28 research outputs found
Ocular Dominance Plasticity after Stroke Was Preserved in PSD-95 Knockout Mice.
No full textNeuronal plasticity is essential to enable rehabilitation when the brain suffers from injury, such as following a stroke. One of the most established models to study cortical plasticity is ocular dominance (OD) plasticity in the primary visual cortex (V1) of the mammalian brain induced by monocular deprivation (MD). We have previously shown that OD-plasticity in adult mouse V1 is absent after a photothrombotic (PT) stroke lesion in the adjacent primary somatosensory cortex (S1). Exposing lesioned mice to conditions which reduce the inhibitory tone in V1, such as raising animals in an enriched environment or short-term dark exposure, preserved OD-plasticity after an S1-lesion. Here we tested whether modification of excitatory circuits can also be beneficial for preserving V1-plasticity after stroke. Mice lacking postsynaptic density protein-95 (PSD-95), a signaling scaffold present at mature excitatory synapses, have lifelong juvenile-like OD-plasticity caused by an increased number of AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) -silent synapses in V1 but unaltered inhibitory tone. In fact, using intrinsic signal optical imaging, we show here that OD-plasticity was preserved in V1 of adult PSD-95 KO mice after an S1-lesion but not in PSD-95 wildtype (WT)-mice. In addition, experience-enabled enhancement of the optomotor reflex of the open eye after MD was compromised in both lesioned PSD-95 KO and PSD-95 WT mice. Basic V1-activation and retinotopic map quality were, however, not different between lesioned PSD-95 KO mice and their WT littermates. The preserved OD-plasticity in the PSD-95 KO mice indicates that V1-plasticity after a distant stroke can be promoted by either changes in excitatory circuitry or by lowering the inhibitory tone in V1 as previously shown. Furthermore, the present data indicate that an increased number of AMPA-silent synapses preserves OD-plasticity not only in the healthy brain, but also in another experimental paradigm of cortical plasticity, namely the long-range influence on V1-plasticity after an S1-lesion
The lack of synapsin alters presynaptic plasticity at hippocampal mossy fibers in male mice
No full textSynapsins are highly abundant presynaptic proteins that play a crucial role in neurotransmission and plasticity via the clustering of synaptic vesicles. The synapsin III isoform is usually downregulated after development, but in hippocampal mossy fiber boutons it persists in adulthood. Mossy fiber boutons express presynaptic forms of short- and long-term plasticity, which are thought to underlie different forms of learning. Previous research on synapsins at this synapse focused on synapsin isoforms I and II. Thus, a complete picture regarding the role of synapsins in mossy fiber plasticity is still missing. Here, we investigated presynaptic plasticity at hippocampal mossy fiber boutons by combining electrophysiological field recordings and transmission electron microscopy in a mouse model lacking all synapsin isoforms. We found decreased short-term plasticity - i.e. decreased facilitation and post-tetanic potentiation - but increased long-term potentiation in male synapsin triple knockout mice. At the ultrastructural level, we observed more dispersed vesicles and a higher density of active zones in mossy fiber boutons from knockout animals. Our results indicate that all synapsin isoforms are required for fine regulation of short- and long-term presynaptic plasticity at the mossy fiber synapse
Funktionelle Rolle Medial Septaler Projektionen zum Parasubiculum
No full textOscillations are a hallmark of brain activity and can be generated by local synchronisation mechanisms. They have been implicated in the communication between brain areas. An important type of oscillations are θ oscillations (4-12 Hz), which are associated with different behaviours, such as movements and navigation, but they also play a crucial role in memory formation and retrieval. One of the major θ rhythm generators in the brain is the medial septum (MS), which with its different types of projecting neurons, innervates many cortical areas and synchronises their activity. I investigated two major projection types of the MS: GABAergic (γ-aminobutyric acid – GABA) and cholinergic (acetylcholine – ACh) projections. Both projections are known to target the medial entorhinal cortex (MEC) and hippocampus. Parvalbumin positive (PV+) projections of the MS, which are GABAergic, are known to synchronise cortical networks via disinhibition often by inhibiting interneurons. In contrast, cholinergic projections of the MS project to a wide range of cell types in the MEC and hippocampus and can have substantially different effects on the target cell (e.g. activation or inhibition). Thus, their function on a network can range from increasing activity through depolarising excitatory cells, to more inhibition of the network by activating interneurons, or even modulating synaptic integration. Previous studies have focussed on identifying projections to the hippocampus and the MEC but did not consider the parasubiculum (PaS), a major input of the MEC. In this study, we electrophysiologically characterised cells in the PaS and demonstrated layer I interneurons to be distinctly different from putative layer II interneurons. The