Search

Abstract The complex neuromuscular network that controls body movements is the target of severe diseases that result in paralysis and death. Here, we report the development of a robust and efficient self-organizing neuromuscular junction (soNMJ) model from human pluripotent stem cells that can be maintained long-term in simple adherent conditions. The timely application of specific patterning signals instructs the simultaneous development and differentiation of position-specific brachial spinal neurons, skeletal muscles, and terminal Schwann cells. High-content imaging reveals self-organized bundles of aligned muscle fibers surrounded by innervating motor neurons that form functional neuromuscular junctions. Optogenetic activation and pharmacological interventions show that the spinal neurons actively instruct the synchronous skeletal muscle contraction. The generation of a soNMJ model from spinal muscular atrophy patient-specific iPSCs reveals that the number of NMJs and muscle contraction is severely affected, resembling the patient’s pathology. In the future, the soNMJ model could be used for high-throughput studies in disease modeling and drug development. Thus, this model will allow us to address unmet needs in the neuromuscular disease field.

Abstract Channelrhodopsins are light-gated ion channels used to control excitability of designated cells in large networks with high spatiotemporal resolution. While ChRs selective for H+, Na+, K+ and anions have been discovered or engineered, Ca2+-selective ChRs have not been reported to date. Here, we analyse ChRs and mutant derivatives with regard to their Ca2+ permeability and improve their Ca2+ affinity by targeted mutagenesis at the central selectivity filter. The engineered channels, termed CapChR1 and CapChR2 for calcium-permeable channelrhodopsins, exhibit reduced sodium and proton conductance in connection with strongly improved Ca2+ permeation at negative voltage and low extracellular Ca2+ concentrations. In cultured cells and neurons, CapChR2 reliably increases intracellular Ca2+ concentrations. Moreover, CapChR2 can robustly trigger Ca2+ signalling in hippocampal neurons. When expressed together with genetically encoded Ca2+ indicators in Drosophila melanogaster mushroom body output neurons, CapChRs mediate light-evoked Ca2+ entry in brain explants.

The intrinsic dynamics of neurons, in particular the generation action potentials, can impact neural network states and processes of encoding information. The authors demonstrate how the elevation of temperature induces a type of action potential dynamics that favors synchronization patterns in neural networks.

Hippocampal ripples are highly synchronous neural events critical for memory consolidation and retrieval. A minority of strong ripples has been shown to be of particular importance in situations of increased memory demands. The propagation dynamics of strong ripples inside the hippocampal formation are, however, still opaque. We analyzed ripple propagation within the hippocampal formation in a large open-access dataset comprising 267 Neuropixel recordings in 49 awake, head-fixed mice. Surprisingly, strong ripples (top 10% in ripple strength) propagate differentially depending on their generation point along the hippocampal longitudinal axis. The septal hippocampal pole is able to generate longer ripples that engage more neurons and elicit spiking activity for an extended time even at considerable distances. Accordingly, a substantial portion of the variance in strong ripple duration (R² = 0.463) is explained by the ripple generation location on the longitudinal axis, in agreement with a possible distinctive role of the hippocampal septal pole in conditions of high-memory demand. Moreover, we observed that the location of the ripple generation has a significant impact on the spiking rate modulation of different hippocampal subfields, even before the onset of the ripple. This finding suggests that ripple generation location plays a crucial role in shaping the neural activity across the hippocampus.

Summary: The recurrent emerging of novel viral variants of concern (VOCs) with evasion of preexisting antibody immunity upholds severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) case numbers and maintains a persistent demand for updated therapies. We selected the patient-derived antibody CV38-142 based on its potency and breadth against the VOCs Alpha, Beta, Gamma, and Delta for preclinical development into a therapeutic. CV38-142 showed in vivo efficacy in a Syrian hamster VOC infection model after post-exposure and therapeutic application and revealed a favorable safety profile in a human protein library screen and tissue cross-reactivity study. Although CV38-142 targets the same viral surface as sotrovimab, which maintains activity against Omicron, CV38-142 did not neutralize the Omicron lineages BA.1 and BA.2. These results highlight the contingencies of developing antibody therapeutics in the context of antigenic drift and reinforce the need to develop broadly neutralizing variant-proof antibodies against SARS-CoV-2.

