Microelectrode array recordings of cultured hippocampal networks reveal a simple model for transcription and protein synthesis‐dependent plasticity

Abstract

A simplified cell culture system was developed to study neuronal plasticity. As changes in synaptic strength may alter network activity patterns, we grew hippocampal neurones on a microelectrode array (MEA) and monitored their collective behaviour with 60 electrodes simultaneously. We found that exposure of the network for 15 min to the GABA A receptor antagonist bicuculline induced an increase in synaptic efficacy at excitatory synapses that was associated with an increase in the frequency of miniature AMPA receptor-mediated EPSCs and a change in network activity from uncoordinated firing of neurones (lacking any recognizable pattern) to a highly organized, periodic and synchronous burst pattern. Induction of recurrent synchronous bursting was dependent on NMDA receptor activation and required extracellular signal-regulated kinase (ERK)1/2 signalling and translation of pre-existing mRNAs. Once induced, the burst pattern persisted for several days; its maintenance phase (> 4 h) was dependent on gene transcription taking place in a critical period of 120 min following induction. Thus, cultured hippocampal neurones display a simple, transcription and protein synthesis-dependent form of plasticity. The non-invasive nature of MEA recordings provides a significant advantage over traditional assays for synaptic connectivity (i.e. long-term potentiation in brain slices) and facilitates the search for activity-regulated genes critical for late-phase plasticity.

Nuclear Calcium Signaling Controls Methyl-CpG-binding Protein 2 (MeCP2) Phosphorylation on Serine 421 following Synaptic Activity

“…1C). To induce AP bursting, we exposed the cultures to the GABA A receptor antagonist bicuculline; this led to the removal of tonic inhibition from the network, giving rise to periodically occurring AP bursts, each of which is associated with an increase in the cytoplasmic and nuclear calcium concentration (31,32) (Fig. 1C).…”

Synaptic Activity Induces Dramatic Changes in the Geometry of the Cell Nucleus: Interplay between Nuclear Structure, Histone H3 Phosphorylation, and Nuclear Calcium Signaling

“…Cultures were plated onto 12 mm glass coverslips at a density of ϳ450 cells (neurons plus glial cells) per mm 2 . Stimulations were done after a culturing period of 9 -10 d during which hippocampal neurons develop a rich network of processes, express functional NMDA-type and AMPA/kainate-type glutamate receptors, and form synaptic contacts (Bading et al, 1995;Arnold et al, 2005). NMDA, D(Ϫ)-APV, MK-801, anisomycin and cycloheximide were from Sigma; bicuculline was from MP Biomedicals; actinomycin D was from AppliChem; tetrodotoxin (TTX) was from Tocris Bioscience; PD98059 and 4-aminopyridine were from Calbiochem (Merck); UO126 was from Axxora; DiD was from Biotrend Chemikalien.…”

“…Synaptic NMDA receptors were stimulated by exposing hippocampal neurons to the GABA A receptor antagonist bicuculline. Because the hippocampal network contains ϳ11% GABA-ergic interneurons that impose a tonic inhibition onto the network, removal of GABA A receptor activity leads to bursts of AP firing (Hardingham et al, 2001b;Arnold et al, 2005). AP bursting causes calcium entry through synaptic NMDA receptors, generates robust calcium transients in the cytosol and nucleus, activates gene transcription, and promotes neuronal plasticity and survival (Hardingham et al, 2002;Hardingham and Bading, 2003;Arnold et al, 2005;Lee et al, 2005;Papadia et al, 2005).…”

“…Dynamics of nuclear calcium signaling in differently sized and nearly equally sized compartments of infolded nuclei. For calcium imaging experiments, cultured hippocampal neurons were exposed to the GABA A receptor blocker bicuculline (50 M); this treatment leads to bursts of AP firing and generates periodic calcium transients in the cytosol and in the nucleus (Hardingham et al, 2001a;Arnold et al, 2005). Image acquisition was done at a rate of 1.5 Hz with a confocal microscope.…”

Network activity-independent coordinated gene expression program for synapse assembly

“…Primary neuronal cultures from an embryonic hippocampus have been extensively used as a model of synaptogenesis, in which cells from different stages of development become resynchronized after dissociation (22), and electrical activity is dependent on the connectivity of the neurons (i.e., network activity), rather than the properties of individual cells (23). Multielectrode arrays provide an efficient, spatially comprehensive, and noninvasive method of recording the development of network spiking activity over time, which has been applied successfully in hippocampal cultures (24). Gene expression arrays can determine the simultaneous profiling of thousands of transcripts.…”