PaS, with its strong θ rhythmic firing cells, was shown to have the highest density of MS PV+ fibres in the parahippocampal formation, suggesting that it is an important target of MS projections and yet MS inputs to the PaS are unknown. Using channelrhodopsin (ChR2), a light sensitive ion channel, expressed in the MS of PV-Cre and ChAT-Cre (choline acetyltransferase) mice in-vivo, I identified GABAergic and cholinergic MS connections to the PaS in-vitro and demonstrated cell type specific projection patterns. I found that PV+ MS projections mainly inhibit interneurons in the PaS, including layer I interneurons, representing a novel cortical target of PV+ MS cells. On the other hand, cholinergic projections depolarise layer I interneurons and have multiple effects on deeper cells of the PaS, leading to a depolarisation or hyperpolarisation. To investigate a potential role of GABAergic projections in θ generation, I recorded local field potentials (LFP) in awake head-fixed mice and entrained oscillations in the PaS by stimulating with light in the MS. In contrast, local stimulation of fibres in the PaS could not entrain oscillation, suggesting that increased activity in the PaS might be required for MS PV+ cells to entrain θ. Taken together, stimulation of PV+ cells in the MS is sufficient to drive oscillations in the PaS, likely via disinhibition in line with other areas as the MEC and hippocampus. However, novel targets in layer I could be involved via cholinergic activation and GABAergic entrainment. Whether cholinergic activation by itself can entrain θ remains to be further investigated.Oszillationen sind ein Kennzeichen von Gehirnaktivität und können durch lokale Synchronisationsmechanismen generiert werden. Sie spielen eine wichtige Rolle bei der Kommunikation zwischen Gehirnarealen. Ein wichtiger Typ von Oszillationen sind θ Oszillationen (4 − 12 Hz), welche mit verschiedenen Verhalten wie Bewegung und Navigation assoziiert sind und eine wichtige Rolle in der Gedächtnisbildung und -abrufung spielen. Einer der wichtigen θ Generatoren im Gehirn ist das Mediale Septum (MS), welches mit seinen verschiedenen projizierenden Neuronen viele kortikale Regionen innerviert. Ich habe zwei Typen von Projektionen des MS untersucht: GABAerge (γ-Aminobuttersäure – GABA) und cholinerge (Acetylcholin – ACh) Projektionen. Beide Typen projizieren zum Medialen Entohinalen Kortex (MEC) und zum Hippocampus. Parvalbumin positive (PV+) Projektionen des MS können kortikale Netzwerke via Disinhibition, durch inhibieren von Interneuronen, synchronisieren. Im Gegensatz dazu projizieren cholinerge Projektionen des MS zu verschiedensten Zelltypen des MEC und des Hippocampus und können unterschiedliche weitreichende Effekte auf Zellen haben (z.B. Aktivierung und Inhibierung). Folglich können die Konsequenzen von Aktivierung des Netzwerkes via Depolarisation von exzitatorischen Zellen, über Inhibierung des Netzwerkes via Aktivierung von Interneuronen bis hin zur Modulation von synaptischer Integration reichen. In der Vergangenheit haben Studien sich auf die Identifizierung von Projektionen zum Hippocampus und MECs fokussiert, jedoch nicht zum Parasubiculum (PaS), eines der bedeutendsten Eingänge des MEC. In dieser Studie haben wir elektrophysiologisch Zellen im PaS charakterisiert und konnten herausstellen, dass Schicht I Zellen sich von anderen vermeintlichen Interneuronen in Schicht II unterscheiden. Das PaS, mit seinen im θ Rhythmus feuernden Zellen, hat die höchste Dichte von MS PV+ Fasern im parahippocampalen Netzwerk, was es als besonderes Ziel für MS Projektionen herausstellt. Dennoch sind Projektionen vom MS zum PaS nicht untersucht worden. Mit Hilfe von Channelrhodopsin (ChR2), einem lichtsensitivem Ionenkanal, welcher im MS von PV-Cre und ChAT-Cre Mäusen exprimiert wurde, konnte ich GABAerge und cholinerge MS Verbindungen zum PaS in-vitro detektieren und Zelltyp-speziefische Projektionen identifizieren. Ich konnte herausstellen, dass PV+ MS Projektionen hauptsächlich Interneurone im PaS inhibieren. Insbesondere Schicht I Interneurone stellen ein neues kortikales Ziel von PV+ MS Zellen dar. Im Gegensatz dazu werden Schicht I Interneurone des PaS durch cholinerge MS Projektionen depolarisiert wohingegen Zellen in tieferen Schichten depolarisiert oder hyperpolarisiert werden können. Um zu zeigen, dass man mit GABAergen Projektionen θ generieren kann, nahm ich das lokale Feldpotential (LFP) in Kopffixierten Mäusen auf und fand, dass man Oszillationen mit MS-Stimulation gleichschalten kann, jedoch eine Stimulation der Fasern im PaS nicht ausreichend ist. Das weist darauf hin, dass eine erhöhte PaS-Aktivität notwendig ist, um θ Oszillationen im PaS zu generieren. Zusammenfassend zeigt sich, dass eine Stimulation der PV+ Zellen im MS ausreichend ist, um im PaS Oszillationen zu generieren. Disinhibierung im PaS ist, ähnlich wie auch im MEC und Hippocampus, ein wahrscheinlicher Mechanismus. Weiterhin könnten jedoch neue Ziele von cholinergen und GABAergen Fasern in Schicht I bei der θ Generierung involviert sein. Ob θ Oszillationen durch cholinerge Projektionen gleichgeschaltet werden kann muss jedoch noch durch weitere Studien gezeigt werden
The presynaptic scaffolding protein Piccolo organizes the readily releasable pool at the calyx of Held
No full textAnoctamin Calcium-Activated Chloride Channels May Modulate Inhibitory Transmission in the Cerebellar Cortex.
