Publications by authors named "Knut Kirmse"

27 Publications

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A limited role of NKCC1 in telencephalic glutamatergic neurons for developing hippocampal network dynamics and behavior.

Proc Natl Acad Sci U S A 2021 Apr;118(14)

Hans-Berger Department of Neurology, Jena University Hospital, 07747 Jena, Germany;

NKCC1 is the primary transporter mediating chloride uptake in immature principal neurons, but its role in the development of in vivo network dynamics and cognitive abilities remains unknown. Here, we address the function of NKCC1 in developing mice using electrophysiological, optical, and behavioral approaches. We report that NKCC1 deletion from telencephalic glutamatergic neurons decreases in vitro excitatory actions of γ-aminobutyric acid (GABA) and impairs neuronal synchrony in neonatal hippocampal brain slices. In vivo, it has a minor impact on correlated spontaneous activity in the hippocampus and does not affect network activity in the intact visual cortex. Moreover, long-term effects of the developmental NKCC1 deletion on synaptic maturation, network dynamics, and behavioral performance are subtle. Our data reveal a neural network function of NKCC1 in hippocampal glutamatergic neurons in vivo, but challenge the hypothesis that NKCC1 is essential for major aspects of hippocampal development.
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http://dx.doi.org/10.1073/pnas.2014784118DOI Listing
April 2021

Optimized photo-stimulation of halorhodopsin for long-term neuronal inhibition.

BMC Biol 2019 11 27;17(1):95. Epub 2019 Nov 27.

Hans-Berger Department of Neurology, Jena University Hospital, Am Klinikum 1, 07747, Jena, Germany.

Background: Optogenetic silencing techniques have expanded the causal understanding of the functions of diverse neuronal cell types in both the healthy and diseased brain. A widely used inhibitory optogenetic actuator is eNpHR3.0, an improved version of the light-driven chloride pump halorhodopsin derived from Natronomonas pharaonis. A major drawback of eNpHR3.0 is related to its pronounced inactivation on a time-scale of seconds, which renders it unsuited for applications that require long-lasting silencing.

Results: Using transgenic mice and Xenopus laevis oocytes expressing an eNpHR3.0-EYFP fusion protein, we here report optimized photo-stimulation techniques that profoundly increase the stability of eNpHR3.0-mediated currents during long-term photo-stimulation. We demonstrate that optimized photo-stimulation enables prolonged hyperpolarization and suppression of action potential discharge on a time-scale of minutes.

Conclusions: Collectively, our findings extend the utility of eNpHR3.0 to the long-lasting inhibition of excitable cells, thus facilitating the optogenetic dissection of neural circuits.
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http://dx.doi.org/10.1186/s12915-019-0717-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6882325PMC
November 2019

Editorial: GABAergic networks in the developing and mature brain.

Authors:
Knut Kirmse

Brain Res 2019 09 25;1718:10-11. Epub 2019 Apr 25.

Hans-Berger Department of Neurology, Jena University Hospital, 07747 Jena, Germany. Electronic address:

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http://dx.doi.org/10.1016/j.brainres.2019.04.029DOI Listing
September 2019

Somatostatin Interneurons Promote Neuronal Synchrony in the Neonatal Hippocampus.

Cell Rep 2019 03;26(12):3173-3182.e5

Hans-Berger Department of Neurology, Jena University Hospital, 07747 Jena, Germany. Electronic address:

Synchronized activity is a universal characteristic of immature neural circuits that is essential for their developmental refinement and strongly depends on GABAergic neurotransmission. A major subpopulation of GABA-releasing interneurons (INs) expresses somatostatin (SOM) and proved critical for rhythm generation in adulthood. Here, we report a mechanism whereby SOM INs promote neuronal synchrony in the neonatal CA1 region. Combining imaging and electrophysiological approaches, we demonstrate that SOM INs and pyramidal cells (PCs) coactivate during spontaneous activity. Bidirectional optogenetic manipulations reveal excitatory GABAergic outputs to PCs that evoke correlated network events in an NKCC1-dependent manner and contribute to spontaneous synchrony. Using a dynamic systems modeling approach, we show that SOM INs affect network dynamics through a modulation of network instability and amplification threshold. Our study identifies a network function of SOM INs with implications for the activity-dependent construction of developing brain circuits.
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http://dx.doi.org/10.1016/j.celrep.2019.02.061DOI Listing
March 2019

Pathogenic role of autoantibodies against inhibitory synapses.

Brain Res 2018 12 8;1701:146-152. Epub 2018 Sep 8.

Hans-Berger Department of Neurology, Jena University Hospital, D-07747 Jena, Germany.

Diverse neuropsychiatric diseases were recently linked to specific anti-neuronal autoantibodies targeting synaptic proteins. Symptoms can range from epileptic seizures to cognitive impairment to movement disorders, commonly responding to treatment with immunotherapy. Several of these autoantibodies target inhibitory synapses that use GABA or glycine as neurotransmitters. Despite their relatively low abundance, inhibitory neurons are extraordinarily diverse in anatomical, electrophysiological and molecular terms, reflecting the variable clinical phenotypes of affected patients. Indeed, data on the antibody effects in neuronal cultures or animals models suggest that most of these antibodies are directly pathogenic by down-regulating synaptic proteins, activating complement or antagonizing ligand binding. The present review summarizes the current achievements in the field of humoral autoimmunity related to inhibitory networks, state-of-the-art diagnostics and clinical characterization of patients. In many instances, the phenotypic spectrum of patients with GABA receptor, glycine receptor, amphiphysin or GAD65 antibodies mirror the experimental findings, suggesting that ongoing work will markedly contribute to the better understanding of pathophysiology in this exciting patient group and might pave the way for disease-specific immunotherapy.
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http://dx.doi.org/10.1016/j.brainres.2018.09.009DOI Listing
December 2018

Human Autoantibodies against the AMPA Receptor Subunit GluA2 Induce Receptor Reorganization and Memory Dysfunction.

