The impact of organisational learning on service excellence in the Department of Science and Technology

The study focuses on organisational learning in the Department of Science and Technology (DST). Attention is paid to the meaning of the concept organisational learning, prerequisites for and factors of organisational learning for service excellence, organisational learning as an important phenomenon in a knowledge based organisation such as the DST. To determine the impact of organisational learning on service excellence in the DST, the study adopted a formalised, communicative, experimental and cross-sectional form of research design. The research methodology adopted in the study is that of qualitative research method in order to find substantial evidence. The study also employs a quantitative research method to complement the qualitative method. Both non-probability and probability sampling methods were employed in the study. The sample included 55 respondents from five programmes of the DST across all levels of the organisational structure. The results indicated that the DST leadership does support service excellence, thus highlighting the importance of communication in organisational learning.Public AdministrationM. P. A. (Public Administration

GABA actions and ionic plasticity in epilepsy

Concepts of epilepsy, based on a simple change in neuronal excitation/inhibition balance, have subsided in face of recent insights into the large diversity and context-dependence of signaling mechanisms at the molecular, cellular and neuronal network level. GABAergic transmission exerts both seizure-suppressing and seizure-promoting actions. These two roles are prone to short-term and long-term alterations, evident both during epileptogenesis and during individual epileptiform events. The driving force of GABAergic currents is controlled by ion-regulatory molecules such as the neuronal K-Cl cotransporter KCC2 and cytosolic carbonic anhydrases. Accumulating evidence suggests that neuronal ion regulation is highly plastic, thereby contributing to the multiple roles ascribed to GABAergic signaling during epileptogenesis and epilepsy.Peer reviewe

KCC2-Mediated Cl- Extrusion Modulates Spontaneous Hippocampal Network Events in Perinatal Rats and Mice

It is generally thought that hippocampal neurons of perinatal rats and mice lack transport-functional K-Cl cotransporter KCC2, and that Cl- regulation is dominated by Cl- uptake via the Na-K-2Cl cotransporter NKCC1. Here, we demonstrate a robust enhancement of spontaneous hippocampal network events (giant depolarizing potentials [GDPs]) by the KCC2 inhibitor VU0463271 in neonatal rats and late-gestation, wildtype mouse embryos, but not in their KCC2-null littermates. VU0463271 increased the depolarizing GABAergic synaptic drive onto neonatal CA3 pyramidal neurons, increasing their spiking probability and synchrony during the rising phase of a GDP. Our data indicate that Cl- extrusion by KCC2 is involved in modulation of GDPs already at their developmental onset during the perinatal period in mice and rats.Peer reviewe

Vasopressin excites interneurons to suppress hippocampal network activity across a broad span of brain maturity at birth

During birth in mammals, a pronounced surge of fetal peripheral stress hormones takes place to promote survival in the transition to the extrauterine environment. However, it is not known whether the hormonal signaling involves central pathways with direct protective effects on the perinatal brain. Here, we show that arginine vasopressin specifically activates interneurons to suppress spontaneous network events in the perinatal hippocampus. Experiments done on the altricial rat and precocial guinea pig neonate demonstrated that the effect of vasopressin is not dependent on the level of maturation (depolarizing vs. hyperpolarizing) of postsynaptic GABA(A) receptor actions. Thus, the fetal mammalian brain is equipped with an evolutionarily conserved mechanism well-suited to suppress energetically expensive correlated network events under conditions of reduced oxygen supply at birth.Peer reviewe

Carbonic anhydrase seven bundles filamentous actin and regulates dendritic spine morphology and density