Introduction: The antibody repertoire from CSF-derived antibody-secreting cells and memory B-cells in patients with encephalitis contains a considerable number of antibodies that do not target the disease-defining autoantigen such as the GABA or NMDA receptors. This study focuses on the functional relevance of autoantibodies to brain blood vessels in patients with GABAA and NMDA receptor encephalitis.Methods: We tested 149 human monoclonal IgG antibodies from the cerebrospinal fluid of six patients with different forms of autoimmune encephalitis on murine brain sections for reactivity to blood vessels using immunohistochemistry. Positive candidates were tested for reactivity with purified brain blood vessels, effects on transendothelial electrical resistance (TEER), and expression of tight junction proteins as well as gene regulation using human brain microvascular endothelial hCMEC/D3 cells as in vitro blood-brain barrier model. One blood-vessel reactive antibody was infused intrathecally by pump injection in mice to study in vivo binding and effects on tight junction proteins such as Occludin. Target protein identification was addressed using transfected HEK293 cells.Results: Six antibodies reacted with brain blood vessels, three were from the same patient with GABAAR encephalitis, and the other three were from different patients with NMDAR encephalitis. One antibody from an NMDAR encephalitis patient, mAb 011-138, also reacted with cerebellar Purkinje cells. In this case, treatment of hCMEC/D3 cells resulted in decreased TEER, reduced Occludin expression, and mRNA levels. Functional relevance in vivo was confirmed as Occludin downregulation was observed in mAb 011-138-infused animals. Unconventional Myosin-X was identified as a novel autoimmune target for this antibody.Discussion: We conclude that autoantibodies to blood vessels occur in autoimmune encephalitis patients and might contribute to a disruption of the blood-brain barrier thereby suggesting a potential pathophysiological relevance of these antibodies.

Communication between the hippocampus and neocortex is organized through high frequency sharp wave ripple activity. Here, the authors report ripple activity coupling between the hippocampus and granular retrosplenial cortex suggesting an involvement in communicating with the neocortex.

Wir erkennen die Wichtigkeit von Forschungsdaten und -software für unsere Forschungsprozesse an und ordnen die Veröffentlichung von Forschungsdaten und -software als wesentlichen Bestandteil der wissenschaftlichen Publikationstätigkeit ein. Dafür unterstützen wir als Verbund unsere Forschenden im Umgang mit Daten und Software nach den FAIR-Prinzipien in Einvernehmen mit dem DFG-Kodex “Leitlinien zur Sicherung guter wissenschaftlicher Praxis”. In Zusammenarbeit mit unseren Institutionen und Fachcommunities stellen wir adäquate Forschungsdatenmanagement-Werkzeuge und -Dienste bereit und befähigen unsere Forschenden zum Umgang damit. Dabei bauen wir vorzugsweise auf existierenden Angeboten auf und bemühen uns im Gegenzug um deren Anpassung an unsere Bedürfnisse. Wir streben Maßnahmen für die Definition und Sicherstellung der Qualität unserer Forschungsdaten und -software an. Wir verwenden vorzugsweise existierende Daten-/Metadatenstandards und vernetzen uns nach Möglichkeit für die Erstellung und Implementierung neuer Standards mit entsprechenden nationalen und internationalen Initiativen. Wir verfolgen die Entwicklungen im Bereich des Forschungsdaten- und -softwaremanagements und prüfen neu entstehende Empfehlungen und Richtlinien zeitnah auf ihre Umsetzbarkeit.