Neuron 2018 10 23;100(1):91-105.e9. Epub 2018 Aug 23.

Hans-Berger Department of Neurology, Jena University Hospital, Am Klinikum 1, 07747 Jena, Germany; Center for Sepsis Control and Care (CSCC), Jena University Hospital, Am Klinikum 1, 07747 Jena, Germany. Electronic address:

AMPA receptors are essential for fast excitatory transmission in the CNS. Autoantibodies to AMPA receptors have been identified in humans with autoimmune encephalitis and severe defects of hippocampal function. Here, combining electrophysiology and high-resolution imaging with neuronal culture preparations and passive-transfer models in wild-type and GluA1-knockout mice, we analyze how specific human autoantibodies against the AMPA receptor subunit GluA2 affect receptor function and composition, synaptic transmission, and plasticity. Anti-GluA2 antibodies induce receptor internalization and a reduction of synaptic GluA2-containing AMPARs followed by compensatory ryanodine receptor-dependent incorporation of synaptic non-GluA2 AMPARs. Furthermore, application of human pathogenic anti-GluA2 antibodies to mice impairs long-term synaptic plasticity in vitro and affects learning and memory in vivo. Our results identify a specific immune-neuronal rearrangement of AMPA receptor subunits, providing a framework to explain disease symptoms.
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http://dx.doi.org/10.1016/j.neuron.2018.07.048DOI Listing
October 2018

Ultra-fast accurate reconstruction of spiking activity from calcium imaging data.

J Neurophysiol 2018 05 21;119(5):1863-1878. Epub 2018 Feb 21.

Department of Psychology, Technische Universität Dresden , Dresden , Germany.

Calcium imaging provides an indirect observation of the underlying neural dynamics and enables the functional analysis of neuronal populations. However, the recorded fluorescence traces are temporally smeared, thus making the reconstruction of exact spiking activity challenging. Most of the established methods to tackle this issue are limited in dealing with issues such as the variability in the kinetics of fluorescence transients, fast processing of long-term data, high firing rates, and measurement noise. We propose a novel, heuristic reconstruction method to overcome these limitations. By using both synthetic and experimental data, we demonstrate the four main features of this method: 1) it accurately reconstructs both isolated spikes and within-burst spikes, and the spike count per fluorescence transient, from a given noisy fluorescence trace; 2) it performs the reconstruction of a trace extracted from 1,000,000 frames in less than 2 s; 3) it adapts to transients with different rise and decay kinetics or amplitudes, both within and across single neurons; and 4) it has only one key parameter, which we will show can be set in a nearly automatic way to an approximately optimal value. Furthermore, we demonstrate the ability of the method to effectively correct for fast and rather complex, slowly varying drifts as frequently observed in in vivo data. NEW & NOTEWORTHY Reconstruction of spiking activities from calcium imaging data remains challenging. Most of the established reconstruction methods not only have limitations in adapting to systematic variations in the data and fast processing of large amounts of data, but their results also depend on the user's experience. To overcome these limitations, we present a novel, heuristic model-free-type method that enables an ultra-fast, accurate, near-automatic reconstruction from data recorded under a wide range of experimental conditions.
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http://dx.doi.org/10.1152/jn.00934.2017DOI Listing
May 2018

Developmental Emergence of Sparse Coding: A Dynamic Systems Approach.

Sci Rep 2017 10 12;7(1):13015. Epub 2017 Oct 12.

Department of Psychology, Technische Universität Dresden, 01187, Dresden, Germany.

During neocortical development, network activity undergoes a dramatic transition from largely synchronized, so-called cluster activity, to a relatively sparse pattern around the time of eye-opening in rodents. Biophysical mechanisms underlying this sparsification phenomenon remain poorly understood. Here, we present a dynamic systems modeling study of a developing neural network that provides the first mechanistic insights into sparsification. We find that the rest state of immature networks is strongly affected by the dynamics of a transient, unstable state hidden in their firing activities, allowing these networks to either be silent or generate large cluster activity. We address how, and which, specific developmental changes in neuronal and synaptic parameters drive sparsification. We also reveal how these changes refine the information processing capabilities of an in vivo developing network, mainly by showing a developmental reduction in the instability of network's firing activity, an effective availability of inhibition-stabilized states, and an emergence of spontaneous attractors and state transition mechanisms. Furthermore, we demonstrate the key role of GABAergic transmission and depressing glutamatergic synapses in governing the spatiotemporal evolution of cluster activity. These results, by providing a strong link between experimental observations and model behavior, suggest how adult sparse coding networks may emerge developmentally.
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http://dx.doi.org/10.1038/s41598-017-13468-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5638906PMC
October 2017

GABAergic Transmission during Brain Development: Multiple Effects at Multiple Stages.

Neuroscientist 2018 02 5;24(1):36-53. Epub 2017 Apr 5.

1 Hans-Berger Department of Neurology, Jena University Hospital, Jena, Germany.

In recent years, considerable progress has been achieved in deciphering the cellular and network functions of GABAergic transmission in the intact developing brain. First, in vivo studies in non-mammalian and mammalian species confirmed the long-held assumption that GABA acts as a mainly depolarizing neurotransmitter at early developmental stages. At the same time, GABAergic transmission was shown to spatiotemporally constrain spontaneous cortical activity, whereas firm evidence for GABAergic excitation in vivo is currently missing. Second, there is a growing body of evidence indicating that depolarizing GABA may contribute to the activity-dependent refinement of neural circuits. Third, alterations in GABA actions have been causally linked to developmental brain disorders and identified as potential targets of timed prophylactic interventions. In this article, we review these major recent findings and argue that both depolarizing and inhibitory GABA actions may be crucial for physiological brain maturation.
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http://dx.doi.org/10.1177/1073858417701382DOI Listing
February 2018

Column-like Ca(2+) clusters in the mouse neonatal neocortex revealed by three-dimensional two-photon Ca(2+) imaging in vivo.

