Cell-Type Specific Inhibition Controls the High-Frequency Oscillations in the Medial Entorhinal Cortex.

The medial entorhinal cortex (mEC) plays a critical role for spatial navigation and memory. While many studies have investigated the principal neurons within the entorhinal cortex, much less is known about the inhibitory circuitries within this structure. Here, we describe for the first time in the mEC a subset of parvalbumin-positive (PV+) interneurons (INs)-stuttering cells (STUT)-with morphological, intrinsic electrophysiological, and synaptic properties distinct from fast-spiking PV+ INs. In contrast to the fast-spiking PV+ INs, the axon of the STUT INs also terminated in layer 3 and showed subthreshold membrane oscillations at gamma frequencies. Whereas the synaptic output of the STUT INs was only weakly reduced by a µ-opioid agonist, their inhibitory inputs were strongly suppressed. Given these properties, STUT are ideally suited to entrain gamma activity in the pyramidal cell population of the mEC. We propose that activation of the µ-opioid receptors decreases the GABA release from the PV+ INs onto the STUT, resulting in disinhibition of the STUT cell population and the consequent increase in network gamma power. We therefore suggest that the opioid system plays a critical role, mediated by STUT INs, in the neural signaling and oscillatory network activity within the mEC.

Outlines to Initiate Epilepsy Surgery in Low- and Middle-Income Countries.

Surgery is an essential treatment option for patients with drug-resistant epilepsy. While most epilepsy patients worldwide live in low- and middle-income countries (LMIC), most of these countries do not have epilepsy surgery, and those that do have surgical epilepsy services lack capacities. The rapidly growing population in LMIC further widens the gap between the number of patients who can potentially benefit from surgery and those who can actually receive it. This makes the initiation of new surgical epilepsy centers in those countries an urgent issue. Epilepsy surgery is feasible in LMIC, even in resource-poor settings, but lack of local expertise is a major obstacle to the introduction of new surgical services. Importantly, expertise deficits can be compensated by collaborating with a well-established epilepsy center for knowledge transfer, skill building and mentoring. Such projects need to be organized in a multidisciplinary team, should focus on the given circumstances, and should use technologies and personnel that are reasonably available and can function sustainably. Local cultural factors and improvement of patients' quality of life are further spotlights reflected by an increasing number of studies. As a general outline for a new surgical epilepsy program in LMIC, it is recommended to initially focus on patients with mesial temporal lobe epilepsy due to hippocampal sclerosis or other well defined pathologies. These constellations have an excellent surgical outcome in terms of seizure control, can be diagnosed by non-invasive methods, and can be reliably identified even under low-resource conditions. Moreover, surgery can be performed with a highly standardized approach and at reasonable costs, and the vast majority of patients will benefit from surgical intervention. The range of services can then be gradually expanded, depending on growing expertise, local needs, prospects and constraints. Although the introduction of surgical epilepsy services in LMIC can face several challenges, none of them should be a permanent barrier for further establishments.

Initiating a new national epilepsy surgery program: Experiences gathered in Georgia.

Surgery is the most effective therapeutic approach for medically refractory epilepsies and a safe and cost-efficient treatment in terms of long-term expenses of direct, indirect, and intangible costs. Georgia is a Caucasian low- to middle-income country with a remarkable effort to deal with epileptic diseases, but without an appropriate epilepsy surgery program. To address the needs for such a service in this country, two joint German-Georgian projects were initiated in 2017 and 2019. In the framework of these projects, a productive exchange program involving German and Georgian experts was undertaken in the past two years. This program included training and mentoring for Georgian clinical colleagues, as well as joint case conferences and workshops with the aim of optimizing presurgical diagnostics and preparing for an epilepsy surgery program in Georgia. Finally, a postsurgical medium- and long-term follow-up scheme was organized as the third component of this comprehensive approach. As a result of our efforts, the first patients underwent anterior temporal lobectomy and all of them remain seizure-free up to the present day. Hence, epilepsy surgery is not only feasible, but also already available in Georgia. In this report, we aim to share our experiences in the initiation and implementation of surgical epilepsy intervention in Georgia and illustrate our recent endeavor and achievements.

Cell-specific synaptic plasticity induced by network oscillations.

