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Abstract: The subiculum has a strategic position in controlling hippocampal activity and is now receiving much experimental attention. However, information regarding this structure remains fragmented and there are important gaps in our knowledge between what we know about the subicular architecture and its biological function. In recent years a substantial amount of in vitro experimentation has explored many aspects of the functional organization of the subicular microcircuits. Here we review these recent findings. We aim to identify the rules that govern the operation of subicular microcircuits in vitro and to relate these to the role of the subiculum in the intact brain. [Copyright &y& Elsevier]

The subiculum, which has a strategic position in controlling hippocampal activity, is receiving significant attention in epilepsy research. However, the functional organization of subicular circuits remains unknown. Here, we combined different recording and analytical methods to study focal and widespread population activity in the isolated subiculum in zero Mg2+ media. Patch and field recordings were combined to examine the contribution of different cell types to population activity. The properties of cells leading field activity were examined. Predictive factors for a cell to behave as leader included exhibiting the bursting phenotype, displaying a low firing threshold, and having more distal apical dendrites. A subset of bursting cells constituted the first glutamatergic type that led a recruitment process that subsequently activated additional excitatory as well as inhibitory cells. This defined a sequence of synaptic excitation and inhibition that was studied by measuring the associated conductance changes and the evolution of the composite reversal potential. It is shown that inhibition was time-locked to excitation, which shunted excitatory inputs and suppressed firing during focal activity. This was recorded extracellularly as a multi-unit ensemble of active cells, the spatial boundaries of which were controlled by inhibition in contrast to widespread epileptiform activity. Focal activity was not dependent on the preparation or the developmental state because it was also recorded under 5 mM [K+]0 and in adult tissue. Our data indicate that the subicular networks can be spontaneously organized as leader-follower local circuits in which excitation is mainly driven by a subset of bursting cells and inhibition controls spatiotemporal firing. [ABSTRACT FROM AUTHOR]

A high percentage of patients with brain metastases frequently develop neurocognitive symptoms; however, understanding how brain metastasis co-opts the function of neuronal circuits beyond a tumor mass effect remains unknown. We report a comprehensive multidimensional modeling of brain functional analyses in the context of brain metastasis. By testing different preclinical models of brain metastasis from various primary sources and oncogenic profiles, we dissociated the heterogeneous impact on local field potential oscillatory activity from cortical and hippocampal areas that we detected from the homogeneous inter-model tumor size or glial response. In contrast, we report a potential underlying molecular program responsible for impairing neuronal crosstalk by scoring the transcriptomic and mutational profiles in a model-specific manner. Additionally, measurement of various brain activity readouts matched with machine learning strategies confirmed model-specific alterations that could help predict the presence and subtype of metastasis., Competing Interests: Declaration of interests The authors declare no conflicts of interest., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

KCC2 regulates neuronal transmembrane chloride gradients and thereby controls GABA signaling in the brain. KCC2 downregulation is observed in numerous neurological and psychiatric disorders. Paradoxical, excitatory GABA signaling is usually assumed to contribute to abnormal network activity underlying the pathology. We tested this hypothesis and explored the functional impact of chronic KCC2 downregulation in the rat dentate gyrus. Although the reversal potential of GABAA receptor currents is depolarized in KCC2 knockdown neurons, this shift is compensated by depolarization of the resting membrane potential. This reflects downregulation of leak potassium currents. We show KCC2 interacts with Task-3 (KCNK9) channels and is required for their membrane expression. Increased neuronal excitability upon KCC2 suppression altered dentate gyrus rhythmogenesis, which could be normalized by chemogenetic hyperpolarization. Our data reveal KCC2 downregulation engages complex synaptic and cellular alterations beyond GABA signaling that perturb network activity thus offering additional targets for therapeutic intervention., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

The proximodistal axis is considered a major organizational principle of the hippocampus. At the interface between the hippocampus and other brain structures, CA2 apparently breaks this rule. The region is involved in social, temporal, and contextual memory function, but mechanisms remain elusive. Here, we reveal cell-type heterogeneity and a characteristic expression gradient of the transcription factor Sox5 within CA2 in the rat. Using intracellular and extracellular recordings followed by neurochemical identification of single cells, we find marked proximodistal trends of synaptic activity, subthreshold membrane potentials, and phase-locked firing coupled to theta and gamma oscillations. Phase-shifting membrane potentials and opposite proximodistal correlations with theta sinks and sources at different layers support influences from different current generators. CA2 oscillatory activity and place coding of rats running in a linear maze reflect proximodistal state-dependent trends. We suggest that the structure and function of CA2 are distributed along the proximodistal hippocampal axis., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