Neuroimage 2016 Sep 21;138:64-75. Epub 2016 May 21.

Hans-Berger Department of Neurology, University Hospital Jena, D-07747 Jena, Germany. Electronic address:

Neuronal network activity in the developing brain is generated in a discontinuous manner. In the visual cortex during the period of physiological blindness of immaturity, this activity mainly comprises retinally triggered spindle bursts or Ca(2+) clusters thought to contribute to the activity-dependent construction of cortical circuits. In spite of potentially important developmental functions, the spatial structure of these activity patterns remains largely unclear. In order to address this issue, we here used three-dimensional two-photon Ca(2+) imaging in the visual cortex of neonatal mice at postnatal days (P) 3-4 in vivo. Large-scale voxel imaging covering a cortical depth of 200μm revealed that Ca(2+) clusters, identified as spindle bursts in simultaneous extracellular recordings, recruit cortical glutamatergic neurons of the upper cortical plate (CP) in a column-like manner. Specifically, the majority of Ca(2+) clusters exhibit prominent horizontal confinement and high intra-cluster density of activation involving the entire depth of the upper CP. Moreover, using simultaneous Ca(2+) imaging from hundreds of neurons at single-cellular resolution, we demonstrate that the degree of neuronal co-activation within Ca(2+) clusters displays substantial heterogeneity. We further provide evidence that co-activated cells within Ca(2+) clusters are spatially distributed in a non-stochastic manner. In summary, our data support the conclusion that dense coding in the form of column-like Ca(2+) clusters is a characteristic property of network activity in the developing visual neocortex. Such knowledge is expected to be relevant for a refined understanding of how specific spatiotemporal characteristics of early network activity instruct the development of cortical circuits.
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http://dx.doi.org/10.1016/j.neuroimage.2016.05.050DOI Listing
September 2016

Functional Indicators of Glutamate Transport in Single Striatal Astrocytes and the Influence of Kir4.1 in Normal and Huntington Mice.

J Neurosci 2016 05;36(18):4959-75

Cluster of Excellence NeuroCure Berlin and Department of Experimental Neurology, University Medicine Charité Berlin, Berlin D-10115, Germany

Unlabelled: This study evaluates single-cell indicators of glutamate transport in sulforhodamine 101-positive astrocytes of Q175 mice, a knock-in model of Huntington's disease (HD). Transport-related fluorescent ratio signals obtained with sodium-binding benzofuran isophtalate (SBFI) AM from unperturbed or voltage-clamped astrocytes and respective glutamate transporter currents (GTCs) were induced by photolytic or synaptic glutamate release and isolated pharmacologically. The HD-induced deficit ranged from -27% (GTC maximum at -100 mV in Ba(2+)) to -41% (sodium transients in astrocytes after loading SBFI-AM). Our specific aim was to clarify the mechanism(s) by which Kir4.1 channels can influence glutamate transport, as determined by either Na(+) imaging or transport-associated electrical signals. A decrease of Kir4.1 conductance was mimicked with Ba(2+) (200 μm), and an increase of Kir4.1 expression was obtained by intravenous administration of AAV9-gfaABC1D-Kir4.1-EGFP. The decrease of Kir4.1 conductance reduced the sodium transients but increased the amplitudes of somatic GTCs. Accordingly, after genetic upregulation of Kir4.1, somatic GTCs were found to be decreased. In individual cells, there was a negative correlation between Kir4.1 currents and GTCs. The relative effect of the Kir4.1 conductance was higher in the astrocyte periphery. These and other results suggest that the Kir4.1 conductance affects glutamate transporter activity in a dual manner: (1) by providing the driving force (voltage dependency of the transport itself) and (2) by limiting the lateral charge transfer (thereby reducing the interference with other electrogenic transporter functions). This leads to the testable prediction that restoring the high conductance state of passive astrocytes will not only normalize glutamate uptake but also restore other astrocytic transporter activities afflicted with HD.

Significance Statement: Insufficiency of astrocytic glutamate uptake is a major element in the pathophysiology of neurodegenerative diseases. Considering the heterogeneity of astrocytes and their differential susceptibility to therapeutic interventions, it becomes necessary to evaluate the determinants of transport activity in individual astroglial cells. We have examined intracellular Na(+) transients and glutamate transporter currents as the most telling indicators of glutamate clearance after synaptic or photolytic release of glutamate in striatal slices. The results show that, in Huntington's disease, glutamate uptake activity critically depends on Kir4.1. These channels enable the high conductance state of the astrocytic plasma membrane, which ensures the driving force for glutamate transport and dumps the transport-associated depolarization along the astrocyte processes. This has significant implications for developing therapeutic targets.
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http://dx.doi.org/10.1523/JNEUROSCI.0316-16.2016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6601850PMC
May 2016

Inferring Neuronal Dynamics from Calcium Imaging Data Using Biophysical Models and Bayesian Inference.

PLoS Comput Biol 2016 Feb 19;12(2):e1004736. Epub 2016 Feb 19.

Department of Psychology, Technische Universität Dresden, Dresden, Germany.