Intracellular pH is a potent modulator of neuronal functions. By catalyzing (de)hydration of CO2, intracellular carbonic anhydrase (CA(i)) isoforms CA2 and CA7 contribute to neuronal pH buffering and dynamics. The presence of two highly active isoforms in neurons suggests that they may serve isozyme-specific functions unrelated to CO2-(de)hydration. Here, we show that CA7, unlike CA2, binds to filamentous actin, and its overexpression induces formation of thick actin bundles and membrane protrusions in fibroblasts. In CA7-overexpressing neurons, CA7 is enriched in dendritic spines, which leads to aberrant spine morphology. We identified amino acids unique to CA7 that are required for direct actin interactions, promoting actin filament bundling and spine targeting. Disruption of CA7 expression in neocortical neurons leads to higher spine density due to increased proportion of small spines. Thus, our work demonstrates highly distinct subcellular expression patterns of CA7 and CA2, and a novel, structural role of CA7.Peer reviewe

A variant of KCC2 from patients with febrile seizures impairs neuronal Cl- extrusion and dendritic spine formation

Genetic variation in SLC12A5 which encodes KCC2, the neuron‐specific cation‐chloride cotransporter that is essential for hyperpolarizing GABAergic signaling and formation of cortical dendritic spines, has not been reported in human disease. Screening of SLC12A5 revealed a co‐segregating variant (KCC2‐R952H) in an Australian family with febrile seizures. We show that KCC2‐R952H reduces neuronal Cl− extrusion and has a compromised ability to induce dendritic spines in vivo and in vitro. Biochemical analyses indicate a reduced surface expression of KCC2‐R952H which likely contributes to the functional deficits. Our data suggest that KCC2‐R952H is a bona fide susceptibility variant for febrile seizures.Peer reviewe

Bumepamine, a brain-permeant benzylamine derivative of bumetanide, does not inhibit NKCC1 but is more potent to enhance phenobarbital's anti seizure efficacy

Correction Volume: 143 Pages: 349-350 DOI: 10.1016/j.neuropharm.2018.10.012Based on the potential role of Na-K-Cl cotransporters (NKCCs) in epileptic seizures, the loop diuretic bumetanide, which blocks the NKCC1 isoforms NKCC1 and NKCC2, has been tested as an adjunct with phenobarbital to suppress seizures. However, because of its physicochemical properties, bumetanide only poorly penetrates through the blood-brain barrier. Thus, concentrations needed to inhibit NKCC1 in hippocampal and neocortical neurons are not reached when using doses (0.1-0.5 mg/kg) in the range of those approved for use as a diuretic in humans. This prompted us to search for a bumetanide derivative that more easily penetrates into the brain. Here we show that bumepamine, a lipophilic benzylamine derivative of bumetanide, exhibits much higher brain penetration than bumetanide and is more potent than the parent drug to potentiate phenobarbital's anticonvulsant effect in two rodent models of chronic difficult-to-treat epilepsy, amygdala kindling in rats and the pilocarpine model in mice. However, bumepamine suppressed NKCC1-dependent giant depolarizing potentials (GDPs) in neonatal rat hippocampal slices much less effectively than bumetanide and did not inhibit GABA-induced Ca2+ transients in the slices, indicating that bumepamine does not inhibit NKCC1. This was substantiated by an oocyte assay, in which bumepamine did not block NKCC1a and NKCC1b after either extra- or intracellular application, whereas bumetanide potently blocked both variants of NKCC1. Experiments with equilibrium dialysis showed high unspecific tissue binding of bumetanide in the brain, which, in addition to its poor brain penetration, further reduces functionally relevant brain concentrations of this drug. These data show that CNS effects of bumetanide previously thought to be mediated by NKCC1 inhibition can also be achieved by a close derivative that does not share this mechanism. Bumepamine has several advantages over bumetanide for CNS targeting, including lower diuretic potency, much higher brain permeability, and higher efficacy to potentiate the anti-seizure effect of phenobarbital.Peer reviewe

A variant of KCC2 from patients with febrile seizures impairs neuronal Cl- extrusion and dendritic spine formation