Gamma rhythms are known to contribute to the process of memory encoding. However, little is known about the underlying mechanisms at the molecular, cellular and network levels. Using local field potential recording in awake behaving mice and concomitant field potential and whole-cell recordings in slice preparations we found that gamma rhythms lead to activity-dependent modification of hippocampal networks, including alterations in sharp wave-ripple complexes. Network plasticity, expressed as long-lasting increases in sharp wave-associated synaptic currents, exhibits enhanced excitatory synaptic strength in pyramidal cells that is induced postsynaptically and depends on metabotropic glutamate receptor-5 activation. In sharp contrast, alteration of inhibitory synaptic strength is independent of postsynaptic activation and less pronounced. Further, we found a cell type-specific, directionally biased synaptic plasticity of two major types of GABAergic cells, parvalbumin- and cholecystokinin-expressing interneurons. Thus, we propose that gamma frequency oscillations represent a network state that introduces long-lasting synaptic plasticity in a cell-specific manner.

Cell type-specific separation of subicular principal neurons during network activities.

The hippocampal output structure, the subiculum, expresses two major memory relevant network rhythms, sharp wave ripple and gamma frequency oscillations. To this date, it remains unclear how the two distinct types of subicular principal cells, intrinsically bursting and regular spiking neurons, participate in these two network rhythms. Using concomitant local field potential and intracellular recordings in an in vitro mouse model that allows the investigation of both network rhythms, we found a cell type-specific segregation of principal neurons into participating intrinsically bursting and non-participating regular spiking cells. However, if regular spiking cells were kept at a more depolarized level, they did participate in a specific manner, suggesting a potential bimodal working model dependent on the level of excitation. Furthermore, intrinsically bursting and regular spiking cells exhibited divergent intrinsic membrane and synaptic properties in the active network. Thus, our results suggest a cell-type-specific segregation of principal cells into two separate groups during network activities, supporting the idea of two parallel streams of information processing within the subiculum.

Changes in neural network homeostasis trigger neuropsychiatric symptoms.

The mechanisms that regulate the strength of synaptic transmission and intrinsic neuronal excitability are well characterized; however, the mechanisms that promote disease-causing neural network dysfunction are poorly defined. We generated mice with targeted neuron type-specific expression of a gain-of-function variant of the neurotransmitter receptor for glycine (GlyR) that is found in hippocampectomies from patients with temporal lobe epilepsy. In this mouse model, targeted expression of gain-of-function GlyR in terminals of glutamatergic cells or in parvalbumin-positive interneurons persistently altered neural network excitability. The increased network excitability associated with gain-of-function GlyR expression in glutamatergic neurons resulted in recurrent epileptiform discharge, which provoked cognitive dysfunction and memory deficits without affecting bidirectional synaptic plasticity. In contrast, decreased network excitability due to gain-of-function GlyR expression in parvalbumin-positive interneurons resulted in an anxiety phenotype, but did not affect cognitive performance or discriminative associative memory. Our animal model unveils neuron type-specific effects on cognition, formation of discriminative associative memory, and emotional behavior in vivo. Furthermore, our data identify a presynaptic disease-causing molecular mechanism that impairs homeostatic regulation of neural network excitability and triggers neuropsychiatric symptoms.

GABA(B) autoreceptor-mediated cell type-specific reduction of inhibition in epileptic mice.

GABA(B) receptors (GABA(B)Rs) mediate slow inhibitory effects on neuronal excitability and synaptic transmission in the brain. However, the GABA(B)R agonist baclofen can also promote excitability and seizure generation in human patients and animals models. Here we show that baclofen has concentration-dependent effects on the hippocampal network in a mouse model of mesial temporal lobe epilepsy. Application of baclofen at a high dose (10 mg/kg i.p.) reduced the power of γ oscillations and the frequency of pathological discharges in the Cornu Ammonis area 3 (CA3) area of freely moving epileptic mice. Unexpectedly, at a lower dose (1 mg/kg), baclofen markedly increased γ activity accompanied by a higher incidence of pathological discharges. Intracellular recordings from CA3 pyramidal cells in vitro further revealed that, although at a high concentration (10 µM), baclofen invariably resulted in hyperpolarization, at low concentrations (0.5 µM), the drug had divergent effects, producing depolarization and an increase in firing frequency in epileptic but not control mice. These excitatory effects were mediated by the selective muting of inhibitory cholecystokinin-positive basket cells (CCK(+) BCs), through enhanced inhibition of GABA release via presynaptic GABA(B)Rs. We conclude that cell type-specific up-regulation of GABA(B)R-mediated autoinhibition in CCK(+) BCs promotes aberrant high frequency oscillations and hyperexcitability in hippocampal networks of chronic epileptic mice.

