Effects of synaptic transmission on network bursts.

<p>(A) Mean firing rate (MFR) as a function of the decreasing AMPA conductance in AG-networks (i.e., networks with AMPA and GABA conductance, dashed lines) and AGN-networks (i.e., networks with AMPA, GABA and NMDA conductance, solid lines). (B) The reduction in AMPA conductance decreases the mean bursting rate (MBR) and increases the number of asynchronous spikes (random spikes). (C) The reduction in AMPA conductance determines a shift in the ISI distribution, from a multi-peak (0% AMPA reduction) to a single-peak distribution (30% AMPA reduction). (D) The model predicts changes in the activity parameters (MFR, MBR, MFIB, MBD), reproducing recordings under pharmacological blockade of inhibition with bicuculline (BIC, 30 μM, p-values: 0.052, 0.473, 0.189, and 0.449, respectively; independent t-tests, n = 5 simulations, n = 4 recordings). (E) AG-networks show single NBs, while AGN-networks show superbursts, both in recordings (n = 3, gray) and in simulations (n = 10, blue). (F) In AGN-networks, (black, n = 10), the ISI distribution has three peaks: P1 relates to the firing within an NB, P2 to the firing between consecutive NBs, and the peak highlighted by a solid arrow (at approximately 100 ms) to the time intervals across consecutive NBs of a superburst. Blockade of NMDA (AG-network) removes the latter peak from the distribution (red, n = 10). (G) Exemplary simulated trace of the membrane potential of a single cell during a superburst and total conductance of AMPA (black) and NMDA (orange) for the same neuron. Note that during the first NB, the NMDA conductance is negligible compared with that of AMPA.</p

The pre-NB activity shares similar temporal motifs for NBs in the same class and is informative of the following coordinated event.

<p>(A) Illustrative representation of the analysis chain used for the pre-NB temporal motifs, where we considered the following: two NBs (<i>α</i>,<i>β</i>), their pre-NB activity (<i>α</i>_<i>PRE</i>,<i>β</i>_<i>PRE</i>), the NB-graphs (<i>α</i>_<i>G</i>,<i>β</i>_<i>G</i>), the largest connected components (<i>α</i>_<i>L</i>,<i>β</i>_<i>L</i>) and the corresponding number of spikes in the shared temporal motifs (<i>α</i>_<i>M</i>,<i>β</i>_<i>M</i>, M = 12 spikes). (B) All NBs of the same cluster (e.g., cluster ID 0) share common pre-NB temporal motifs (cyan spikes). For instance, the NB occurring at time <i>t</i>₀ shares a temporal motif with the event at time <i>t</i>₁ but differs from the temporal motif shared between NBs occurring at time <i>t</i>₂ and <i>t</i>₃. (C) Normalized similarity matrix among pre-NBs activities (M-motif, with M > 5) for clustered NBs. The clustered NBs share the highest number of motifs (i.e., higher values on the diagonal), indicating that the pre-NB activity is informative of the following NB. (D) Cumulative similarity plot corresponding to the data in panel C but normalized with respect to the cluster size (per-cluster, dashed line) or to the total number of NBs (solid lines). The similarity is consistently higher for the pre-NB activity of NBs belonging to the same cluster (IN) than for the NBs of other clusters (ACROSS). (E) Reordered similarity matrix of pre-NB activity of a simulated network, showing a block structure. Each block is relative to an NB cluster characterized by similar pre-NB spiking patterns. (F) Reordered similarity matrix of recorded pre-NB activity in a cultured network, still shows a block structure.</p

Analysis of the structural connectivity motifs in the functional communities of the network.

<p>Clustering coefficient of structural motifs tested (up to six nodes and isomorphic subgraphs) against the relative abundance of structural motifs in fCOMs and the null-models: rndCOMs (A), rCOMs (B) and sCOMs (C) (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005672#sec009" target="_blank">Materials and Methods</a> section). Red dots mark the structural motifs occurring with a significant frequency difference (p-value<0.05, t-test) for all null-models. Regression lines visualize the trend (where <i>a</i> and <i>p</i> denote the value of the slope and the p-value respectively) among all tested motifs (gray) and the statistically different ones (red). (D) Illustration of overabundant (positive index) and under-represented (negative indexes) structural motifs found in fCOMs with respect to the null models depicted in A-C. As highlighted by the red lines, a high clustering coefficient characterizes the structural motifs that are significantly over-expressed in fCOMs; conversely, significantly under-represented structural motifs resulted in low clustering coefficients (0.56±0.30 vs. 0.19±0.27, respectively).</p

Ready-to-fire state of the functional communities.

<p>The fCOMs of the simulated networks (n = 10 networks simulations) were perturbed with mild-subthreshold stimuli to test their sensitivity in eliciting NBs (see the <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005672#sec009" target="_blank">Materials and Methods</a> section). (A) Raster plot of the activity (black dots) with overlaid electrical stimuli (red dots). As shown, stimuli can either marginally affect the spontaneous activity (B.1) or evoke an NB (B.2). When an NB is elicited, almost all of the stimulated neurons fire (B.3, eNB) a spike within 50 ms. In contrast, when an NB is not elicited, only a small fraction of stimulated neurons fire a spike. In this case, the neuronal activity is not significantly different between the probed regions. (C) Representation of the spatial arrangement of the fCOMs and reference regions that were probed in this example. Neurons are colour coded according to the strength of the stimulus. Stimuli are delivered to fCOMs I, II, III and IV (c.f. <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005672#pcbi.1005672.g004" target="_blank">Fig 4A</a>) and to reference regions R_I, R_II, P_II. (D) The perturbation of the fCOMs evokes NBs with a higher probability than the reference sites.</p

Network regions initiating spontaneous NBs correspond to functional communities.

<p>Spatial maps of the functional communities (fCOMs) and CATs (for clarity, CATs are only depicted up to 50 ms after NB initiation) computed for NBs in simulations (A) and recordings (B), show that the ignition sites (ISs, blue dots) of the NBs and the fCOMs (regions delimited by black solid curves) tend to overlap. A further quantification confirms that the overlap is statistically significant. (C) Fraction of NBs with ISs in a specific fCOM over the total number of NBs, for simulated (black) and experimental (blue) data. (D) Fraction of NBs with ISs located in an area defined by an increasing number of fCOMs (‘area covered’) over the total number of NBs (black: n = 20 simulations, blue: n = 15 recordings, mean value: solid line, standard deviation: shaded area). As shown, NBs originate from <50% of the network area. (E) Boxplots of simulated and experimental areas covered by fCOMs with respect to the total network area, showing that even though not all fCOMs are associated with an IS, the mean total area covered by all fCOMs is <40%.</p

Spontaneous spiking activity recorded with CMOS-MEAs in hippocampal cultures.

<p>(A) High-density CMOS-MEAs simultaneously record extracellular electrical signals from an array of 64 x 64 electrodes covering a 5.12mm x 5.12mm area (three representative traces in black). Coordinated spontaneous spiking activity, or network bursts (NBs), propagates through the network, as indicated by the temporal differences in the spiking activity (black ticks). The three traces are part of the propagating activity shown on the right. (B) Raster plot of 40 s of spontaneous activity. The spike count (blue line, 5-ms time bins) displays a peak in correspondence of the NBs. (C) Raster plots and (D) Centre-of-activity trajectories (CATs, time of the propagation is colour coded) of two consecutive NBs exhibiting different propagations lasting approximately 100 ms. (E) The cross-correlation matrix of NBs shows that events of the same cluster do not occur with a periodicity. (F) Instead, the reordered cross-correlation matrix of NBs shows that the NBs are clustered in a few classes.</p