Genetic variation in SLC12A5 which encodes KCC2, the neuron‐specific cation‐chloride cotransporter that is essential for hyperpolarizing GABAergic signaling and formation of cortical dendritic spines, has not been reported in human disease. Screening of SLC12A5 revealed a co‐segregating variant (KCC2‐R952H) in an Australian family with febrile seizures. We show that KCC2‐R952H reduces neuronal Cl− extrusion and has a compromised ability to induce dendritic spines in vivo and in vitro. Biochemical analyses indicate a reduced surface expression of KCC2‐R952H which likely contributes to the functional deficits. Our data suggest that KCC2‐R952H is a bona fide susceptibility variant for febrile seizures.Peer reviewe

K(2P)18.1 translates T cell receptor signals into thymic regulatory T cell development

It remains largely unclear how thymocytes translate relative differences in T cell receptor (TCR) signal strength into distinct developmental programs that drive the cell fate decisions towards conventional (Tconv) or regulatory T cells (Treg). Following TCR activation, intracellular calcium (Ca2+) is the most important second messenger, for which the potassium channel K(2P)18.1 is a relevant regulator. Here, we identify K(2P)18.1 as a central translator of the TCR signal into the thymus-derived Treg (tTreg) selection process. TCR signal was coupled to NF-kappa B-mediated K(2P)18.1 upregulation in tTreg progenitors. K(2P)18.1 provided the driving force for sustained Ca2+ influx that facilitated NF-kappa B- and NFAT-dependent expression of FoxP3, the master transcription factor for Treg development and function. Loss of K(2P)18.1 ion-current function induced a mild lymphoproliferative phenotype in mice, with reduced Treg numbers that led to aggravated experimental autoimmune encephalomyelitis, while a gain-of-function mutation in K(2P)18.1 resulted in increased Treg numbers in mice. Our findings in human thymus, recent thymic emigrants and multiple sclerosis patients with a dominant-negative missense K(2P)18.1 variant that is associated with poor clinical outcomes indicate that K(2P)18.1 also plays a role in human Treg development. Pharmacological modulation of K(2P)18.1 specifically modulated Treg numbers in vitro and in vivo. Finally, we identified nitroxoline as a K(2P)18.1 activator that led to rapid and reversible Treg increase in patients with urinary tract infections. Conclusively, our findings reveal how K(2P)18.1 translates TCR signals into thymic T cell fate decisions and Treg development, and provide a basis for the therapeutic utilization of Treg in several human disorders.Peer reviewe