Recent advances in imaging technology have highlighted that scaffold proteins and receptors are arranged in subsynaptic nanodomains. The synaptic membrane-associated guanylate kinase (MAGUK) scaffold protein membrane protein palmitoylated 2 (MPP2) is a component of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-associated protein complexes and also binds to the synaptic cell adhesion molecule SynCAM 1. Using superresolution imaging, we show that-like SynCAM 1-MPP2 is situated at the periphery of the postsynaptic density (PSD). In order to explore MPP2-associated protein complexes, we used a quantitative comparative proteomics approach and identified multiple γ-aminobutyric acid (GABA)A receptor subunits among novel synaptic MPP2 interactors. In line with a scaffold function for MPP2 in the assembly and/or modulation of intact GABAA receptors, manipulating MPP2 expression had effects on inhibitory synaptic transmission. We further show that GABAA receptors are found together with MPP2 in a subset of dendritic spines and thus highlight MPP2 as a scaffold that serves as an adaptor molecule, linking peripheral synaptic elements critical for inhibitory regulation to central structures at the PSD of glutamatergic synapses.

Rapid removal of glutamate from the sites of glutamate release is an essential step in excitatory synaptic transmission. However, despite many years of research, the molecular mechanisms underlying the intracellular regulation of glutamate transport at tripartite synapses have not been fully uncovered. This limits the options for pharmacological treatment of glutamate-related motor disorders, including Huntington’s disease (HD). We therefore investigated the possible binding partners of transgenic EAAT2 and their alterations under the influence of mutant huntingtin (mHTT). Mass spectrometry analysis after pull-down of striatal YFP-EAAT2 from wild-type (WT) mice and heterozygote (HET) Q175 mHTT-knock-in mice identified a total of 148 significant (FDR < 0.05) binders to full-length EAAT2. Of them 58 proteins exhibited mHTT-related differences. Most important, in 26 of the 58 mHTT-sensitive cases, protein abundance changed back toward WT levels when the mice expressed a C-terminal-truncated instead of full-length variant of EAAT2. These findings motivated new attempts to clarify the role of astrocytic EAAT2 regulation in cortico-basal movement control. Striatal astrocytes of Q175 HET mice were targeted by a PHP.B vector encoding EAAT2 with different degree of C-terminal modification, i.e., EAAT2-S506X (truncation at S506), EAAT2-4KR (4 lysine to arginine substitutions) or EAAT2 (full-length). The results were compared to HET and WT injected with a tag-only vector (CTRL). It was found that the presence of a C-terminal-modified EAAT2 transgene (i) increased the level of native EAAT2 protein in striatal lysates and perisynaptic astrocyte processes, (ii) enhanced the glutamate uptake of transduced astrocytes, (iii) stimulated glutamate clearance at individual corticostriatal synapses, (iv) increased the glutamate uptake of striatal astrocytes and (iv) alleviated the mHTT-related hypokinesia (open field indicators of movement initiation). In contrast, over-expression of full-length EAAT2 neither facilitated glutamate uptake nor locomotion. Together, our results support the new hypothesis that preventing abnormal protein-protein interactions at the C-terminal of EAAT2 could eliminate the mHTT-related deficits in corticostriatal synaptic glutamate clearance and movement initiation.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that leads to the death of upper and lower motor neurons. While most cases of ALS are sporadic, some of the familial forms of the disease are caused by mutations in the gene encoding for the RNA-binding protein FUS. Under physiological conditions, FUS readily phase separates into liquid-like droplets in vivo and in vitro. ALS-associated mutations interfere with this process and often result in solid-like aggregates rather than fluid condensates. Yet, whether cells recognize and triage aberrant condensates remains poorly understood, posing a major barrier to the development of novel ALS treatments. Using a combination of ALS-associated FUS mutations, optogenetic manipulation of FUS condensation, chemically induced stress, and pH-sensitive reporters of organelle acidity, we systematically characterized the cause-effect relationship between the material state of FUS condensates and the sequestering of lysosomes. From our data, we can derive three conclusions. First, regardless of whether we use wild-type or mutant FUS, expression levels (i.e., high concentrations) play a dominant role in determining the fraction of cells having soluble or aggregated FUS. Second, chemically induced FUS aggregates recruit LAMP1-positive structures. Third, mature, acidic lysosomes accumulate only at FUS aggregates but not at liquid-condensates. Together, our data suggest that lysosome-degradation machinery actively distinguishes between fluid and solid condensates. Unraveling these aberrant interactions and testing strategies to manipulate the autophagosome-lysosome axis provides valuable clues for disease intervention.