Segregation of axonal and somatic activity during fast network oscillations.

In central neurons, information flows from the dendritic surface toward the axon terminals. We found that during in vitro gamma oscillations, ectopic action potentials are generated at high frequency in the distal axon of pyramidal cells (PCs) but do not invade the soma. At the same time, axo-axonic cells (AACs) discharged at a high rate and tonically inhibited the axon initial segment, which can be instrumental in preventing ectopic action potential back-propagation. We found that activation of a single AAC substantially lowered soma invasion by antidromic action potential in postsynaptic PCs. In contrast, activation of soma-inhibiting basket cells had no significant impact. These results demonstrate that AACs can separate axonal from somatic activity and maintain the functional polarization of cortical PCs during network oscillations.

Proper synaptic vesicle formation and neuronal network activity critically rely on syndapin I.

Synaptic transmission relies on effective and accurate compensatory endocytosis. F-BAR proteins may serve as membrane curvature sensors and/or inducers and thereby support membrane remodelling processes; yet, their in vivo functions urgently await disclosure. We demonstrate that the F-BAR protein syndapin I is crucial for proper brain function. Syndapin I knockout (KO) mice suffer from seizures, a phenotype consistent with excessive hippocampal network activity. Loss of syndapin I causes defects in presynaptic membrane trafficking processes, which are especially evident under high-capacity retrieval conditions, accumulation of endocytic intermediates, loss of synaptic vesicle (SV) size control, impaired activity-dependent SV retrieval and defective synaptic activity. Detailed molecular analyses demonstrate that syndapin I plays an important role in the recruitment of all dynamin isoforms, central players in vesicle fission reactions, to the membrane. Consistently, syndapin I KO mice share phenotypes with dynamin I KO mice, whereas their seizure phenotype is very reminiscent of fitful mice expressing a mutant dynamin. Thus, syndapin I acts as pivotal membrane anchoring factor for dynamins during regeneration of SVs.

Cell-type-specific modulation of feedback inhibition by serotonin in the hippocampus.

Midbrain raphe nuclei provide strong serotonergic projections to the hippocampus, in which serotonin (5-HT) exerts differential effects mediated by multiple 5-HT receptor subtypes. The functional relevance of this diversity of information processing is poorly understood. Here we show that serotonin via 5-HT(1B) heteroreceptors substantially reduces synaptic excitation of cholecystokinin-expressing interneurons in area CA1 of the rat hippocampus, in contrast to parvalbumin-expressing basket cells. The reduction is input specific, affecting only glutamatergic synaptic transmission originating from CA1 pyramidal cells. As a result, serotonin selectively decreases feedback inhibition via 5-HT(1B) receptor activation and subsequently increases the integration time window for spike generation in CA1 pyramidal cells. Our data imply an important role for serotonergic modulation of GABAergic action in subcortical control of hippocampal output.

Low-frequency stimulation of the temporoammonic pathway induces heterosynaptic disinhibition in the subiculum.

The subiculum (Sub) is the principal target of CA1 pyramidal cells. It serves as the final relay of hippocampal output and thus mediates hippocampal-cortical interaction. In addition, the Sub receives direct input from the entorhinal cortex via the temporoammonic pathway. In this study, we demonstrate that low-frequency stimulation of the temporoammonic pathway results in the disinhibition of excitatory synaptic transmission at CA1-Sub synapses. We provide evidence that this disinhibition is mediated by an NMDA receptor-dependent long-term depression (LTD) of GABAergic inhibition. This mechanism might bear physiological significance for the stabilization and processing of mnemonic information at hippocampal output synapses and underpins the functional role of hippocampal-entorhinal interaction in memory formation.

Altered Excitatory-Inhibitory Balance in the NMDA-Hypofunction Model of Schizophrenia.