effects on neuronal signaling and on learning

1\. EINLEITUNG 1.1. Kationen-Chlorid-Kotransporter 1.1.1. Struktur und molekulare Diversität der Kationen-Chlorid-Kotransporter 1.1.2. Expressionsmuster der neuronalen Kationen-Chlorid-Kotransporter 1.1.2.1. Kalium-Chlorid-Kotransporter 2 (Kcc2) 1.1.2.2. Kalium-Chlorid-Kotransporter 3 (Kcc3) 1.1.2.3. Natrium-Kalium-Chlorid-Kotransporter 1 (Nkcc1) 1.1.3. Der Einfluss von Kcc2 auf die Morphologie von Neuronen 1.1.4. CCCs und die Polarität der GABAA-Rezeptorströme 1.1.4.1. Die Polarität der GABA-Antwort 1.1.4.2. Intraneuronale Cl--Gradienten 1.2. GABAerge und glycinerge Neurotransmission 1.2.1. Glycin- und GABA-Rezeptoren 1.2.1.1. GABAA-Rezeptoren 1.2.1.2. Glycinrezeptoren 1.2.2. GABAerge Inhibition 1.2.2.1. Klassische Hyperpolarisations-Inhibition 1.2.2.2. Shunting Inhibition 1.2.2.3. Tonische Inhibition 1.2.3. GABAerge Exzitation 1.2.3.1. Exzitatorisches GABA 1.2.3.2. Neuronale Maturation durch depolarisierendes GABA 1.3. Cerebellum 1.3.1. Neuronales Netzwerk des Cerebellums 1.3.2. Vorwärtshemmung und negative Rückkopplung 1.3.3. Kompensatorische Augenbewegungen 1.3.3.1. Vestibulo- okulärer Reflex 1.3.3.2. Adaption des vestibulo-okulären Reflexes 2\. FRAGESTELLUNG 3\. ERGEBNISSE 3.1. Kalium-Chlorid-Kotransporter 2 3.1.1. Konditionaler Knockout von Kcc2 in Körnerzellen und Purkinjezellen 3.1.1.1. Verwendete Cre-Linien 3.1.1.1.1. Analyse der L7/pcp-2:Cre-Maus 3.1.1.1.2. Analyse der Δα6Cre-Maus 3.1.1.2. Verifizierung des zellspezifischen Knockouts von Kcc2 3.1.1.2.1. Verpaarung der konditionalen Kcc2lox/lox- und L7-Cre- Mäusen 3.1.1.2.2. Zeitverlauf von Kcc2 Expression und Deletion in PC- ΔKcc2-Mäusen 3.1.1.2.3. Verpaarung der konditionalen Kcc2lox/lox- und Δα6Cre- Mäusen 3.1.1.2.4. Zeitverlauf von Kcc2 Expression und Deletion in GC- ΔKcc2-Mäusen 3.1.1.2.5. Verpaarung der konditionalen Kcc2lox/lox-Mäuse mit L7-Cre/Δα6Cre- Mäusen 3.1.1.2.6. Muster der Deletion von Kcc2 in PC-ΔKcc2- und GC-ΔKcc2- Mäusen 3.1.2. Der Einfluss von Kcc2 auf die Ausbildung von Synapsen 3.1.2.1. Histologische Untersuchung des Cerebellums 3.1.2.2. Untersuchung dendritischer Spines von Purkinjezellen 3.1.2.3. Immunohistochemische Quantifizierung exzitatorischer und inhibitorischer Synapsen im cerebellären Kortex von konditionalen Kcc2-Knockoutmäusen 3.1.2.4. Ultrastrukturelle Untersuchung der Synapsen im cerebellären Kortex 3.1.2.5. Elektrophysiologische Untersuchung inhibitorischer und exzitatorischer Postsynapsen von Purkinjezellen 3.1.3. Elektrophysiologische Untersuchung von PC-ΔKcc2-Mäusen 3.1.3.1. Ruhemembranpotential im Purkinjezellen und GABAerge Ströme 3.1.3.2. Determination von EGABA in Purkinjezellen 3.1.3.3. Chloridextrusion nach akuter Erhöhung von [Cl-]i 3.1.4. Elektrophysiologische Untersuchung von GC-ΔKcc2 Mäusen 3.1.4.1. Einzelkanalmessungen von GABAARs zur Bestimmung von DFGABA 3.1.4.2. Die Bestimmung des Ruhemembranpotentials von Körnerzellen 3.1.4.3. Cl--Konzentration und Permeabilität beeinflussen das Ruhemembranpotential von Körnerzellen 3.1.4.4. Nkcc1 in adulten Körnerzellen 3.1.4.5. Die Aktivierung von GABAARs führt zu Shunting Inhibition 3.1.5. Auswirkungen des konditionalen Kcc2-Knockouts auf das Netzwerk des Cerebellums 3.1.5.1. Spontane postsynaptische Ströme von Purkinjezellen in GC-ΔKcc2 