Abstract Background Given the global increase in the aging population and age-related diseases, the promotion of healthy aging is one of the most crucial public health issues. This trial aims to contribute to the establishment of effective approaches to promote cognitive and brain health in older individuals with subjective cognitive decline (SCD). Presence of SCD is known to increase the risk of objective cognitive decline and progression to dementia due to Alzheimer’s disease. Therefore, it is our primary goal to determine whether spermidine supplementation has a positive impact on memory performance in this at-risk group, as compared with placebo. The secondary goal is to examine the effects of spermidine intake on other neuropsychological, behavioral, and physiological parameters. Methods The SmartAge trial is a monocentric, randomized, double-blind, placebo-controlled phase IIb trial. The study will investigate 12 months of intervention with spermidine-based nutritional supplementation (target intervention) compared with 12 months of placebo intake (control intervention). We plan to recruit 100 cognitively normal older individuals with SCD from memory clinics, neurologists and general practitioners in private practice, and the general population. Participants will be allocated to one of the two study arms using blockwise randomization stratified by age and sex with a 1:1 allocation ratio. The primary outcome is the change in memory performance between baseline and post-intervention visits (12 months after baseline). Secondary outcomes include the change in memory performance from baseline to follow-up assessment (18 months after baseline), as well as changes in neurocognitive, behavioral, and physiological parameters (including blood and neuroimaging biomarkers), assessed at baseline and post-intervention. Discussion The SmartAge trial aims to provide evidence of the impact of spermidine supplementation on memory performance in older individuals with SCD. In addition, we will identify possible neurophysiological mechanisms of action underlying the anticipated cognitive benefits. Overall, this trial will contribute to the establishment of nutrition intervention in the prevention of Alzheimer’s disease. Trial registration ClinicalTrials.gov, NCT03094546. Registered 29 March 2017—retrospectively registered. Protocol version Based on EA1/250/16 version 1.5

Chitinase 3-like-3 (Chi3l3) is expressed in microglia, but its function is not clear. Here the authors show that Chi3l3 is expressed in the subventricular zone in mouse experimental immune encephalitis, which induces oligodendrogenesis.

Synaptic plasticity is a cellular model for learning and memory. However, the expression mechanisms underlying presynaptic forms of plasticity are not well understood. Here, we investigate functional and structural correlates of presynaptic potentiation at large hippocampal mossy fiber boutons induced by the adenylyl cyclase activator forskolin. We performed 2-photon imaging of the genetically encoded glutamate sensor iGluu that revealed an increase in the surface area used for glutamate release at potentiated terminals. Time-gated stimulated emission depletion microscopy revealed no change in the coupling distance between P/Q-type calcium channels and release sites mapped by Munc13-1 cluster position. Finally, by high-pressure freezing and transmission electron microscopy analysis, we found a fast remodeling of synaptic ultrastructure at potentiated boutons: Synaptic vesicles dispersed in the terminal and accumulated at the active zones, while active zone density and synaptic complexity increased. We suggest that these rapid and early structural rearrangements might enable long-term increase in synaptic strength.

Summary: Sharp wave-ripples (SWRs) represent synchronous discharges of hippocampal neurons and are believed to play a major role in memory consolidation. A large body of evidence suggests that SWRs are exclusively generated in the CA3-CA2 network. In contrast, here, we provide several lines of evidence showing that the subiculum can function as a secondary SWRs generator. SWRs with subicular origin propagate forward into the entorhinal cortex as well as backward into the hippocampus proper. Our findings suggest that the output structures of the hippocampus are not only passively facilitating the transfer of SWRs to the cortex, but they also can actively contribute to the genesis of SWRs. We hypothesize that SWRs with a subicular origin may be important for the consolidation of information conveyed to the hippocampus via the temporoammonic pathway.