The brain is a complex system composed of networks of interacting elements, from genes to circuits, whose function (and dysfunction) is not derivable from the superposition of individual components. Epilepsy is frequently described as a network disease, but to date, there is no standardized framework within which network concepts applicable to all levels from genes to whole brain can be used to generate deeper insights into the pathogenesis of seizures or the associated morbidities. To address this shortcoming, the Neurobiology Commission of the International League Against Epilepsy dedicated a Workshop on Neurobiology of Epilepsy (XIV WONOEP 2017) with the aim of formalizing network concepts as they apply to epilepsy and to critically discuss whether and how such concepts could augment current research endeavors. Here, we review concepts and strategies derived by considering epilepsy as a disease of different network hierarchies that range from genes to clinical phenotypes. We propose that the concept of networks is important for understanding epilepsy and is critical for developing new study designs. These approaches could ultimately facilitate the development of novel diagnostic and therapeutic strategies., (Wiley Periodicals, Inc. © 2018 International League Against Epilepsy.)

Memory traces are reactivated selectively during sharp-wave ripples. The mechanisms of selective reactivation, and how degraded reactivation affects memory, are poorly understood. We evaluated hippocampal single-cell activity during physiological and pathological sharp-wave ripples using juxtacellular and intracellular recordings in normal and epileptic rats with different memory abilities. CA1 pyramidal cells participate selectively during physiological events but fired together during epileptic fast ripples. We found that firing selectivity was dominated by an event- and cell-specific synaptic drive, modulated in single cells by changes in the excitatory/inhibitory ratio measured intracellularly. This mechanism collapses during pathological fast ripples to exacerbate and randomize neuronal firing. Acute administration of a use- and cell-type-dependent sodium channel blocker reduced neuronal collapse and randomness and improved recall in epileptic rats. We propose that cell-specific synaptic inputs govern firing selectivity of CA1 pyramidal cells during sharp-wave ripples., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Recent reports in human demonstrate a role of theta-gamma coupling in memory for spatial episodes and a lack of coupling in people experiencing temporal lobe epilepsy, but the mechanisms are unknown. Using multisite silicon probe recordings of epileptic rats engaged in episodic-like object recognition tasks, we sought to evaluate the role of theta-gamma coupling in the absence of epileptiform activities. Our data reveal a specific association between theta-gamma (30-60 Hz) coupling at the proximal stratum radiatum of CA1 and spatial memory deficits. We targeted the microcircuit mechanisms with a novel approach to identify putative interneuronal types in tetrode recordings (parvalbumin basket cells in particular) and validated classification criteria in the epileptic context with neurochemical identification of intracellularly recorded cells. In epileptic rats, putative parvalbumin basket cells fired poorly modulated at the falling theta phase, consistent with weaker inputs from Schaffer collaterals and attenuated gamma oscillations, as evaluated by theta-phase decomposition of current-source density signals. We propose that theta-gamma interneuronal rhythmopathies of the temporal lobe are intimately related to episodic memory dysfunction in this condition.

Pathological high-frequency oscillations (HFOs) (80-800 Hz) are considered biomarkers of epileptogenic tissue, but the underlying complex neuronal events are not well understood. Here, we identify and discuss several outstanding issues or conundrums in regards to the recording, analysis, and interpretation of HFOs in the epileptic brain to critically highlight what is known and what is not about these enigmatic events. High-frequency oscillations reflect a range of neuronal processes contributing to overlapping frequencies from the lower 80 Hz to the very fast spectral frequency bands. Given their complex neuronal nature, HFOs are extremely sensitive to recording conditions and analytical approaches. We provide a list of recommendations that could help to obtain comparable HFO signals in clinical and basic epilepsy research. Adopting basic standards will facilitate data sharing and interpretation that collectively will aid in understanding the role of HFOs in health and disease for translational purpose.