No full textCalcium-activated chloride channels of the anoctamin (alias TMEM16) protein family fulfill critical functions in epithelial fluid transport, smooth muscle contraction and sensory signal processing. Little is known, however, about their contribution to information processing in the central nervous system. Here we examined the recent finding that a calcium-dependent chloride conductance impacts on GABAergic synaptic inhibition in Purkinje cells of the cerebellum. We asked whether anoctamin channels may underlie this chloride conductance. We identified two anoctamin channel proteins, ANO1 and ANO2, in the cerebellar cortex. ANO1 was expressed in inhibitory interneurons of the molecular layer and the granule cell layer. Both channels were expressed in Purkinje cells but, while ANO1 appeared to be retained in the cell body, ANO2 was targeted to the dendritic tree. Functional studies confirmed that ANO2 was involved in a calcium-dependent mode of ionic plasticity that reduces the efficacy of GABAergic synapses. ANO2 channels attenuated GABAergic transmission by increasing the postsynaptic chloride concentration, hence reducing the driving force for chloride influx. Our data suggest that ANO2 channels are involved in a Ca2+-dependent regulation of synaptic weight in GABAergic inhibition. Thus, in balance with the chloride extrusion mechanism via the co-transporter KCC2, ANO2 appears to regulate ionic plasticity in the cerebellum
ANO1 and ANO2 expression levels in the cerebellum.
No full text<p><b>(A)</b> Membrane topology model for anoctamin Ca<sup>2+</sup>-activated Cl<sup>-</sup> channels based on the X-ray structure of a fungal TMEM16 protein [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref042" target="_blank">42</a>]. The transmembrane domains 5 and 6 are thought to provide the pore-lining region in the homodimeric channel [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref095" target="_blank">95</a>]. Five negatively charged amino-acid residues <i>(E</i>, <i>D)</i> and an asparagine residue <i>(N)</i> in transmembrane domains 6–8 serve as Ca<sup>2+</sup>-binding sites involved in channel gating [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref039" target="_blank">39</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref041" target="_blank">41</a>]. Four alternatively spliced segments <i>(a—d)</i> determine the apparent Ca<sup>2+</sup>-sensitivity of the ANO1 channel [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref005" target="_blank">5</a>]. ANO2 has two isoforms <i>A</i> and <i>B</i> and a regulatory motif at a position homologous to segment <i>c</i> in ANO1 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.ref006" target="_blank">6</a>]. <b>(B)</b> RT-PCR analysis from mouse olfactory epithelium <i>(OE)</i> and mouse cerebellum <i>(CB)</i> yield similarly strong ANO1 signals in cerebellum but weaker signals for ANO2. <b>(C)</b> Immunoblots obtained from lysates of cerebellum <i>(CB)</i> and main olfactory epithelium <i>(OE)</i> from wild-type and Ano2<sup>-/-</sup> mice show an ANO1-specific signal at ~120 kDa with the ANO1<sub>in</sub> antiserum. <b>(D)</b> Rabbit anti-ANO2<sub>ex</sub> serum stains ANO2-specific bands <i>(asterisks)</i> in immunoblots obtained from lysates of main olfactory epithelium <i>(OE)</i> and eye, as well as in membrane-protein preparations of main olfactory bulb <i>(OB)</i> and cerebellum <i>(CB)</i>. ANO2 bands are not present in immunoblots from Ano2<sup>-/-</sup> mice.</p
PCR primer pairs used to characterize cerebellar ANO1 and ANO2.
No full text<p>PCR primer pairs used to characterize cerebellar ANO1 and ANO2.</p
The experience-enabled enhancement of the optomotor reflex of the open eye after monocular deprivation (MD) was compromised in both PT-lesioned PSD-95 WT and PSD-95 KO mice.