Serotonin (5-HT) is one of the major neuromodulators present in the mammalian brain and has been shown to play a role in multiple physiological processes. The mechanisms by which 5-HT modulates cortical network activity, however, are not yet fully understood. We investigated the effects of 5-HT on slow oscillations (SOs), a synchronized cortical network activity universally present across species. SOs are observed during anesthesia and are considered to be the default cortical activity pattern. We discovered that (±)3,4-methylenedioxymethamphetamine (MDMA) and fenfluramine, two potent 5-HT releasers, inhibit SOs within the entorhinal cortex (EC) in anesthetized mice. Combining opto- and pharmacogenetic manipulations with in vitro electrophysiological recordings, we uncovered that somatostatin-expressing (Sst) interneurons activated by the 5-HT2A receptor (5-HT2AR) play an important role in the suppression of SOs. Since 5-HT2AR signaling is involved in the etiology of different psychiatric disorders and mediates the psychological effects of many psychoactive serotonergic drugs, we propose that the newly discovered link between Sst interneurons and 5-HT will contribute to our understanding of these complex topics.

In the course of a day, brain states fluctuate, from conscious awake information-acquiring states to sleep states, during which previously acquired information is further processed and stored as memories. One hypothesis is that memories are consolidated and stored during “offline” states such as sleep, a process thought to involve transfer of information from the hippocampus to other cortical areas. Up and Down states (UDS), patterns of activity that occur under anesthesia and sleep states, are likely to play a role in this process, although the nature of this role remains unclear. Here we review what is currently known about these mechanisms in three anatomically distinct but interconnected cortical areas: somatosensory cortex, entorhinal cortex, and the hippocampus. In doing so, we consider the role of this activity in the coordination of “replay” during sleep states, particularly during hippocampal sharp-wave ripples. We conclude that understanding the generation and propagation of UDS may provide key insights into the cortico-hippocampal dialogue linking archi- and neocortical areas during memory formation.

Optogenetic tools enable precise experimental control of the behaviour of cells. Here, the authors introduce a genetically-encoded two-protein system that enables silencing of excitable cells such as neurons and cardiomyocytes using blue light, and demonstrate its utility both in vitro and In vivo.

The role of mossy cells (MCs) of the hippocampal dentate area has long remained mysterious. Recent research has begun to unveil their significance in spatial computation of the hippocampus. Here, we used an in vitro model of sharp wave-ripple complexes (SWRs), which contribute to hippocampal memory formation, to investigate MC involvement in this fundamental population activity. We find that a significant fraction of MCs (∼47%) is recruited into the active neuronal network during SWRs in the CA3 area. Moreover, MCs receive pronounced, ripple-coherent, excitatory and inhibitory synaptic input. Finally, we find evidence for SWR-related synaptic activity in granule cells that is mediated by MCs. Given the widespread connectivity of MCs within and between hippocampi, our data suggest a role for MCs as a hub functionally coupling the CA3 and the DG during ripple-associated computations.

The distinctive firing pattern of grid cells in the medial entorhinal cortex (MEC) supports its role in the representation of space. It is widely believed that the hexagonal firing field of grid cells emerges from neural dynamics that depend on the local microcircuitry. However, local networks within the MEC are still not sufficiently characterized. Here, applying up to eight simultaneous whole-cell recordings in acute brain slices, we demonstrate the existence of unitary excitatory connections between principal neurons in the superficial layers of the MEC. In particular, we find prevalent feed-forward excitation from pyramidal neurons in layer III and layer II onto stellate cells in layer II, which might contribute to the generation or the inheritance of grid cell patterns.