Developmental cortical malformations comprise a large spectrum of histopathological brain abnormalities and syndromes. Their genetic, developmental and clinical complexity suggests they should be better understood in terms of the complementary action of independently timed perturbations (i.e., the multiple-hit hypothesis). However, understanding the underlying biological processes remains puzzling. Here we induced developmental cortical malformations in offspring, after intraventricular injection of methylazoxymethanol (MAM) in utero in mice. We combined extensive histological and electrophysiological studies to characterize the model. We found that MAM injections at E14 and E15 induced a range of cortical and hippocampal malformations resembling histological alterations of specific genetic mutations and transplacental mitotoxic agent injections. However, in contrast to most of these models, intraventricularly MAM-injected mice remained asymptomatic and showed no clear epilepsy-related phenotype as tested in long-term chronic recordings and with pharmacological manipulations. Instead, they exhibited a non-specific reduction of hippocampal-related brain oscillations (mostly in CA1); including theta, gamma and HFOs; and enhanced thalamocortical spindle activity during non-REM sleep. These data suggest that developmental cortical malformations do not necessarily correlate with epileptiform activity. We propose that the intraventricular in utero MAM approach exhibiting a range of rhythmopathies is a suitable model for multiple-hit studies of associated neurological disorders.

Hippocampal high-frequency oscillations (HFOs) are prominent in physiological and pathological conditions. During physiological ripples (100-200 Hz), few pyramidal cells fire together coordinated by rhythmic inhibitory potentials. In the epileptic hippocampus, fast ripples (>200 Hz) reflect population spikes (PSs) from clusters of bursting cells, but HFOs in the ripple and the fast ripple range are vastly intermixed. What is the meaning of this frequency range? What determines the expression of different HFOs? Here, we used different concentrations of Ca(2+) in a physiological range (1-3 mM) to record local field potentials and single cells in hippocampal slices from normal rats. Surprisingly, we found that this sole manipulation results in the emergence of two forms of HFOs reminiscent of ripples and fast ripples recorded in vivo from normal and epileptic rats, respectively. We scrutinized the cellular correlates and mechanisms underlying the emergence of these two forms of HFOs by combining multisite, single-cell and paired-cell recordings in slices prepared from a rat reporter line that facilitates identification of GABAergic cells. We found a major effect of extracellular Ca(2+) in modulating intrinsic excitability and disynaptic inhibition, two critical factors shaping network dynamics. Moreover, locally modulating the extracellular Ca(2+) concentration in an in vivo environment had a similar effect on disynaptic inhibition, pyramidal cell excitability, and ripple dynamics. Therefore, the HFO frequency band reflects a range of firing dynamics of hippocampal networks.

While novel influential concepts in neuroscience bring the focus to local activities generated within a few tens of cubic micrometers in the brain, we are still devoid of appropriate tools to record and manipulate pharmacologically neuronal activity at this fine scale. Here we designed, fabricated and encapsulated microprobes for simultaneous depth recording and drug delivery using exclusively the polymer SU-8 as structural material. A tetrode- and linear-like electrode patterning was combined for the first time with single and double fluidic microchannels for independent drug delivery. The device was tested experimentally using the in vivo anesthetized rat preparation. Both probe types successfully recorded detailed spatiotemporal features of local field potentials and single-cell activity at a resolution never attained before with integrated fluidic probes. Drug delivery was achieved with high spatial and temporal precision in a range from tens of nanoliters to a few microliters, as confirmed histologically. These technological advancements will foster a wide range of neural applications aimed at simultaneous monitoring of brain activity and delivery at a very precise micrometer scale.