No full textIn contrast, enhancements of spatial vision were present in nonlesioned PSD-95 WT and PSD-95 KO mice. (A, B) Spatial frequency threshold of the optomotor response of the open eye in cycles per degree (cyc/deg) plotted against days after MD. After 7 days of MD, nonlesioned PSD-95 KO mice (A) as well as sham-treated control mice (B; data from Greifzu et al., 2011) showed a significant increase in the spatial frequency threshold of the optomotor reflex of the open eye. This experience-enabled increase was abolished by a PT in S1 (A, B). (C-F) Contrast sensitivity thresholds of the optomotor reflex of the open eye at 6 different spatial frequencies before (day 0) and 7 days after MD. For both nonlesioned PSD-95 KO (C) and PSD-95 WT mice (D), there was an increase in contrast sensitivity after 7 days of MD. After PT, this experience-enabled increase was absent in both groups (E, F).</p
Involvement of ANO2 in depolarization-induced depression of inhibition.
No full text<p><b>(A)</b> A cerebellar Purkinje cell loaded with the fluorescent dye Alexa Fluor 568. Scale bar: 10 μm. <b>(B)</b> Spontaneous postsynaptic currents in a Purkinje cell with E<sub>Cl</sub> near 0 mV and V<sub>hold</sub> = -69 mV. Overlay of 764 current traces showing similar time courses but differing amplitudes, probably reflecting distinct positions of GABAergic synapses on the Purkinje cell dendritic tree. <b>(C)</b> Postsynaptic currents were completely blocked by 50 μM picrotoxin, an inhibitor of GABA<sub>A</sub>-receptor chloride channels. <b>(D)</b> Protocol for activation of climbing fibers: Ten 0.1-ms current pulses were applied to the area near the proximal dendrite of a Purkinje cell while recording the whole-cell current of that cell at -70 mV. CF-activation produced characteristic complex spikes, as shown in the inset. <b>(E)</b><i>Upper traces</i>: GABAergic inhibitory postsynaptic currents recorded from a Purkinje cell at V<sub>hold</sub> = -48 mV and with 5 mM Cl<sup>-</sup> in the pipette solution. The positive polarity of IPSCs indicates Cl<sup>-</sup> influx. <i>Lower traces</i>: postsynaptic currents, recorded immediately after the climbing-fiber stimulation, displayed decreased amplitudes. <b>(F)</b> IPSCs recorded from a Purkinje cell of an Ano2<sup>-/—</sup>mouse before <i>(upper traces)</i> and immediately after <i>(lower traces)</i> CF-activation. <b>(G)</b> The number of detectable IPSC signals decreased by ~47% through climbing-fiber stimulation (before CF: 30.7 ± 6.5 min<sup>-1</sup>; after CF: 14.6 ± 3.4 min<sup>-1</sup>; 8 cells; <i>ctrl</i>). In slices from Ano2<sup>-/-</sup> mice, more IPSCs were detected (54.5 ± 18.5 min<sup>-1</sup>; 4 cells), and the activation of climbing fibers had no effect (52.5 ± 16.2 min<sup>-1</sup>; 4 cells).</p
Identification of an inhibitor specific for cerebellar ANO2 channels.
No full text<p><b>(A)</b> Whole-cell recording from an HEK293 cell transfected with mouse cerebellar ANO2 isoform <i>B</i> at -70 mV. Chloride inward current was activated by diffusion of 7.5 μM Ca<sup>2+</sup> from the pipette into the cell immediately after whole-cell breakthrough <i>(arrow)</i>. <b>(B)</b> Results from current recordings without <i>(black bars)</i> and with <i>(hatched bars) T16Ainh-A01</i> revealed that neither mouse ANO1<i>ac</i> nor ANO1<i>abc</i> were significantly inhibited by 5 μM of the compound, while the ANO2 isoform <i>B</i> showed a significantly reduced current density. At 25 μM <i>T16Ainh-A01</i>, significant inhibition was also observed with ANO1<i>ac</i>. Results were averaged from 19–30 cells for each condition. <b>(C)</b> When applied at the ANO2-specific concentration of 5 μM, <i>T16Ainh-A01</i> blocked <i>DDI</i>: The number of detectable IPSCs was 48.9 ± 12.1 min<sup>-1</sup> before and 53.4 ± 15.4 min<sup>-1</sup> after CF stimulation. ANO2<sup>+/+</sup> control data from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142160#pone.0142160.g004" target="_blank">Fig 4G</a> are included for comparison. <b>(D)</b> IPSC traces from wildtype mice in the presence of 5 μM <i>T16Ainh-A01</i> show no difference in IPSC shapes and amplitudes before <i>(left traces)</i> and after <i>(right traces)</i> CF-activation.</p