Summary: At presynaptic terminals, neurotransmitters are released by synaptic vesicle exocytosis at the active zone. In order to maintain efficient neurotransmission and proper synaptic structure, sites of vesicle fusion must be cleared rapidly by endocytosis. Therefore, the coupling of exo- and endocytosis is crucial. Despite many years of research, the exact molecular and biophysical requirements for the coupling of exo- and endocytosis remain unclear. We investigate whether endocytosis can be triggered in a calcium-independent fashion by evoking calcium-independent exocytosis using a hypertonic sucrose solution. We demonstrate that endocytosis can be triggered, in the absence of calcium influx, in a clathrin-independent manner that relies on actin polymerization. Our findings point to a central role of membrane tension dependent on actin for efficient coupling of exo- and endocytosis. : As Ca2+ is necessary for action-potential-evoked synaptic vesicle fusion, the role of Ca2+ in triggering endocytosis is difficult to discern. Orlando et al. find that Ca2+-independent exocytosis triggered by hyperosmotic shock results in the formation of endocytic pits in hippocampal synapses, which depend on actin polymerization. Keywords: endocytosis, calcium, presynaptic terminal, synapse, synaptic transmission, ultrastructure, exo-endocytosis coupling

All synapses require fusion-competent vesicles and coordinated Ca2+-secretion coupling for neurotransmission, yet functional and anatomical properties are diverse across different synapse types. We show that the presynaptic protein RIM-BP2 has diversified functions in neurotransmitter release at different central murine synapses and thus contributes to synaptic diversity. At hippocampal pyramidal CA3-CA1 synapses, RIM-BP2 loss has a mild effect on neurotransmitter release, by only regulating Ca2+-secretion coupling. However, at hippocampal mossy fiber synapses, RIM-BP2 has a substantial impact on neurotransmitter release by promoting vesicle docking/priming and vesicular release probability via stabilization of Munc13-1 at the active zone. We suggest that differences in the active zone organization may dictate the role a protein plays in synaptic transmission and that differences in active zone architecture is a major determinant factor in the functional diversity of synapses.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The subiculum is the gatekeeper between the hippocampus and cortical areas. Yet, the lack of a pyramidal cell-specific marker gene has made the analysis of the subicular area very difficult. Here we report that the vesicular-glutamate transporter 2 (VGLUT2) functions as a specific marker gene for subicular burst-firing neurons, and demonstrate that VGLUT2-Cre mice allow for Channelrhodopsin-2 (ChR2)-assisted connectivity analysis.

The majority of excitatory synapses are located on dendritic spines of cortical glutamatergic neurons. In spines, compartmentalized Ca2+ signals transduce electrical activity into specific long-term biochemical and structural changes. Action potentials (APs) propagate back into the dendritic tree and activate voltage gated Ca2+ channels (VGCCs). For spines, this global mode of spine Ca2+ signaling is a direct biochemical feedback of suprathreshold neuronal activity. We previously demonstrated that backpropagating action potentials (bAPs) result in long-term enhancement of spine VGCCs. This activity-dependent VGCC plasticity results in a large interspine variability of VGCC Ca2+ influx. Here, we investigate how spine VGCCs affect glutamatergic synaptic transmission. We combined electrophysiology, two-photon Ca2+ imaging and two-photon glutamate uncaging in acute brain slices from rats. T- and R-type VGCCs were the dominant depolarization-associated Ca2+conductances in dendritic spines of excitatory layer 2 neurons and do not affect synaptic excitatory postsynaptic potentials (EPSPs) measured at the soma. Using two-photon glutamate uncaging, we compared the properties of glutamatergic synapses of single spines that express different levels of VGCCs. While VGCCs contributed to EPSP mediated Ca2+ influx, the amount of EPSP mediated Ca2+ influx is not determined by spine VGCC expression. On a longer timescale, the activation of VGCCs by bAP bursts results in downregulation of spine NMDAR function.