High frequency oscillations (HFO) have a variety of characteristics: band-limited or broad-band, transient burst-like phenomenon or steady-state. HFOs may be encountered under physiological or under pathological conditions (pHFO). Here we review the underlying mechanisms of oscillations, at the level of cells and networks, investigated in a variety of experimental in vitro and in vivo models. Diverse mechanisms are described, from intrinsic membrane oscillations to network processes involving different types of synaptic interactions, gap junctions and ephaptic coupling. HFOs with similar frequency ranges can differ considerably in their physiological mechanisms. The fact that in most cases the combination of intrinsic neuronal membrane oscillations and synaptic circuits are necessary to sustain network oscillations is emphasized. Evidence for pathological HFOs, particularly fast ripples, in experimental models of epilepsy and in human epileptic patients is scrutinized. The underlying mechanisms of fast ripples are examined both in the light of animal observations, in vivo and in vitro, and in epileptic patients, with emphasis on single cell dynamics. Experimental observations and computational modeling have led to hypotheses for these mechanisms, several of which are considered here, namely the role of out-of-phase firing in neuronal clusters, the importance of strong excitatory AMPA-synaptic currents and recurrent inhibitory connectivity in combination with the fast time scales of IPSPs, ephaptic coupling and the contribution of interneuronal coupling through gap junctions. The statistical behaviour of fast ripple events can provide useful information on the underlying mechanism and can help to further improve classification of the diverse forms of HFOs., (Copyright © 2012. Published by Elsevier Ltd.)

Here, we describe new fabrication methods aimed to integrate planar tetrode-like electrodes into a polymer SU-8 based microprobe for neuronal recording applications. New concepts on the fabrication sequences are introduced in order to eliminate the typical electrode-tissue gap associated to the passivation layer. Optimization of the photolithography technique and high step coverage of the sputtering process have been critical steps in this new fabrication process. Impedance characterization confirmed the viability of the electrodes for reliable neuronal recordings with values comparable to commercial probes. Furthermore, a homogeneous sensing behavior was obtained in all the electrodes of each probe. Finally, in vivo action potential and local field potential recordings were successfully obtained from the rat dorsal hippocampus. Peak-to-peak amplitude of action potentials ranged from noise level to up to 400-500 μV. Moreover, action potentials of different amplitudes and shapes were recorded from all the four recording sites, suggesting improved capability of the tetrode to distinguish from different neuronal sources., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Affective symptoms such as anxiety and depression are frequently observed in patients with epilepsy. The mechanisms of comorbidity of epilepsy and affective disorders, however, remain unclear. Diverse models are traditionally used in epilepsy research, including the status epilepticus (SE) model in rats, which are aimed at generating chronic epileptic animals; however, the implications of different SE models and rat strains in emotional behaviors has not been reported. To address this issue, we examined the emotional sequelae of two SE models of temporal lobe epilepsy (TLE)--the lithium-pilocarpine (LIP) model and the kainic acid (KA) model--in two different rat strains (Wistar and Sprague-Dawley), which differ significantly in the pattern and extent of TLE-associated brain lesions. We found differences between LIP- and KA-treated animals in tests for depression-like and anxiety-like behaviors, as well as differences in plasma corticosterone levels. Whereas only LIP-treated rats displayed increased motivation to consume saccharin, both SE models led to reduced motivation for social contact, with LIP-treated animals being particularly affected. Evaluation of behavior in the open field test indicated very low levels of anxiety in LIP-treated rats and a mild decrease in KA-treated rats compared to controls. After exposure to a battery of behavioral tests, plasma corticosterone levels were increased only in LIP-treated animals. This hyperactivity in the hypothalamus-pituitary-adrenocortical (HPA) axis was highly correlated with performance in the open field test and the social interaction test, suggesting that comorbidity of epilepsy and emotional behaviors might also be related to other factors such as HPA axis function. Our results indicate that altered emotional behaviors are not inherent to the epileptic condition in experimental TLE; instead, they likely reflect alterations in anxiety levels related to model-dependent dysregulation of the HPA axis.

A major goal in epilepsy research is to understand the cellular basis of pathological forms of network oscillations, particularly those classified as high-frequency activity. What are the underlying mechanisms, and how do they arise? The topic of this review is the pattern of high-frequency oscillations that have been recorded in epileptic tissue, and how they might differ from physiological activity. We discuss recent experimental and clinical data with a major focus on the diverse sources of extracellular signals and the contribution of different neuronal populations, including GABAergic interneurons and glutamatergic principal cells., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The mechanisms involved in the transition to an epileptic seizure remain unclear. To examine them, we used tissue slices from human subjects with mesial temporal lobe epilepsies. Ictal-like discharges were induced in the subiculum by increasing excitability along with alkalinization or low Mg(2+). During the transition, distinct pre-ictal discharges emerged concurrently with interictal events. Intracranial recordings from the mesial temporal cortex of subjects with epilepsy revealed that similar discharges before seizures were restricted to seizure onset sites. In vitro, pre-ictal events spread faster and had larger amplitudes than interictal discharges and had a distinct initiation site. These events depended on glutamatergic mechanisms and were preceded by pyramidal cell firing, whereas interneuron firing preceded interictal events that depended on both glutamatergic and depolarizing GABAergic transmission. Once established, recurrence of these pre-ictal discharges triggered seizures. Thus, the subiculum supports seizure generation, and the transition to seizure involves an emergent glutamatergic population activity.