Calcium imaging has been used as a promising technique to monitor the dynamic activity of neuronal populations. However, the calcium trace is temporally smeared which restricts the extraction of quantities of interest such as spike trains of individual neurons. To address this issue, spike reconstruction algorithms have been introduced. One limitation of such reconstructions is that the underlying models are not informed about the biophysics of spike and burst generations. Such existing prior knowledge might be useful for constraining the possible solutions of spikes. Here we describe, in a novel Bayesian approach, how principled knowledge about neuronal dynamics can be employed to infer biophysical variables and parameters from fluorescence traces. By using both synthetic and in vitro recorded fluorescence traces, we demonstrate that the new approach is able to reconstruct different repetitive spiking and/or bursting patterns with accurate single spike resolution. Furthermore, we show that the high inference precision of the new approach is preserved even if the fluorescence trace is rather noisy or if the fluorescence transients show slow rise kinetics lasting several hundred milliseconds, and inhomogeneous rise and decay times. In addition, we discuss the use of the new approach for inferring parameter changes, e.g. due to a pharmacological intervention, as well as for inferring complex characteristics of immature neuronal circuits.
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http://dx.doi.org/10.1371/journal.pcbi.1004736DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4760968PMC
February 2016

Method to quantify accuracy of position feedback signals of a three-dimensional two-photon laser-scanning microscope.

Biomed Opt Express 2015 Oct 1;6(10):3678-93. Epub 2015 Sep 1.

Experimentelle Neurologie, Hans-Berger-Klinik für Neurologie, Universitätsklinikum Jena, Erlanger Allee 101, D-07747 Jena, Germany.

Two-photon laser-scanning microscopy enables to record neuronal network activity in three-dimensional space while maintaining single-cellular resolution. One of the proposed approaches combines galvanometric x-y scanning with piezo-driven objective movements and employs hardware feedback signals for position monitoring. However, readily applicable methods to quantify the accuracy of those feedback signals are currently lacking. Here we provide techniques based on contact-free laser reflection and laser triangulation for the quantification of positioning accuracy of each spatial axis. We found that the lateral feedback signals are sufficiently accurate (defined as <2.5 µm) for a wide range of scan trajectories and frequencies. We further show that axial positioning accuracy does not only depend on objective acceleration and mass but also its geometry. We conclude that the introduced methods allow a reliable quantification of position feedback signals in a cost-efficient, easy-to-install manner and should be applicable for a wide range of two-photon laser scanning microscopes.
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http://dx.doi.org/10.1364/BOE.6.003678DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4605029PMC
October 2015

GABA depolarizes immature neurons and inhibits network activity in the neonatal neocortex in vivo.

Nat Commun 2015 Jul 16;6:7750. Epub 2015 Jul 16.

Hans-Berger Department of Neurology, University Hospital Jena, D-07747 Jena, Germany.

A large body of evidence from in vitro studies suggests that GABA is depolarizing during early postnatal development. However, the mode of GABA action in the intact developing brain is unknown. Here we examine the in vivo effects of GABA in cells of the upper cortical plate using a combination of electrophysiological and Ca(2+)-imaging techniques. We report that at postnatal days (P) 3-4, GABA depolarizes the majority of immature neurons in the occipital cortex of anaesthetized mice. At the same time, GABA does not efficiently activate voltage-gated Ca(2+) channels and fails to induce action potential firing. Blocking GABA(A) receptors disinhibits spontaneous network activity, whereas allosteric activation of GABA(A) receptors has the opposite effect. In summary, our data provide evidence that in vivo GABA acts as a depolarizing neurotransmitter imposing an inhibitory control on network activity in the neonatal (P3-4) neocortex.
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http://dx.doi.org/10.1038/ncomms8750DOI Listing
July 2015

Impact of heme and heme degradation products on vascular diameter in mouse visual cortex.

J Am Heart Assoc 2014 Aug 28;3(4). Epub 2014 Aug 28.

Hans-Berger Department of Neurology, University Hospital Jena, Germany (A.J., S.G.W., A.W., K.K., O.W.W., K.H.).

Background: Delayed cerebral vasospasm is the most common cause of mortality and severe neurological impairment in patients who survive subarachnoid hemorrhage. Despite improvements in the field of diagnostic imaging, options for prevention and medical treatment-primarily with the calcium channel antagonist nimodipine or hemodynamic manipulations-are insufficient. Previous studies have suggested that heme and bilirubin oxidation end products, originating from degraded hemoglobin around ruptured blood vessels, are involved in the development of vasospasm by inhibiting large conductance BKC a potassium channels in vascular smooth muscle cells. In this study, we identify individual heme degradation products regulating arteriolar diameter in dependence of BKC a channel activity.

Methods And Results: Using differential interference contrast video microscopy in acute brain slices, we determined diameter changes of intracerebral arterioles in mouse visual cortex. In preconstricted vessels, the specific BKC a channel blockers paxilline and iberiotoxin as well as iron-containing hemin caused vasoconstriction. In addition, the bilirubin oxidation end product Z-BOX A showed a stronger vasoconstrictive potency than its regio-isomer Z-BOX B. Importantly, Z-BOX A had the same vasoconstrictive effect, independent of its origin from oxidative degradation or chemical synthesis. Finally, in slices of Slo1-deficient knockout mice, paxilline and Z-BOX A remained ineffective in changing arteriole diameter.

Conclusions: We identified individual components of the oxidative bilirubin degradation that led to vasoconstriction of cerebral arterioles. The vasoconstrictive effect of Z-BOX A and Z-BOX B was mediated by BKC a channel activity that might represent a signaling pathway in the occurrence of delayed cerebral vasospasm in subarachnoid hemorrhage patients.
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http://dx.doi.org/10.1161/JAHA.114.001220DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4310418PMC
August 2014

Reliable in vivo identification of both GABAergic and glutamatergic neurons using Emx1-Cre driven fluorescent reporter expression.

