Auswirkungen des Off Pump Coronary Artery Bypass Grafting (OPCAB) auf die Kontraktilität des Herzmuskels : ein Vergleich mit der Schädigung durch die extrakorporale Zirkulation am Tiermodell

ZIELSETZUNG: In heutiger Zeit beobachtet man in allen Bereichen der Chirurgie den Trend zu weniger traumatischen Operationen im Sinne einer minimalinvasiven Chirurgie. In der Herzchirurgie ist eines dieser Verfahren die OPCAB-Methode, eine Bypassoperation ohne Einsatz der Herz-Lungen-Maschine (off pump coronary artery grafting). Um eventuelle Vorteile dieser Methode auf die Kontraktilität des Herzmuskels zu untersuchen, wurde sie mit einer Bypassoperation unter Verwendung der extrakorporalen Zirkulation verglichen. Dazu wurde ein Leitfähigkeitskatheter verwendet, welcher die linksventrikuläre Funktion mit Hilfe von Druck-Volumen-Beziehungen vor und nach der Operation erfassen kann. METHODEN: 34 Yorkshire Duroc Schweine wurden sternotomiert und anschließend der jeweiligen OP-Methode zugeführt. Der Leitfähigkeitskatheter wurde in den linken Ventrikel eingeführt. Die Kontraktilitätsparameter wurden prä- und postoperativ gemessen. Eine Gruppe (n=11) wurde für eine Stunde einer normothermen extrakorporalen Zirkulation ausgesetzt. Eine zweite Gruppe (n=8) wurde nach der OPCAB-Methode operiert. Die dritte Gruppe (n=15) diente als Kontrollgruppe ohne Operation und extrakorporale Zirkulation. ERGEBNISSE: In der EKZ-Gruppe zeigt sich postoperativ bei allen bestimmten Kontraktilitätsparametern (ESPVR [p=0,01], dP/dt max [p<0,0001] & EF [p<0,0002]) ein signifikanter Kontraktilitätsverlust des Herzmuskels. Hinzu kommt ein signifikanter Abfall des Herzindex [p=0,0004]. In der OPCAB-Gruppe ist kein signifikanter Unterschied bezüglich der ESPVR [p=0,06] sowie der EF [p<0,65] nachzuweisen. Ebenso kommt es nicht zu einem Abfall des Herzindex [p=0,34]. Nicht ganz eindeutig stellt sich in unseren Versuchen das Ergebnis von dP/dt max [p=0,02] dar. Es zeigt einen signifikanten Unterschied, obwohl dieser von der ESPVR sowie der EF nicht wiedergegeben wird. Im intraoperativen Vergleich zeichnet sich insgesamt ein signifikanter Abfall der Herzmuskelkontraktilität während des Anlegens der Anastomose ab. In der Kontrollgruppe ist bei keinem der bestimmten Parameter im prä- und postoperativen Vergleich eine Änderung nachzuweisen (ESPVR [p=0,94], dP/dt max [p=0,75], EF [p=0,65], CI [p=0,78]). SCHLUSSFOLGERUNG: Der Einsatz der extrakorporalen Zirkulation führt zu einer signifikanten postoperativen Einschränkung der linksventrikulären Funktion. Die OPCAB-Methode führt in der sensiblen Phase des Anlegens der Gefäßanastomose ebenfalls zu einer Einschränkung der Herzmuskelkontraktilität, dennoch sind die Auswirkungen im Vergleich zur extrakorporalen Zirkulation deutlich reduziert. Im prä- /postoperativen Vergleich lässt sich kein Kontraktilitätsverlust nachweisen. Es kommt nicht zu einem Abfall der Herzleistung. Diese Studie zeigt somit einen eindeutigen Vorteil der OPCAB-Methode hinsichtlich Kontraktilität und liefert damit ein zusätzliches Argument zur weiteren Verbesserung dieser OP-Methode

Neuroligin 1 is dynamically exchanged at postsynaptic sites

Neuroligins are postsynaptic cell adhesion molecules that associate with presynaptic neurexins. Both factors form a transsynaptic connection, mediate signaling across the synapse, specify synaptic functions, and play a role in synapse formation. Neuroligin dysfunction impairs synaptic transmission, disrupts neuronal networks, and is thought to participate in cognitive diseases. Here we report that chemical treatment designed to induce long-term potentiation or long-term depression (LTD) induces neuroligin 1/3 turnover, leading to either increased or decreased surface membrane protein levels, respectively. Despite its structural role at a crucial transsynaptic position, GFP-neuroligin 1 leaves synapses in hippocampal neurons over time with chemical LTD-induced neuroligin internalization depending on an intact microtubule cytoskeleton. Accordingly, neuroligin 1 and its binding partner postsynaptic density protein-95 (PSD-95) associate with components of the dynein motor complex and undergo retrograde cotransport with a dynein subunit. Transgenic depletion of dynein function in mice causes postsynaptic NLG1/3 and PSD-95 enrichment. In parallel, PSD lengths and spine head sizes are significantly increased, a phenotype similar to that observed upon transgenic overexpression of NLG1 (Dahlhaus et al., 2010). Moreover, application of a competitive PSD-95 peptide and neuroligin 1 C-terminal mutagenesis each specifically alter neuroligin 1 surface membrane expression and interfere with its internalization. Our data suggest the concept that synaptic plasticity regulates neuroligin turnover through active cytoskeleton transport