Cognitive impairment is a major concern in temporal lobe epilepsy (TLE). While different experimental models have been used to characterize TLE-related cognitive deficits, little is known on whether a particular deficit is more associated with the underlying brain injuries than with the epileptic condition per se. Here, we look at the relationship between the pattern of brain damage and spatial memory deficits in two chronic models of TLE (lithium-pilocarpine, LIP and kainic acid, KA) from two different rat strains (Wistar and Sprague-Dawley) using the Morris water maze and the elevated plus maze in combination with MRI imaging and post-morten neuronal immunostaining. We found fundamental differences between LIP- and KA-treated epileptic rats regarding spatial memory deficits and anxiety. LIP-treated animals from both strains showed significant impairment in the acquisition and retention of spatial memory, and were unable to learn a cued version of the task. In contrast, KA-treated rats were differently affected. Sprague-Dawley KA-treated rats learned less efficiently than Wistar KA-treated animals, which performed similar to control rats in the acquisition and in a probe trial testing for spatial memory. Different anxiety levels and the extension of brain lesions affecting the hippocampus and the amydgala concur with spatial memory deficits observed in epileptic rats. Hence, our results suggest that hippocampal-dependent spatial memory is not necessarily affected in TLE and that comorbidity between spatial deficits and anxiety is more related with the underlying brain lesions than with the epileptic condition per se.

Objective: Drug-resistant temporal lobe epilepsy (TLE) can be treated by tailored surgery guided by electrocorticography (ECoG). Although its value is still controversial, ECoG activity can provide continuous information on intracortical interactions that may be useful to understand the pathophysiology of TLE. The goal of this study is to characterize local interactions in multichannel ECoG recordings of the lateral cortex of TLE patients using three synchronization measures and to link this information with surgical outcome., Methods: Intraoperative ECoG recordings from 29 TLE patients were obtained using grids of 20 electrodes (4 x 5) covering regions T1, T2, and T3 of the lateral temporal lobe. Linear correlation, mutual information, and phase synchronization were calculated to quantify lateral intracortical interactions. Surrogate data files were generated to test results statistically., Results: By distributing locally the interactions between the electrodes, we characterized the spatial patterns of ECoG activity. We found clusters of synchronized activity at specific areas of the lateral temporal cortex in most patients. Methodologically, linear correlation and phase synchronization performed better than mutual information for cluster discrimination. ROC analysis suggested that surgical removal of sharply defined synchronization clusters correlated with seizure control., Conclusions: Our results show that synchronous intraoperative ECoG activity emerges from specific cortical areas that are highly differentiated from the rest of the temporal cortex. This suggests that synchronization analysis could be used to functionally map into the temporal cortex of TLE patients. Moreover, our results suggest that these sites might be involved in the circuits that participate in clinical seizures.

Ripples are sharp-wave-associated field oscillations (100-300 Hz) recorded in the hippocampus during behavioral immobility and slow-wave sleep. In epileptic rats and humans, a different and faster oscillation (200-600 Hz), termed fast ripples, has been described. However, the basic mechanisms are unknown. Here, we propose that fast ripples emerge from a disorganized ripple pattern caused by unreliable firing in the epileptic hippocampus. Enhanced synaptic activity is responsible for the irregular bursting of CA3 pyramidal cells due to large membrane potential fluctuations. Lower field interactions and a reduced spike-timing reliability concur with decreased spatial synchronization and the emergence of fast ripples. Reducing synaptically driven membrane potential fluctuations improves both spike-timing reliability and spatial synchronization and restores ripples in the epileptic hippocampus. Conversely, a lower spike-timing reliability, with reduced potassium currents, is associated with ripple shuffling in normal hippocampus. Therefore, fast ripples may reflect a pathological desynchronization of the normal ripple pattern.