Cell Calcium 2012 Aug 31;52(2):182-9. Epub 2012 May 31.

Experimentelle Neurologie, Hans-Berger-Klinik für Neurologie, Universitätsklinikum Jena, Germany.

The development of genetically modified mice in which subpopulations of cortical neurons are labelled by fluorescent proteins has greatly facilitated single-cellular imaging and electrophysiology studies in vitro and in vivo. However, the parallel visualization of both inhibitory and excitatory neocortical neurons remains problematic. We here provide an alternative approach to identify GABAergic neurons in the context of in vivo calcium imaging. The method relies on the Emx1(IREScre) recombinase driven expression of a red fluorescent protein in excitatory neurons and glia. We quantitatively examined the upper layers of the visual neocortex in vivo and found that due to pronounced neuropil staining Emx1(IREScre)-negative and Emx1(IREScre)-positive neurons can be reliably differentiated based on negative and positive contrast, respectively. Immunohistochemical analyses confirmed that the entire population of GABAergic interneurons is represented by Emx1(IREScre)-negative cells. The potential usefulness of the method is exemplified by calcium imaging of sensory-evoked responses in the primary visual cortex. We conclude that the proposed method extends the repertoire of strategies aimed at discriminating two major neocortical neuron populations in situ.
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http://dx.doi.org/10.1016/j.ceca.2012.05.004DOI Listing
August 2012

GABAergic depolarization during early cortical development and implications for anticonvulsive therapy in neonates.

Epilepsia 2011 Sep 10;52(9):1532-43. Epub 2011 Jun 10.

Hans Berger Department of Neurology, Jena University Hospital, Jena, Germany.

Epileptic seizures rank among the most frequent neurologic symptoms during the neonatal period. Accumulating data from experimental animal studies and clinical trials in humans suggest that neonatal seizures could adversely affect normal brain development and result in long-term neurologic sequelae. Unfortunately, currently used anticonvulsive drugs are often ineffective in the neonatal period. One particularity of the immature neuronal network during neonatal development is that the neurotransmitter γ-aminobutyric acid (GABA) is mainly depolarizing, rather than hyperpolarizing as commonly observed in adults. This might, in part, explain not only the higher seizure propensity of the immature neuronal network, but also the limited anticonvulsive efficacy of GABA-enhancing drugs during early postnatal life. Accordingly, pharmacologic attenuation of GABAergic depolarization has been proposed as a strategy for neonatal seizure control. However, the underlying conjecture of a depolarizing mode of GABA action has been seriously challenged recently. In the present review, we will summarize the state of knowledge regarding GABAergic depolarization in early life and discuss how these data might impact a currently tested anticonvulsive strategy.
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http://dx.doi.org/10.1111/j.1528-1167.2011.03128.xDOI Listing
September 2011

GABA depolarizes immature neocortical neurons in the presence of the ketone body ß-hydroxybutyrate.

J Neurosci 2010 Nov;30(47):16002-7

Experimentelle Neurologie, Hans-Berger-Klinik für Neurologie, Universitätsklinikum Jena, 07747 Jena, Germany.

A large body of evidence suggests that the neurotransmitter GABA undergoes a developmental switch from being predominantly depolarizing-excitatory to predominantly hyperpolarizing-inhibitory. Recently published data, however, point to the possibility that the presumed depolarizing mode of GABA action during early development might represent an artifact due to an insufficient energy supply of the in vitro preparations used. Specifically, addition of the ketone body dl-β-hydroxybutyrate (βHB) to the extracellular medium was shown to prevent GABA from exerting excitatory effects. Applying a complementary set of minimally invasive optical and electrophysiological techniques in brain slices from neonatal mice, we investigated the effects of βHB on GABA actions in immature cells of the upper cortical plate. Fluorescence imaging revealed that GABA-mediated somatic [Ca(2+)] transients, that required activation of GABA(A) receptors and voltage-gated Ca(2+) channels, remained unaffected by βHB. Cell-attached current-clamp recordings showed that, in the presence of βHB, GABA still induced a membrane potential depolarization. To estimate membrane potential changes quantitatively, we used cell-attached recordings of voltage-gated potassium currents and demonstrated that the GABA-mediated depolarization was independent of supplementation of the extracellular solution with βHB. We conclude that, in vitro, GABA depolarizes immature cells of the upper cortical plate in the presence of the ketone body βHB. Our data thereby support the general concept of an excitatory-to-inhibitory switch of GABA action during early development.
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http://dx.doi.org/10.1523/JNEUROSCI.2534-10.2010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6633760PMC
November 2010

Estimation of ambient GABA levels in layer I of the mouse neonatal cortex in brain slices.

J Physiol 2010 Jul 26;588(Pt 13):2351-60. Epub 2010 Apr 26.

Institute of Physiology and Pathophysiology, University Medical Center of the Johannes Gutenberg University, Duesbergweg 6, Mainz, Germany.