Completion of neuronal remodeling prompts myelination along developing motor axon branches

Neuronal remodeling and myelination are two fundamental processes during neurodevelopment. How they influence each other remains largely unknown, even though their coordinated execution is critical for circuit function and often disrupted in neuropsychiatric disorders. It is unclear whether myelination stabilizes axon branches during remodeling or whether ongoing remodeling delays myelination. By modulating synaptic transmission, cytoskeletal dynamics, and axonal transport in mouse motor axons, we show that local axon remodeling delays myelination onset and node formation. Conversely, glial differentiation does not determine the outcome of axon remodeling. Delayed myelination is not due to a limited supply of structural components of the axon–glial unit but rather is triggered by increased transport of signaling factors that initiate myelination, such as neuregulin. Further, transport of promyelinating signals is regulated via local cytoskeletal maturation related to activity-dependent competition. Our study reveals an axon branch–specific fine-tuning mechanism that locally coordinates axon remodeling and myelination

Absence of Whisker-Related Pattern Formation in Mice with NMDA Receptors Lacking Coincidence Detection Properties and Calcium Signaling

Precise refinement of synaptic connectivity is the result of activity-dependent mechanisms in which coincidence-dependent calcium signaling by NMDA receptors (NMDARs) under control of the voltage-dependent Mg2+ block might play a special role. In the developing rodent trigeminal system, the pattern of synaptic connections between whisker-specific inputs and their target cells in the brainstem is refined to form functionally and morphologically distinct units (barrelettes). To test the role of NMDA receptor signaling in this process, we introduced the N598R mutation into the native NR1 gene. This leads to the expression of functional NMDARs that are Mg2+ insensitive and Ca2+impermeable. Newborn mice expressing exclusively NR1 N598R-containing NMDARs do not show any whisker-related patterning in the brainstem, whereas the topographic projection of trigeminal afferents and gross brain morphology appear normal. Furthermore, the NR1 N598R mutation does not affect expression levels of NMDAR subunits and other important neurotransmitter receptors. Our results show that coincidence detection by, and/or Ca2+ permeability of, NMDARs is necessary for the development of somatotopic maps in the brainstem and suggest that highly specific signaling underlies synaptic refinement

Single-channel properties of glycine receptors of juvenile rat spinal motoneurones in vitro

An essential step in understanding fast synaptic transmission is to establish the activation mechanism of synaptic receptors. The purpose of this work was to extend our detailed single-channel kinetic characterization of α1β glycine channels from rat recombinant receptors to native channels from juvenile (postnatal day 12–16) rat spinal cord slices. In cell-attached patches from ventral horn neurones, 1 mm glycine elicited clusters of channel openings to a single conductance level (41 ± 1 pS, n=12). This is similar to that of recombinant heteromers. However, fewer than 1 in 100 cell-attached patches from spinal neurones contained glycine channels. Outside-out patches gave a much higher success rate, but glycine channels recorded in this configuration appeared different, in that clusters opened to three conductance levels (28 ± 2, 38 ± 1 and 46 ± 1 pS, n=7, one level per cluster, all levels being detected in each patch). Furthermore, open period properties were different for the different conductances. As a consequence of this, the only recordings suitable for kinetic analysis were the cell-attached ones. Low channel density precluded recording at glycine concentrations other than 1 mm, but the 1 mm data allowed us to estimate the fully bound gating constants by global model fitting of the ‘flip’ mechanism of Burzomato and co-workers. Our results suggest that glycine receptors on ventral horn neurones in the juvenile rat are heteromers and have fast gating, similar to that of recombinant α1β receptors

An Essential Role for the Tetraspanin LHFPL4 in the Cell-Type-Specific Targeting and Clustering of Synaptic GABAReceptors