Schizophrenia is a common psychiatric disorder of high incidence, affecting approximately 1% of the world population. The essential neurotransmitter pathology of schizophrenia remains poorly defined, despite huge advances over the past half-century in identifying neurochemical and pathological abnormalities in the disease. The dopamine/serotonin hypothesis has originally provided much of the momentum for neurochemical research in schizophrenia. In recent years, the attention has, however, shifted to the glutamate system, the major excitatory neurotransmitter in the CNS and towards a concept of functional imbalance between excitatory and inhibitory transmission at the network level in various brain regions in schizophrenia. The evidence indicating a central role for the NMDA-receptor subtype in the aetiology of schizophrenia has led to the NMDA-hypofunction model of this disease and the use of phencyclidines as a means to induce the NMDA-hypofunction state in animal models. The purpose of this review is to discuss recent findings highlighting the importance of the NMDA-hypofunction model of schizophrenia, both from a clinical perspective, as well as in opening a line of research, which enables electrophysiological studies at the cellular and network level in vitro. In particular, changes in excitation-inhibition (E/I) balance in the NMDA-hypofunction model of the disease and the resulting changes in network behaviours, particularly in gamma frequency oscillatory activity, will be discussed.

Glycinergic tonic inhibition of hippocampal neurons with depolarizing GABAergic transmission elicits histopathological signs of temporal lobe epilepsy.

An increasing number of epilepsy patients are afflicted with drug-resistant temporal lobe epilepsy (TLE) and require alternative therapeutic approaches. High-affinity glycine receptors (haGlyRs) are functionally adapted to tonic inhibition due to their response to hippocampal ambient glycine, and their synthesis is activity-dependent. Therefore, in our study, we scanned TLE hippocampectomies for expression of haGlyRs and characterized the effects mediated by these receptors using primary hippocampal neurons. Increased haGlyR expression occurred in TLE hippocampi obtained from patients with a severe course of disease. Furthermore, in TLE patients, haGlyR and potassium chloride cotransporter 2 (KCC2) expressions were inversely regulated. To examine this potential causal relationship with respect to TLE histopathology, we established a hippocampal cell culture system utilising tonic inhibition mediated by haGlyRs in response to hippocam-pal ambient glycine and in the context of a high Cl equilibrium potential, as is the case in TLE hippocampal neurons. We showed that hypoactive neurons increase their ratio between glutamatergic and GABAergic synapses, reduce their dendrite length and finally undergo excitotoxicity. Pharmacological dissection of the underlying processes revealed ionotropic glutamate and TrkB receptors as critical mediators between neuronal hypoactivity and the emergence of these TLE-characteristic histopathological signs. Moreover, our results indicate a beneficial role for KCC2, because decreasing the Cl- equilibrium potential by KCC2 expression also rescued hypoactive hippocampal neurons. Thus, our data support a causal relationship between increased haGlyR expression and the emergence of histopathological TLE-characteristic signs, and they establish a pathophysiological role for neuronal hypoactivity in the context of a high Cl- equilibrium potential.

Impaired hippocampal rhythmogenesis in a mouse model of mesial temporal lobe epilepsy.

Mesial temporal lobe epilepsy (mTLE) is one of the most common forms of epilepsy, characterized by hippocampal sclerosis and memory deficits. Injection of kainic acid (KA) into the dorsal hippocampus of mice reproduces major electrophysiological and histopathological characteristics of mTLE. In extracellular recordings from the morphologically intact ventral hippocampus of KA-injected epileptic mice, we found that theta-frequency oscillations were abolished, whereas gamma oscillations persisted both in vivo and in vitro. Whole-cell recordings further showed that oriens-lacunosum-moleculare (O-LM) interneurons, key players in the generation of theta rhythm, displayed marked changes in their intrinsic and synaptic properties. Hyperpolarization-activated mixed cation currents (Ih) were significantly reduced, resulting in an increase in the input resistance and a hyperpolarizing shift in the resting membrane potential. Additionally, the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) was increased, indicating a stronger excitatory input to these neurons. As a consequence, O-LM interneurons increased their firing rate from theta to gamma frequencies during induced network activity in acute slices from KA-injected mice. Thus, our physiological data together with network simulations suggest that changes in excitatory input and synaptic integration in O-LM interneurons lead to impaired rhythmogenesis in the hippocampus that in turn may underlie memory deficit.

On the formation of gamma-coherent cell assemblies by oriens lacunosum-moleculare interneurons in the hippocampus.