The subiculum, which provides the major hippocampal output, contains different cell types including weak/strong bursting and regular-spiking cells, and fast-spiking interneurons. These cellular populations play different roles in the generation of physiological rhythms and epileptiform activity. However, their intrinsic connectivity and the synaptic regulation of their discharge patterns remain unknown. In the present study, the local synaptic responses of subicular cell types were examined in vitro. To this purpose, slices were prepared at a specific orientation that permitted the antidromic activation of projection cells as a tool to examine local circuits. Patch recordings in cell-attached and whole-cell configurations were combined with neurobiotin labelling to classify cell types. Strong (approximately 75 %), but not weak (approximately 22 %), bursting cells typically fired bursts in response to local synaptic excitation, whereas the majority of regular-spiking cells (approximately 87 %) remained silent. Local excitation evoked single spikes in more than 70 % of fast-spiking interneurons. This different responsiveness was determined by intrinsic membrane properties and not by the amplitude and pharmacology of synaptic currents. Inhibitory GABAergic responses were also detected in some cells, typically as a component of an excitatory/inhibitory sequence. A positive correlation between the latency of the excitatory and inhibitory responses, together with the glutamatergic control (via non-NMDA receptors) of inhibition, suggested a local mechanism. The effect of local inhibition on synaptically activated firing of different cell types was evaluated. It is shown that projection bursting cells of the subiculum are strongly controlled by local inhibitory circuits.

We combined visual-assisted whole-cell recordings with Lucifer Yellow labelling to record in vitro from interneuron-like cells from human epileptic neocortical tissue that exhibited spiking activity in situ. Information from intraoperative electrocorticography, which was performed to tailor resection, were used to select those areas that participate in interictal spiking. Whole-cell recordings from slices prepared from these areas showed that both pyramidal and interneuron-like cells were present and retained their main intrinsic firing patterns, i.e. regular spiking and fast spiking, respectively. Moreover, non-pyramidal interneuron-like cells remained innervated by excitatory inputs as both spontaneous and evoked non-NMDA mediated excitatory postsynaptic potentials were recorded. We conclude that putative inhibitory GABAergic cells are present and functional in the human epileptic tissue.

Rhythmic spontaneous bursting is a fundamental hallmark of the immature hippocampal activity recorded in vitro. These bursts or giant depolarizing potentials (GDPs) are GABA- and glutamatergic-driven events. The mechanisms of GDPs generation are still controversial, since although a hilar origin has been suggested, GDPs were also recorded from isolated CA3 area. Here, we have investigated the origin of GDPs in hippocampal slices from newborn rabbits. Simultaneous intracellular recordings were performed in CA3, CA1 and the fascia dentata. We found a high degree of correlation between the spontaneous GDPs present in CA3 and CA1 regions. Cross-correlation analysis demonstrated that CA3 firing precedes CA1 by about 192 ms, although a significant population of discharges was recorded first in CA1 (20%). Granule cells (GCs) in the fascia dentata also showed GDPs. The frequency of these events (1.46 +/- 1.25 GDPs/min, n = 7) is significantly lower when compared with that from CA3 (3.13 +/- 1.43 GDPs/min, n = 10) or CA1 (2.94 +/- 1.36 GDPs/min, n = 17). Dual recordings from CA3 and fascia dentata cells showed synchronous bursts in both regions with no prevalent preceding area. By recording from isolated areas we found that CA1, CA3 and the fascia dentata can produce GDPs, suggesting that they emerge as a property of local circuits present throughout the hippocampus.

We have analyzed the postnatal evolution of the spontaneous electrical activity in pyramidal neurons from rabbit hippocampal slices. The firing mode of CA1 neurons changes from bursting to regular spiking along the first postnatal month. Interspike intervals (ISIs) were used to account for the dynamical structure of the firing behavior. Histograms and joint interval scattergrams show that the firing mode from (P0-P7) cells has a different distribution from that obtained in (P15-P25) neurons. We have used a mathematical measure called the product of inertia to quantify this difference. Our findings demonstrate that the spontaneous activity changes along the maturational process.