GABAergic synapses on Cajal-Retzius neurons in layer I of the murine neocortex experience GABA(B) receptor (GABA(B)R)-mediated tonic inhibition. Extracellular GABA concentration ([GABA](o)) that determines the strength of GABA(B)R-mediated inhibition is controlled by GABA transporters (GATs). In this study, we hypothesized that the strength of presynaptic GABA(B)R activation reflects [GABA](o) in the vicinity of synaptic contacts. Slices obtained from two age groups were used, namely postnatal days (P)2-3 and P5-7. GABAergic postsynaptic currents (IPSCs) were recorded using the whole-cell patch-clamp technique. Minimal electrical stimulation in layer I was applied to elicit evoked IPSCs (eIPSCs) using a paired-pulse protocol. Three parameters were selected for comparison: the mean eIPSC amplitude, paired-pulse ratio, and failure rate. When GAT-1 and GAT-2/3 were blocked by NO-711 (10 microM) and SNAP-5114 (40 microM), respectively, no tonic GABA(B)R-mediated inhibition was observed. In order to restore the control levels of GABA(B)R-mediated inhibition, 250 and 125 nm exogenous GABA was required at P2-3 and P5-7, respectively. Addition of 3-mercaptopropionic acid, a glutamate decarboxylase inhibitor, did not significantly change the obtained values arguing against the suggestion that a mechanism different from GATs contributes to [GABA](o) control. We conclude that juxtasynaptic [GABA](o) is higher (about 250 nM) at P2-3 than at P5-7 (about 125 nM). As both radial cell migration and corticogenesis in general are strongly dependent on [GABA](o) and the formation of the last layer 2/3 is finished by P4 in rodents, the observed [GABA](o) reduction in layer I might reflect this crucial event in the cortical development.
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http://dx.doi.org/10.1113/jphysiol.2010.187054DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2915512PMC
July 2010

Role of GABA transporter 3 in GABAergic synaptic transmission at striatal output neurons.

Synapse 2009 Oct;63(10):921-9

Institute of Neurophysiology, Johannes Mueller Centre of Physiology, Charité-University Medicine Berlin, Berlin, Germany.

Striatal GABAergic signaling has been shown to be essential for basal ganglia output and proper motor performance. In the mouse neostriatum GABA transporter 1 (GAT-1) was previously found to assist in the clearance of GABA from the extracellular space and influence both phasic and tonic GABAergic inhibition of medium-sized striatal output neurons (SONs). It currently remains unknown whether GAT subtypes other than GAT-1 participate in the modulation of GABAergic transmission in this brain structure. In this study, we aimed at assessing the role of GAT-3 in the mouse neostriatum. To this end, we recorded GABAergic inhibitory postsynaptic currents (IPSCs) from SONs in brain slices at different developmental stages (postnatal days (P) 7-9, 12-14, and 28-34) using the whole-cell patch-clamp technique. When applied under control conditions, SNAP-5114 (40 microM), a specific GAT-3 blocker, did not affect miniature or evoked IPSCs (m/eIPSCs) and produced no significant effect on tonic GABAA receptor-mediated conductances in SONs. However, in the presence of NO-711 (10 microM), a specific GAT-1 blocker, SNAP-5114 reduced mIPSC frequencies without affecting mIPSC amplitudes or kinetics. In addition, SNAP-5114 reduced the mean amplitude of eIPSCs and increased the paired-pulse ratio. These effects were entirely abolished by CGP55845 (1 microM), a specific GABAB receptor blocker, indicating that they were mediated by presynaptic GABAB receptors. Similar results were obtained from all age groups. We conclude that GAT-3 is functionally expressed in the mouse neostriatum. Whereas an acute block of GAT-3 under resting conditions is fully compensated for by GAT-1, GAT-3 might provide an additional uptake capacity when neuronal activity and GABA release are increased.
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http://dx.doi.org/10.1002/syn.20675DOI Listing
October 2009

GABA transporter 1 tunes GABAergic synaptic transmission at output neurons of the mouse neostriatum.

J Physiol 2008 Dec 2;586(23):5665-78. Epub 2008 Oct 2.

Institute of Neurophysiology, Johannes Müller Centre of Physiology, Charité - University Medicine Berlin, Tucholskystr. 2, 10117 Berlin, Germany.

GABAergic medium-sized striatal output neurons (SONs) provide the principal output for the neostriatum. In vitro and in vivo data indicate that spike discharge of SONs is tightly controlled by effective synaptic inhibition. Although phasic GABAergic transmission critically depends on ambient GABA levels, the role of GABA transporters (GATs) in neostriatal GABAergic synaptic transmission is largely unknown. In the present study we aimed at elucidating the role of GAT-1 in the developing mouse neostriatum (postnatal day (P) 7-34). We recorded GABAergic postsynaptic currents (PSCs) using the whole-cell patch-clamp technique. Based on the effects of NO-711, a specific GAT-1 blocker, we demonstrate that GAT-1 is operative at this age and influences GABAergic synaptic transmission by presynaptic and postsynaptic mechanisms. Presynaptic GABA(B)R-mediated suppression of GABA release was found to be functional at all ages tested; however, there was no evidence for persistent GABA(B)R activity under control conditions, unless GAT-1 was blocked (P12-34). In addition, whereas no tonic GABA(A)R-mediated conductances were detected in SONs until P14, application of a specific GABA(A)R antagonist caused distinct tonic outward currents later in development (P19-34). In the presence of NO-711, tonic GABA(A)R-mediated currents were also observed at P7-14 and were dramatically increased at more mature stages. Furthermore, GAT-1 block reduced the median amplitude of GABAergic miniature PSCs indicating a decrease in quantal size. We conclude that in the murine neostriatum GAT-1 operates in a net uptake mode. It prevents the persistent activation of presynaptic GABA(B)Rs (P12-34) and prevents (P7-14) or reduces (P19-34) tonic postsynaptic GABA(A)R activity.
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http://dx.doi.org/10.1113/jphysiol.2008.161943DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2655402PMC
December 2008

Postsynaptically different inhibitory postsynaptic currents in Cajal-Retzius cells in the developing neocortex.

Neuroreport 2008 Aug;19(12):1213-6

Institute of Neurophysiology, Johannes-Mueller-Center of Physiology, Charité-University-Medicine Berlin, Berlin, Germany.