The measurement of activity in vivo and in vitro has shifted from electrical to optical methods. While the indicators for imaging activity have improved significantly over the last decade, tools for analysing optical data have not kept pace. Most available analysis tools are limited in their flexibility and applicability to datasets obtained at different spatial scales. Here, we present SamuROI (Structured analysis of multiple user-defined ROIs), an open source Python-based analysis environment for imaging data. SamuROI simplifies exploratory analysis and visualization of image series of fluorescence changes in complex structures over time and is readily applicable at different spatial scales. In this paper, we show the utility of SamuROI in Ca2+-imaging based applications at three spatial scales: the micro-scale (i.e., sub-cellular compartments including cell bodies, dendrites and spines); the meso-scale, (i.e., whole cell and population imaging with single-cell resolution); and the macro-scale (i.e., imaging of changes in bulk fluorescence in large brain areas, without cellular resolution). The software described here provides a graphical user interface for intuitive data exploration and region of interest (ROI) management that can be used interactively within Jupyter Notebook: a publicly available interactive Python platform that allows simple integration of our software with existing tools for automated ROI generation and post-processing, as well as custom analysis pipelines. SamuROI software, source code and installation instructions are publicly available on GitHub and documentation is available online. SamuROI reduces the energy barrier for manual exploration and semi-automated analysis of spatially complex Ca2+ imaging datasets, particularly when these have been acquired at different spatial scales.

The entorhinal cortices in the temporal lobe of the brain are key structures relaying memory related information between the neocortex and the hippocampus. The medial entorhinal cortex (MEC) routes spatial information whereas the lateral entorhinal cortex (LEC) routes predominantly olfactory information to the hippocampus. Gamma oscillations are known to coordinate information transfer between brain regions by precisely timing population activity of neuronal ensembles. Here we studied the organization of in vitro gamma oscillations in the MEC & LEC of the transgenic (tg) APP-PS1 mouse model of Alzheimer’s Disease (AD) at 4 – 5 months of age. In vitro gamma oscillations using the kainate model peaked between 30 - 50 Hz and therefore we analyzed the oscillatory properties in the 20 – 60 Hz range. Our results indicate that the LEC shows clear alterations in frequency and power of gamma oscillations early-on in AD as compared to the MEC. The gamma-frequency oscillation slows down in the LEC and also the gamma power in dorsal LEC is decreased as early as 4 – 5 months in the tg APP-PS1 mice. The results of this study suggest that the timing of olfactory inputs from LEC to the hippocampus might be affected early on in AD, resulting in a possible erroneous integration of the information carried by the two input pathways to the hippocampal subfields.

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.

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.

A key feature of signalling in dendritic spines is the synapse-specific transduction of short electrical signals into biochemical responses. Ca2+ is a major upstream effector in this transduction cascade, serving both as a depolarising electrical charge carrier at the membrane and an intracellular second messenger. Upon action potential firing, the majority of spines are subject to global back-propagating action potential (bAP) Ca2+ transients. These transients translate neuronal suprathreshold activation into intracellular biochemical events. Using a combination of electrophysiology, two-photon Ca2+ imaging, and modelling, we demonstrate that bAPs are electrochemically coupled to Ca2+ release from intracellular stores via ryanodine receptors (RyRs). We describe a new function mediated by spine RyRs: the activity-dependent long-term enhancement of the bAP-Ca2+ transient. Spines regulate bAP Ca2+ influx independent of each other, as bAP-Ca2+ transient enhancement is compartmentalized and independent of the dendritic Ca2+ transient. Furthermore, this functional state change depends exclusively on bAPs travelling antidromically into dendrites and spines. Induction, but not expression, of bAP-Ca2+ transient enhancement is a spine-specific function of the RyR. We demonstrate that RyRs can form specific Ca2+ signalling nanodomains within single spines. Functionally, RyR mediated Ca2+ release in these nanodomains induces a new form of Ca2+ transient plasticity that constitutes a spine specific storage mechanism of neuronal suprathreshold activity patterns.

