spike timing from the 5th WIV onwards. These developmental changes are discussed in the light of structural changes in the network and activity-dependent plasticity mechanisms. Preliminary findings are presented on the pattern of spike sequences within network burst, as well as the effect of external stimulation on the spatio-temporal organization within network bursts.

Michael A. Corner

Papers

Long-term characterization of firing dynamics of spontaneous bursts in cultured neural networks

Longterm stability and developmental changes in spontaneous network burst firing patterns in dissociated rat cerebral cortex cell cultures on multielectrode arrays.

Dynamics and plasticity in developing neuronal networks in vitro.

Implications of activity dependent neurite outgrowth for neuronal morphology and network development.

Spontaneous neuronal discharge patterns in developing organotypic mega-co-cultures of neonatal rat cerebral cortex.