Alessandro Vato

Dissociated cortical networks show spontaneously correlated activity patterns during in vitro development.

In vitro cultured neuronal networks coupled to microelectrode arrays (MEAs) constitute a valuable experimental model for studying changes in the neuronal dynamics at different stages of development. After a few days in culture, neurons start to connect each other with functionally active synapses, forming a random network and displaying spontaneous electrophysiological activity. The patterns of collective rhythmic activity change in time spontaneously during in vitro development. Such activity-dependent modifications play a key role in the maturation of the network and reflect changes in the synaptic efficacy, fact widely recognized as a cellular basis of learning, memory and developmental plasticity. Getting advantage from the possibilities offered by the MEAs, the aim of our study is to analyze and characterize the natural changes in dynamics of the electrophysiological activity at different ages of the culture, identifying peculiar steps of the spontaneous evolution of the network. The main finding is that between the second and the third week of culture, the network completely changes its electrophysiological patterns, both in terms of spiking and bursting activity and in terms of cross-correlation between pairs of active channels. Then the maturation process can be characterized by two main phases: modulation and shaping in the synaptic functional connectivity of the network (within the first and second week) and general moderate correlated activity, spread over the entire network, with connections properly formed and stabilized (within the fourth and fifth week).

NETWORK DYNAMICS AND SYNCHRONOUS ACTIVITY IN CULTURED CORTICAL NEURONS

Neurons extracted from specific areas of the Central Nervous System (CNS), such as the hippocampus, the cortex and the spinal cord, can be cultured in vitro and coupled with a micro-electrode array (MEA) for months. After a few days, neurons connect each other with functionally active synapses, forming a random network and displaying spontaneous electrophysiological activity. In spite of their simplified level of organization, they represent an useful framework to study general information processing properties and specific basic learning mechanisms in the nervous system. These experimental preparations show patterns of collective rhythmic activity characterized by burst and spike firing. The patterns of electrophysiological activity may change as a consequence of external stimulation (i.e., chemical and/or electrical inputs) and by partly modifying the randomness of the network architecture (i.e., confining neuronal sub-populations in clusters with micro-machined barriers). In particular we investigated how the spontaneous rhythmic and synchronous activity can be modulated or drastically changed by focal electrical stimulation, pharmacological manipulation and network segregation. Our results show that burst firing and global synchronization can be enhanced or reduced; and that the degree of synchronous activity in the network can be characterized by simple parameters such as cross-correlation on burst events.

Networks of neurons coupled to microelectrode arrays: a neuronal sensory system for pharmacological applications

Two main features make microelectrode arrays (MEAs) a valuable tool for electrophysiological measurements under the perspective of pharmacological applications, namely: (i) they are non-invasive and permit, under appropriate conditions, to monitor the electrophysiological activity of neurons for a long period of time (i.e. from several hours up to months); (ii) they allow a multi-site recording (up to tens of channels). Thus, they should allow a high-throughput screening while reducing the need for animal experiments. In this paper, by taking advantages of these features, we analyze the changes in activity pattern induced by the treatment with specific substances, applied on dissociated neurons coming from the chick-embryo spinal cord. Following pioneering works by Gross and co-workers (see e.g. Gross and Kowalski, 1991. Neural Networks, Concepts, Application and Implementation, vol. 4. Prentice Hall, NJ, pp. 47-110; Gross et al., 1992. Sensors Actuators, 6, 1-8.), in this paper analysis of the drugs effects (e.g. NBQX, CTZ, MK801) to the collective electrophysiological behavior of the neuronal network in terms of burst activity, will be presented. Data are simultaneously recorded from eight electrodes and besides variations induced by the drugs also the correlation between different channels (i.e. different area in the neural network) with respect to the chemical stimuli will be introduced (Bove et al., 1997. IEEE Trans. Biomed. Eng., 44, 964-977.). Cultured spinal neurons from the chick embryo were chosen as a neurobiological system for their relative simplicity and for their reproducible spontaneous electrophysiological behavior. It is well known that neuronal networks in the developing spinal cord are spontaneously active and that the presence of a significant and reproducible bursting activity is essential for the proper formation of muscles and joints (Chub and ODonovan, 1998. J. Neurosci., 1, 294-306.). This fact, beside a natural variability among different biological preparations, allows a comparison also among different experimental session giving reliable results and envisaging a definition of a bioelectronic neuronal sensory system.