Fast and slowly rising inhibitory postsynaptic currents (IPSCs, IPSCF and IPSCS) in neocortical Cajal-Retzius cells are observed. In this study, zolpidem, a benzodiazepine agonist that specifically modulates gamma-aminobutyric acid type A receptors (GABAARs) containing gamma2 subunit, was used to characterize GABAARs mediating IPSCF and IPSCS. One-hundred-nanomolar zolpidem prolonged IPSCS, increased evoked IPSCS (eIPSCS) amplitude, and decreased paired-pulse ratio (PPR) of eIPSCS. Two micromolar zolpidem prolonged both IPSCF and IPSCS, increased miniature IPSCF and eIPSCF amplitudes, increased eIPSCS amplitude but not miniature IPSCS amplitude, decreased PPR of eIPSCS, but failed to affect PPR of eIPSCF. We conclude that IPSCF are mediated by alpha2/3-containing GABAARs, which are not saturated by synaptic GABA release, whereas IPSCS are mediated by alpha1-containing and alpha2/3-containing GABAARs, which are saturated by quantal GABA release.
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http://dx.doi.org/10.1097/WNR.0b013e328308daa0DOI Listing
August 2008

Cajal Retzius cells in the mouse neocortex receive two types of pre- and postsynaptically distinct GABAergic inputs.

J Physiol 2007 Dec 25;585(Pt 3):881-95. Epub 2007 Oct 25.

Institute of Neurophysiology, Johannes-Mueller-Center of Physiology, Charité-University-Medicine Berlin, Tucholskystrasse 2, 10117 Berlin, Germany.

Cajal-Retzius (CR) cells are principal cells of layer I in the developing neocortex. They are able to generate action potentials, make synaptic contacts in layer I and receive excitatory GABAergic inputs before birth. Although CR cells participate in neuronal network activity in layer I, the properties of their synaptic inputs are not yet characterized. We recorded miniature (mIPSCs) and evoked (eIPSCs) postsynaptic currents using the whole-cell patch-clamp technique. Most of CR cells displayed two types of mIPSCs, namely those with fast (mIPSC(F)) and slow (mIPSC(S)) rise kinetics. The mIPSC(F) mean amplitude was significantly larger than that of mIPSC(S), while their decay rates were not different. Peak-scaled non-stationary noise analysis revealed that mIPSC(S) and mIPSC(F) differed in their weighted single-channel conductance. In addition, zolpidem (100 nm), a modulator of alpha(1) subunit-containing GABA(A) receptors, selectively affected mIPSC(S) suggesting that different postsynaptic GABA(A) receptors mediate mIPSC(F) and mIPSC(S). eIPSCs also split into two populations with different rise kinetics. Fast eIPSCs (eIPSC(F)) displayed higher paired-pulse ratio (PPR) and lower GABA release probability than slowly rising eIPSCs (eIPSC(S)). As CGP55845, a GABA(B) receptor antagonist, eliminated the observed difference in PPR, the lower release probability at IPSC(F) connections probably reflects a stronger tonic GABA(B) receptor-mediated inhibition of IPSC(F) synapses. At low (0.1 Hz) stimulation frequency both inputs can effectively convert presynaptic action potentials into postsynaptic ones; however, only IPSC(F) connections reliably transfer the presynaptic activity patterns at higher stimulation rates. Thus, CR cells receive two GABAergic inputs, which differ in the quantal amplitude, the probability of GABA release and the frequency dependence of signal transfer.
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http://dx.doi.org/10.1113/jphysiol.2007.145003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2375528PMC
December 2007

Developmental downregulation of excitatory GABAergic transmission in neocortical layer I via presynaptic adenosine A(1) receptors.

Cereb Cortex 2008 Feb 7;18(2):424-32. Epub 2007 Jun 7.

Institute of Neurophysiology, Johannes-Mueller-Center of Physiology, Charité-University-Medicine Berlin, Tucholskystr. 2, 10117 Berlin, Germany.

Layer I of the developing cortex contains a dense GABAergic fiber plexus. These fibers provide excitatory inputs to Cajal-Retzius (CR) cells, the early born neurons in layer I. CR cells possess an extensive axonal projection and form synaptic contacts with excitatory, presumably pyramidal, neurons before birth. Interestingly, activity of CR cells declines during the first postnatal week, but mechanism(s) underlying this phenomenon is not yet known. Here we recorded inhibitory postsynaptic currents (IPSCs) in CR cells at postnatal day (P) 1-2 and P5-7. Blockade of adenosine A(1) receptors (A(1)Rs) increased the amplitude of evoked IPSCs (eIPSCs) and decreased paired-pulse ratio at P5-7 but not at P1-2. A(1)R activation decreased the mean eIPSC amplitude at P5-7, but failed to affect eIPSCs at P1-2. Ecto-adenosine triphosphatase (ATPase) inhibition completely abolished the A(1)R-mediated effects suggesting that extracellular ATP is the main source of adenosine. Because A(1)R blockade did not affect the median miniature IPSC amplitude, our results demonstrate that adenosine reduces gamma-aminiobutyric acid (GABA) release probability via presynaptic A(1)Rs at P5-7. As neuronal activity in layer I can depolarize pyramidal neurons influencing thereby glutamatergic synaptogenesis in the lower cortical layers, postnatal weakening of GABAergic transmission by adenosinergic system might reflect a developmental downregulation of this excitatory drive when glutamatergic synapses are formed.
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http://dx.doi.org/10.1093/cercor/bhm077DOI Listing
February 2008

N-ethylmaleimide increases release probability at GABAergic synapses in layer I of the mouse visual cortex.

Eur J Neurosci 2006 Nov;24(10):2741-8

Sensory and Developmental Physiology Group, Institute of Neurophysiology, Johannes-Mueller-Center of Physiology, Charité-University Medicine Berlin, Berlin, Germany.