The subiculum (SUB) is a pivotal structure positioned between the hippocampus proper and various cortical and subcortical areas. Despite the growing body of anatomical and intrinsic electrophysiological data of subicular neurons, modulation of synaptic transmission in the SUB is not well understood. In the present study we investigated the role of group II metabotropic glutamate receptors (mGluRs), which have been shown to be involved in the regulation of synaptic transmission by suppressing presynaptic cAMP activity. Using field potential and patch-clamp whole cell recordings we demonstrate that glutamatergic transmission at CA1-SUB synapses is depressed by group II mGluRs in a cell-type specific manner. Application of the group II mGluR agonist (2S,1'R,2'R,3'R)-2-(2, 3-dicarboxycyclopropyl)glycine (DCG-IV) led to a significantly higher reduction of excitatory postsynaptic currents in subicular bursting cells than in regular firing cells. We further used low-frequency stimulation protocols and brief high-frequency bursts to test whether synaptically released glutamate is capable of activating presynaptic mGluRs. However, neither frequency facilitation is enhanced in the presence of the group II mGluR antagonist LY341495, nor is a test stimulus given after a high-frequency burst. In summary, we present pharmacological evidence for presynaptic group II mGluRs targeting subicular bursting cells, but both low- and high-frequency stimulation protocols failed to activate presynaptically located mGluRs.

Synapses exhibit strikingly different forms of plasticity over a wide range of time scales, from milliseconds to hours. Studies on synaptic plasticity typically use constant-frequency stimulation to activate synapses, whereas in vivo activity of neurons is irregular.Using extracellular and whole-cell electrophysiological recordings, we have here studied the synaptic responses at hippocampal mossy fiber synapses in vitro to stimulus patterns obtained from in vivo recordings of place cell firing of dentate gyrus granule cells in behaving rodents. We find that synaptic strength is strongly modulated on short- and long-lasting time scales during the presentation of the natural stimulus trains.We conclude that dynamic short- and long-term synaptic plasticity at the hippocampal mossy fiber synapse plays a prominent role in normal synaptic function.

BackgroundAmong the various hippocampal network patterns, sharp wave-ripples (SPW-R) are currently the mechanistically least understood. Although accurate information on synaptic interactions between the participating neurons is essential for comprehensive understanding of the network function during complex activities like SPW-R, such knowledge is currently notably scarce.Methodology/principal findingsWe demonstrate an in vitro approach to SPW-R that offers a simple experimental tool allowing detailed analysis of mechanisms governing the sharp wave-state of the hippocampus. We combine interface storage of slices with modifications of a conventional submerged recording system and established in vitro SPW-R comparable to their in vivo counterpart. We show that slice storage in the interface chamber close to physiological temperature is the required condition to preserve network integrity that is necessary for the generation of SPW-R. Moreover, we demonstrate the utility of our method for studying synaptic and network properties of SPW-R, using electrophysiological and imaging methods that can only be applied in the submerged system.Conclusions/significanceThe approach presented here demonstrates a reliable and experimentally simple strategy for studying hippocampal sharp wave-ripples. Given its utility and easy application we expect our model to foster the generation of new insight into the network physiology underlying SPW-R.

Recent analysis of genetically modified mice deficient in different kainate receptor (KAR) subunits have strongly pointed to a role of the GluK2 subunit, mediating the vulnerability of the brain towards seizures. Research concerning this issue has focused mainly on the hippocampus. However, several studies point to a potential role of other parts of the hippocampal formation, in particular the entorhinal cortex, in the development of epileptic seizures. There is extensive cell death after such seizures in layer III of the medial entorhinal cortex (LIII mEC), making this region of special interest for investigation into related pathological conditions. We therefore characterized KAR mediated currents in LIII mEC pyramidal neurons by several different approaches. Using patch-clamp technique, in combination with glutamate uncaging in horizontal brain slices, we show that LIII mEC neurons exhibit KAR currents. Use of genetically modified mice reveal that these currents are mediated by GluK2 containing KARs. The IV curve indicates the predominant presence of a Ca(2+) impermeable and edited form of the KAR. Finally, we show that GluK2 containing kainate receptors are essential for kainate-induced gamma oscillations within the entorhinal cortex.