Burst detection algorithms for the analysis of spatio-temporal patterns in cortical networks of neurons

Cortical neurons extracted from the developing rat central nervous system and put in culture, show, after a few days, spontaneous activity with a typical electrophysiological pattern ranging from stochastic spiking to synchronized bursting. Using microelectrode arrays (MEA), on which dissociated cultures can be grown for long-term measurements, we recorded the electrophysiological activity of cortical networks during development, in order to monitor their responses at different stages of the maturation process. Employing algorithms for detection and analysis of bursts in single-channel spike trains and of synchronized network bursts in multi-channel spike trains, significant changes have been revealed in the firing dynamics at different stages of the developmental process.

Behaviors from an electrically stimulated spinal cord neuronal network cultured on microelectrode arrays

Abstract The spontaneous electrophysiological activity of neural networks seems to play an important role in the Central Nervous System (CNS) developing, subsequent maturation and learning. Learning a new behavior is an exploration process that involves the modulation and the formation of association set between stimuli and responses. Here, we analyze how the electrophysiological activity of cultured spinal cord neurons (14 DIV) from the chick embryo is affected by electrical stimulation. Active neurons show a typical high frequency activity pattern called burst. Induced changes in the patterns of electrophysiological activity are described.

Burst analysis of chemically stimulated spinal cord neuronal networks cultured on microelectrode arrays

Planar microelectrode arrays allow the recording of the electrophysiological activity of cultured neurons for long time periods and from different sites of the network. In the developing Central Nervous System, several neural networks are spontaneously active and show a typical high frequency activity pattern called burst. Such behavior seems to play an important role in subsequent maturation. Here we analyze how the bursting electrophysiological activity of cultured spinal cord neurons (14-18 DIV) from the chick embryo is affected by cyclothiazide (CTZ) which acts on AMPA receptors. Changes in the patterns of electrophysiological activity are described in detail.

Bioartificial Neuronal Networks: Coupling Networks of Biological Neurons to Microtransducer Arrays

In this chapter we introduce the concept of bioartificial neuronal networks, that is networks of biological neurons cultured in-vitro and coupled to Micro Transducer Arrays (MTAs). In-vitro cultured neurons extracted from rats or mice embryos form a bi-dimensional physical model of the brain and, in spite of their simplified level of organization, are an useful framework to study information processing in the nervous system. One of the peculiar aspects is the possibility to chronically (i.e., for several days or weeks) stimulate at and record from multiple sites at the same time, thus establishing a bi-directional interface between a neuronal (i.e. biological) system and an artificial device. This experimental framework can be utilized as a new paradigm for studying novel and advanced neuro-electronic interface.

Modulating neural networks dynamics: multi-site electrical stimulation of in-vitro cortical neurons coupled to MEA devices

Networks of neurons extracted from the developing central nervous system (CNS) are spontaneously active and show a typical electrophysiological pattern called burst. In vitro cultured neurons represent a simplified level of organization where the collective and functional electrophysiological properties emerge and can be experimentally characterized for a better understanding on how brain processes information. Using microelectrode arrays (MEA), on which a cell culture can be grown and kept alive for a long time (from weeks up to months), we recorded the electrophysiological activity of cortical cultures of neurons extracted from an embryonic rat. This activity pattern can be, at some extent, modified by an electrical manipulation, in order to define distinct functional collective states of the network related to specific pattern of stimulation delivered through the network. In order to represent these behaviors from a quantitative point of view, we described the network activity both at burst and at spike level, analyzing the electrophysiological pattern under different stimulation conditions and providing information about network behavior employing custom developed algorithms of burst analysis and standard statistical procedures for spike analysis.