Gamma frequency (30-80 Hz) network oscillations have been observed in the hippocampus during several behavioral paradigms in which they are often modulated by a theta frequency (4-12 Hz) oscillation. Interneurons of the hippocampus have been shown to be crucially involved in rhythms generation, and several subtypes with distinct anatomy and physiology have been described. In particular, the oriens lacunosum-moleculare (O-LM) interneurons were shown to synapse on distal apical dendrites of pyramidal cells and to spike preferentially at theta frequency, even in the presence of gamma-field oscillations. O-LM cells have also recently been shown to present higher axonal ramification in the longitudinal axis of the hippocampus. By using a hippocampal network model composed of pyramidal cells and two types of interneurons (O-LM and basket cells), we show here that the O-LM interneurons lead to gamma coherence between anatomically distinct cell modules. We thus propose that this could be a mechanism for coupling longitudinally distant cells excited by entorhinal cortex inputs into gamma-coherent assemblies.

Increased inhibitory input to CA1 pyramidal cells alters hippocampal gamma frequency oscillations in the MK-801 model of acute psychosis.

The phencyclidine compound MK-801 can induce psychosis with symptoms which closely resemble those observed in an acute schizophrenic episode. Here we used an in vitro model of psychosis after systemic administration of MK-801. We found that kainate-induced gamma frequency field oscillations in animals previously exposed to MK-801 have significantly higher power than in control animals. The intrinsic membrane properties of pyramidal cells, such as membrane input resistance and time constant, were not found to be different. In contrast, the MK-801 cells exhibited significantly more depolarized resting membrane potentials than control cells. We propose cellular alterations in Na+-K+-pump activity and increases in phasic inhibition in MK-801 cells to be the respective underlying mechanisms responsible for the more depolarized resting membrane potentials and the increased power of gamma frequency oscillations observed in MK-801 pretreated animals.

Orthogonal arrangement of rhythm-generating microcircuits in the hippocampus.

As a structure involved in learning and memory, the hippocampus functions as a network. The functional differentiation along the longitudinal axis of the hippocampus is poorly demarcated in comparison with the transverse axis. Using patch clamp recordings in conjunction with post hoc anatomy, we have examined the pattern of connectivity and the functional differentiation along the long axis of the hippocampus. Here, we provide anatomical and physiological evidence that the prominent rhythmic network activities of the hippocampus, the behavior-specific gamma and theta oscillations, are seen predominantly along the transverse and longitudinal axes respectively. This orthogonal relationship is the result of the axonal field trajectories and the consequential interaction of the principal cells and major interneuron subtypes involved in generating each rhythm. Thus, the axonal arborization patterns of hippocampal inhibitory cells may represent a structural framework for the spatiotemporal distribution of activity observed within the hippocampus.

Differential involvement of oriens/pyramidale interneurones in hippocampal network oscillations in vitro.

Using whole-cell patch-clamp recordings in conjunction with post hoc anatomy we investigated the physiological properties of hippocampal stratum oriens and stratum pyramidale inhibitory interneurones, before and following the induction of pharmacologically evoked gamma frequency network oscillations. Prior to kainate-induced transient epochs of gamma activity, two distinct classes of oriens interneurones, oriens lacunosum-moleculare (O-LM) and trilaminar cells, showed prominent differences in their membrane and firing properties, as well as in the amplitude and kinetics of their excitatory postsynaptic events. In the active network both types of neurone received a phasic barrage of gamma frequency excitatory inputs but, due to their differential functional integration, showed clear differences in their output patterns. While O-LM cells fired intermittently at theta frequency, trilaminar interneurones discharged on every gamma cycle and showed a propensity to fire spike doublets. Two other classes of fast spiking interneurones, perisomatic targeting basket and bistratified cells, in the active network discharged predominantly single action potentials on every gamma cycle. Thus, within a locally excited network, O-LM cells are likely to provide a theta-frequency patterned output to distal dendritic segments, whereas basket and bistratified cells are involved in the generation of locally synchronous gamma band oscillations. The anatomy and output profile of trilaminar cells suggest they are involved in the projection of locally generated gamma rhythms to distal sites. Therefore a division of labour appears to exist whereby different frequencies and spatiotemporal properties of hippocampal rhythms are mediated by different interneurone subtypes.