The sulphydryl alkylating agent N-ethylmaleimide (NEM) has been often used as an uncoupler of pertussis toxin-sensitive G-proteins. However, the effects of NEM on gamma-aminobutyric acid (GABA)ergic synaptic transmission remain controversial. Using the whole-cell patch-clamp technique, GABA(A) receptor-mediated postsynaptic currents (IPSCs) have been recorded from Cajal-Retzius (CR) cells in layer I of the neonatal mouse visual cortex. NEM increased the frequencies of both spontaneous and miniature IPSCs (mIPSCs) without an effect on the median mIPSC amplitudes or mIPSC kinetics. The NEM actions on mIPSCs did not depend on the extracellular Ca(2+), Ca(2+) release from intracellular stores, adenylyl cyclase and protein kinase A activities. NEM increased the mean amplitudes of evoked IPSCs and strongly decreased the paired-pulse ratio. The size of the readily releasable pool of presynaptic vesicles (RRP) was estimated using a high-frequency stimulation protocol. The RRP size was not affected by NEM. In addition, NEM significantly decreased the latency between the stimulus and the onset of GABA release. These results suggest that NEM selectively increases GABA release probability. At postnatal day 2, mIPSCs were observed only in about 30% of CR cells. NEM application revealed, however, that more than 90% of CR cells receive GABAergic inputs. Therefore, NEM seems to be a useful tool to verify the existence of 'silent' GABAergic synapses.
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http://dx.doi.org/10.1111/j.1460-9568.2006.05179.xDOI Listing
November 2006

Ambient GABA constrains the strength of GABAergic synapses at Cajal-Retzius cells in the developing visual cortex.

J Neurosci 2006 Apr;26(16):4216-27

Sensory and Developmental Physiology Group, Institute of Neurophysiology, Johannes-Mueller-Center of Physiology, Charité-University Medicine Berlin, 10117 Berlin, Germany.

At early stages of brain development, GABA plays a dual role. It fulfills important trophic functions and provides a major excitatory drive for the immature neuronal network. Here, we investigated whether GABA itself can limit the strength of excitatory GABAergic synapses on Cajal-Retzius (CR) cells in sagittal slices from the mouse visual cortex. (2S)-3-[[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl](phenylmethyl)phosphinic acid (CGP55845), a specific GABAB receptor (GABABR) blocker, increased the frequency of spontaneous Ca2+ transients and spontaneous and miniature IPSCs (mIPSCs) but did not affect mIPSC amplitudes or kinetics. CGP55845 significantly increased evoked IPSC (eIPSC) amplitudes and decreased the paired-pulse ratio (PPR). Baclofen, a specific GABABR agonist, produced opposite effects. The size of the readily releasable pool was not affected by these GABABR modulators. The same CGP55845 actions were observed at physiological temperatures, but they were abolished after glutamate decarboxylase block with 3-mercaptopropionic acid (3-MP). These results indicate that presynaptic GABABRs dynamically regulate GABA release probability. SNAP-5114, a specific GABA transporter-2/3 (GAT-2/3) blocker, enhanced mIPSC frequencies, decreased PPR, and increased eIPSC amplitudes without changing eIPSC kinetics. These effects were blocked by CGP55845 and 3-MP. NO-711, a specific GAT-1 blocker, prolonged eIPSC decay and decreased eIPSC/mIPSC amplitudes. These NO-711-mediated effects were not sensitive to CGP55845 and 3-MP. We conclude that the strength of GABAergic inputs to CR cells is constrained by GABABRs that are persistently activated by ambient GABA. The latter is also provided by GAT-2/3 operating in the reversed mode. Presynaptic GAT-1 functions in the uptake mode and possibly provides GABA for presynaptic vesicle filling.
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http://dx.doi.org/10.1523/JNEUROSCI.0589-06.2006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6674013PMC
April 2006

Developmental downregulation of low-voltage-activated Ca2+ channels in Cajal-Retzius cells of the mouse visual cortex.

Eur J Neurosci 2005 Jun;21(12):3269-76

Developmental Physiology, Johannes Müller Centre of Physiology, Institute of Neurophysiology, 10117 Berlin, Germany.

Cajal-Retzius (CR) cells have been demonstrated to fulfil an important secretory function in the developing neocortex. On the other hand, the contribution of CR cell electrical activity during cortex development is still unclear. Using the whole-cell patch-clamp technique, we studied low-voltage-activated (LVA) Ca(2+) channels in CR cells in the layer I of the mouse visual cortex. CR cells were found to display a transient Ca(2+) current (I(T)) in response to a depolarization step from -100 mV to -40 mV. I(T) showed: (i) typical for LVA Ca(2+) channels voltage dependence of activation (half-activation at -55 mV) and inactivation (half-inactivation at -76 mV); (ii) fast activation and inactivation kinetics, with time constants of 1.4 and 28 ms, respectively, at -40 mV; (iii) fast recovery from steady-state inactivation (time constant: 290 ms); (iv) a complete block by 1 microm mibefradil; and (v) a partial block (to 55%) by 100 microm Ni(2+). The density of I(T) dramatically decreased between postnatal day (P) 1 and P9. Immunostaining demonstrated the presence and postnatal downregulation of the alpha(1G)-subunit of LVA Ca(2+) channels in CR cells. Experiments performed in the current-clamp mode revealed that mibefradil delayed an action potential generation in response to a suprathreshold depolarizing current at P1, but not at P8-9. We suggest that LVA Ca(2+) channels might influence CR cell excitability during the first postnatal week and thereby contribute to the shaping of synaptic connectivity in the neocortical layer I.
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http://dx.doi.org/10.1111/j.1460-9568.2005.04171.xDOI Listing
June 2005