Mäusen 3.1.5.2. Spontanaktivität von Purkinjezellen 3.1.5.3. Vestibulo-okulärer Reflex 3.1.5.3.1. Funktionsprüfung der okulären Reflexe 3.1.5.3.2. Kurzfristige Adaption des vestibulo-okulären Reflexes 3.1.5.3.3. Langzeitadaptation des vestibulo-okulären Reflexes 3.2. Kalium-Chlorid- Kotransporter 3 3.2.1. Konditionaler Knockout von Kcc3 in Purkinjezellen 3.2.2. Verifizierung des zellspezifischen Knockouts von Kcc3 3.2.3. Ruhemembranpotential im Purkinjezellen und GABAerge Ströme 3.2.4. Die Beteiligung von Kcc3 an Chloridextrusion nach akuter Erhöhung von [Cl-]i 60 4\. DISKUSSION 4.1. Kcc2 und Synapsenmaturation im Cerebellum 4.2. Kcc2 ist der dominierende Cl--Extruder von Purkinje- und Körnerzellen 4.2.1. Kcc2 senkt [Cl-]i von Purkinjezellen 4.2.2. [Cl-]i beeinflusst das Membranpotential von Körnerzellen 4.3. Signaltransduktion im cerebellären Kortex 4.3.1. Eine erhöhte neuronale [Cl-]i hat verschiedene Konsequenzen für Purkinjezellen und Körnerzellen 4.3.2. Die Auswirkungen der zellspezifischen Deletion von Kcc2 auf das neuronale Netzwerk des Cerebellums 4.4. Die Adaptation des vestibulo- okulären Reflexes ist durch zellspezifische Deletion von Kcc2 beeinträchtigt 5\. MATERIAL UND METHODEN 5.1. Material 5.1.1. Chemikalien und Enzyme 5.1.2. Filmmaterialien und bildgebende Verfahren 5.1.3. Zusammensetzung verwendeter Puffer und Lösungen 5.1.4. Bakterienstämme 5.1.5. Vektoren 5.1.6. Oligonukleotide 5.1.6.1. Genotypisierung 5.1.6.2. Sequenzierung 5.1.6.3. Sonden für In-situ Hybridisierungen 5.1.7. Antikörper 5.1.8. Mauslinien 5.2. Methoden 5.2.1. Mikrobiologische Methoden 5.2.1.1. Herstellung chemisch kompetenter Bakterien 5.2.1.2. Transformation von Bakterien 5.2.2. Molekularbiologische Methoden 5.2.2.1. Isolierung genomischer DNA aus Schwanzbiopsien 5.2.2.2. Polymerase-Kettenreaktion (PCR) 5.2.2.3. Präparation von Plasmid DNA aus 2 ml-Kulturen (Miniprep) 5.2.2.4. Präparation von Plasmid- DNA aus 100 ml-Kulturen (Midiprep) 5.2.2.5. Sequenzierung von DNA 5.2.2.6. Konzentrationsbestimmung von Nukleinsäuren 5.2.2.7. Restriktion von DNA 5.2.2.8. DNA-Gelelektrophorese 5.2.2.9. Isolierung von DNA-Fragmenten aus Agarosegelen 5.2.2.10. Ligation von DNA-Fragmenten 5.2.2.11. Phenol /Chloroform-Extraktion von DNA 5.2.2.12. in vitro-Transkription der Digitoxin- markierten RNA Sonden 5.2.2.13. Aufreinigung von RNA-Sonden 5.2.3. Proteinbiochemische Methoden 5.2.3.1. Herstellung von Gewebeextrakten für Western Blot 5.2.3.2. Polyacrylamidgel-Elektrophorese und Western Blot 5.2.3.3. Aufreinigung polyklonaler Antiseren 5.2.4. Histologische Methoden 5.2.4.1. Gelatinierung von Objektträgern 5.2.4.2. Perfusion von Mäusen 5.2.4.3. Kryotomschnitte 5.2.4.4. Frei schwimmende Schnitte 5.2.4.5. Hämatoxylin-Eosin-Färbung 5.2.4.6. X-Gal Färbung 5.2.4.7. Alkalische Phosphatase (AP) Färbung 5.2.4.8. Immunofluoreszenzfärbung von frei schwimmenden Schnitten 5.2.4.9. In situ Hybridisierung auf Gehirnschnitten 5.2.4.10. Elektronenmikroskopische Aufnahmen 5.2.5. Elektrophysiologische Methoden 5.2.5.1. Geräte 5.2.5.2. Akute Schnittpräparation für Elektrophysiologie 5.2.5.3. Patch clamp Messungen im akuten Gewebeschnitt 5.2.5.4. Gramicidin Perforated-Patch 5.2.5.5. Korrektur des Grenzflächenpotentials (Liquid junction potential) 5.2.5.6. Befüllung von Neuronen mit Biocytin 