Inhibitory synaptic transmission requires the targeting and stabilization of GABAA receptors (GABAARs) at synapses. The mechanisms responsible remain poorly understood, and roles for transmembrane accessory proteins have not been established. Using molecular, imaging, and electrophysiological approaches, we identify the tetraspanin LHFPL4 as a critical regulator of postsynaptic GABAAR clustering in hippocampal pyramidal neurons. LHFPL4 interacts tightly with GABAAR subunits and is selectively enriched at inhibitory synapses. In LHFPL4 knockout mice, there is a dramatic cell-type-specific reduction in GABAAR and gephyrin clusters and an accumulation of large intracellular gephyrin aggregates in vivo. While GABAARs are still trafficked to the neuronal surface in pyramidal neurons, they are no longer localized at synapses, resulting in a profound loss of fast inhibitory postsynaptic currents. Hippocampal interneuron currents remain unaffected. Our results establish LHFPL4 as a synapse-specific tetraspanin essential for inhibitory synapse function and provide fresh insights into the molecular make-up of inhibitory synapses

Myosin VI Drives Clathrin-Mediated AMPA Receptor Endocytosis to Facilitate Cerebellar Long-Term Depression

Myosin VI is an actin-based cytoskeletal motor implicated in various steps of membrane trafficking. Here, we investigated whether this myosin is crucial for synaptic function and plasticity in neurons. We find that myosin VI localizes at cerebellar parallel fiber to Purkinje cell synapses and that the myosin is indispensable for long-term depression of AMPA-receptor-mediated synaptic signal transmission at this synapse. Moreover, direct visualization of GluA2-containing AMPA receptors in Purkinje cells reveals that the myosin drives removal of AMPA receptors from the surface of dendritic spines in an activity-dependent manner. Co-immunoprecipitation and super-resolution microscopy indicate that specifically the interaction of myosin VI with the clathrin adaptor component alpha-adaptin is important during long-term depression. Together, these data suggest that myosin VI directly promotes clathrin-mediated endocytosis of AMPA receptors in Purkinje cells to mediate cerebellar long-term depression. Our results provide insights into myosin VI function and the molecular mechanisms underlying synaptic plasticity

Collybistin and gephyrin are novel components of the eukaryotic translation initiation factor 3 complex

<p>Abstract</p> <p>Background</p> <p>Collybistin (CB), a neuron-specific guanine nucleotide exchange factor, has been implicated in targeting gephyrin-GABA_Areceptors clusters to inhibitory postsynaptic sites. However, little is known about additional CB partners and functions.</p> <p>Findings</p> <p>Here, we identified the p40 subunit of the eukaryotic translation initiation factor 3 (eIF3H) as a novel binding partner of CB, documenting the interaction in yeast, non-neuronal cell lines, and the brain. In addition, we demonstrated that gephyrin also interacts with eIF3H in non-neuronal cells and forms a complex with eIF3 in the brain.</p> <p>Conclusions</p> <p>Together, our results suggest, for the first time, that CB and gephyrin associate with the translation initiation machinery, and lend further support to the previous evidence that gephyrin may act as a regulator of synaptic protein synthesis.</p

Isoform Heterogeneity of the Human Gephyrin Gene (GPHN), Binding Domains to the Glycine Receptor, and Mutation Analysis in Hyperekplexia

Gephyrin (GPHN) is an organizational protein that clusters and localizes the inhibitory glycine (GlyR) and GABAA receptors to the microtubular matrix of the neuronal postsynaptic membrane. Mice deficient in gephyrin develop a hereditary molybdenum cofactor deficiency and a neurological phenotype that mimics startle disease (hyperekplexia). This neuromotor disorder is associated with mutations in the GlyR α1 and β subunit genes (GLRA1 and GLRB). Further genetic heterogeneity is suspected, and we hypothesized that patients lacking mutations in GLRA1 and GLRB might have mutations in the gephyrin gene (GPHN). In addition, we adopted a yeast two-hybrid screen, using the GlyR β subunit intracellular loop as bait, in an attempt to identify further GlyR-interacting proteins implicated in hyperekplexia. Gephyrin cDNAs were isolated, and subsequent RT-PCR analysis from human tissues demonstrated the presence of five alternatively spliced GPHN exons concentrated in the central linker region of the gene. This region generated 11 distinct GPHN transcript isoforms, with 10 being specific to neuronal tissue. Mutation analysis of GPHN exons in hyperekplexia patients revealed a missense mutation (A28T) in one patient causing an amino acid substitution (N10Y). Functional testing demonstrated that GPHNN10Y does not disrupt GlyR-gephyrin interactions or collybistininduced cell-surface clustering. We provide evidence that GlyR-gephyrin binding is dependent on the presence of an intact C-terminal MoeA homology domain. Therefore, the N10Y mutation and alternative splicing of GPHN transcripts do not affect interactions with GlyRs but may affect other interactions with the cytoskeleton or gephyrin accessory proteins