5.2.5.7. Messung GABAerger Miniaturströme 5.2.5.8. Messung glutamaterger Miniaturströme 5.2.5.9. Messung spontaner IPSCs (sIPSCs) 5.2.5.10. Messung spontaner EPSCs (sEPSCs) 5.2.5.11. Bestimmung des Ruhemembranpotentials im Perforated-Patch 5.2.5.12. Bestimmung des GABA-Umkehr Potentials 5.2.5.13. Messung der Chloridextrusion nach akuter Cl- Beladung 5.2.5.14. Einzelkanalmessungen von GABAA-Rezeptoren 5.2.5.15. Cell attached Messung des Membranpotentials 5.2.5.16. Bestimmung von [Cl-]i 5.2.5.17. Spontane Aktivität von Purkinjezellen 5.2.6. Verhalten 5.2.6.1. Aufnahme von Augenbewegungen 5.2.6.2. Optokinetischer Reflex (OKR) 5.2.6.3. Vestibulo- okulärer Reflex (VOR) 5.2.6.4. Lernparadigma 6\. REFERENZEN 7\. PUBLIKATION 8\. DANKSAGUNGENDas Cerebellum kontrolliert Bewegungen sowie das Erlernen motorischer Fähigkeiten. Alle Informationen, die im Cerebellum verarbeitet werden, durchlaufen das sehr gleichförmig aufgebaute neuronale Netzwerk des cerebellären Kortex. Hier konvergieren alle Signale auf Purkinjezellen, den einzigen Neuronen, deren Axone aus dem cerebellären Kortex heraus projizieren. Der Informationsfluss durch den cerebellären Kortex wird mittels Vorwärtshemmung und negativer Rückkopplung durch Interneuronen moduliert. Allerdings ist die Relevanz dieser inhibitorischen neuronalen Schaltkreise für motorisches Lernen und dessen Konsolidierung nicht aufgeklärt. Die schnelle Komponente der GABAergen Inhibition wird durch GABAA-Rezeptoren vermittelt. Da GABAA-Rezeptoren ligandenaktivierte Anionenkanäle sind, hängt der von ihnen vermittelte Strom von der Verteilung der leitbaren Anionen, vor allem Chlorid, über die Membran ab. Um die schnelle Inhibition zu vermitteln, ist eine niedrige intraneuronale Cl--Konzentration essentiell. Es besteht Konsens, dass der neuronenspezifische Kalium-Chlorid-Kotransporter Kcc2 die intrazelluläre Chloridkonzentration unter das elektrochemische Chloridgleichgewichtspotential absenkt, und so für eine hyperpolarisierende GABA-Antwort sorgt. Aufgrund der Interaktion des C-Terminus von Kcc2 mit einem Zytoskelett-assoziierten Protein, wurde Kcc2 ausserdem eine Schlüsselrolle in der Maturation dendritischer Spines zugeschrieben. Diese Interaktion beeinflusst ausserdem die Aggregation von AMPA-Rezeptoren in dendritischen Spines von adulten Neuronen, und dadurch die Effizienz glutamaterger Neurotransmission. Um die physiologische Funktion von Inhibition im cerebellären Netzwerk zu untersuchen, wurde Kcc2 spezifisch in cerebellären Purkinjezellen und Körnerzellen deletiert. In diesen Mausmodellen war weder die Morphologie der dendritischen Spines, noch die Dichte der Synapsen oder die glutamaterge Transmission messbar verändert. Die Deletion von Kcc2 führte in beiden Zelltypen zu einer Erhöhung der intrazellulären Chloridkonzentration um den Faktor zwei. In Purkinjezellen wurde hierdurch die GABAerge Inhibition zwar nicht vollständig unterbunden, aber immerhin drastisch reduziert. In Körnerzellen war zwar die elektrische Antwort auf GABA unverändert, doch die Verdopplung der intrazellulären Chloridkonzentration führte zu einer konstanten Depolarisation des Membranpotentials. Die Reduktion der GABAergen Inhibition von Purkinjezellen führte zu einer beeinträchtigeten Feinabstimmung der Motorik an veränderte Bedingungen. Dies wurde am Beispiel des vestibulo- okulären Reflexes untersucht. Sowohl das die Amplitude von kompensatorischen Augenbewegungen, als auch die zeitliche Feinabstimmung war reduziert. Dies legt eine fundamentale Rolle der Vorwärtshemmung von Purkinjezellen bei der Induktion von neuronaler Plastizität nahe. Die Deletion von Kcc2 in Körnerzellen hatte keinen Effekt auf die Adaption von kompensatorischen Augenbewegungen. Einzig die Übernacht-Konsolidierung von zeitlichen Abstimmungen des vestibulo-okulären Reflexes war drastisch eingeschränkt. Die GABAerge Inhibition war durch Deletion von Kcc2 nicht verändert. Daher liegt nahe, dass die Erregbarkeit von Körnerzellen eine sehr spezifische Rolle in der Konsolidierung von zeitlicher Feinabstimmung, nicht jedoch der Amplitude der kompensatorischen Augenbewegungen spielt.The cerebellum controls movement and learning of motor skills through a uniformly patterned neuronal circuitry, where all information converges onto a single type of neuron, the Purkinje cell. Purkinje cells receive glutamatergic input from climbing fibers and granule cells. Interneurons modulate information flow through the cerebellar cortex by feedforward and feedback inhibition. Their contribution to motor learning and consolidation is not fully understood. For fast inhibitory action of GABA, mediated by the GABAA receptor anion channels, low intracellular Cl- concentration is required. The neuron-specific KCl cotransporter Kcc2 is thought to lower the intracellular Cl- concentration below its electrochemical equilibrium potential and thereby rendering GABA hyperpolarizing. Additionally, Kcc2 was described as a key factor in the maturation of dendritic spines owed to interaction of its C-terminus with a cytoskeleton-associated protein. Furthermore it was reported that Kcc2 affects AMPA receptor aggregation in dendritic spines of adult neurons and thereby influences glutamatergic transmission. To investigate the functional roles of specific inhibitory pathways in cerebellar circuits, we specifically disrupted Kcc2 in cerebellar Purkinje cells and granule cells. We neither found abnormalities in dendritic spine morphology, nor in synapse density or glutamatergic transmission in these mouse models. The disruption of Kcc2 increased the intracellular Cl- concentration roughly twofold in both cell types. This drastically reduced, but not abolished, GABAergic inhibition on Purkinje cells. In granule cells it led to a constitutive depolarized membrane potential. Decreasing GABAergic inhibition on Purkinje cells in mice affected their ability to adjust their eye movements during vestibulo-ocular mismatch training. Furthermore, the consolidation of learned gain and phase adaptations was strongly compromised. This sugessts a fundamental role of feedforward inhibition onto Purkinje cells in the induction of plasticity. In contrast to this, Kcc2 ablation in granule cells specifically impaired overnight consolidation of phase learning of the vestibulo-ocular reflex, while gain modulation and short term adaptation of gain and phase remained intact. As GABAergic inhibition on Granule cells was unchanged in these mice, this suggests that Granule cell excitability plays a specific role in the consolidation of phase learning