Karl Heinz Smalla
Research interests
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InterestsNeurobiology, Neurobiochemistry, Neurodegenerative Diseases, Neuropharmacology
Publications
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5.33Impact points
The cell adhesion molecule neuroplastin-65 is a novel interaction partner of γ- Aminobutyric acid type A receptors.
The Journal of biological chemistry. 03/2012;
γ-Aminobutyric acid(A) (GABA)(A) receptors are pentameric ligand-gated ion channels that mediate fast inhibition in the central nervous system. Depending on their subunit composition, these receptors exhibit distinct pharmacological properties and differ in their ability to interact with proteins in... [more] γ-Aminobutyric acid(A) (GABA)(A) receptors are pentameric ligand-gated ion channels that mediate fast inhibition in the central nervous system. Depending on their subunit composition, these receptors exhibit distinct pharmacological properties and differ in their ability to interact with proteins involved in receptor anchoring at synaptic or extra-synaptic sites. Whereas GABA(A) receptors containing α1, α2, or α3 subunits are mainly located synaptically where they interact with the sub-membraneous scaffolding protein gephyrin, receptors containing α5 subunits are predominantly found extra-synaptically and seem to interact with radixin for anchorage. Neuroplastin is a cell adhesion molecule of the immunoglobulin superfamily that is involved in hippocampal synaptic plasticity. Our results reveal that neuroplastin and GABA(A) receptors can be co-purified from rat brain and exhibit a direct physical interaction as demonstrated by co-precipitation and Forster Resonance Energy Transfer (FRET) analysis in a heterologous expression system. The brain-specific isoform neuroplastin-65 co-localizes with GABA(A) receptors as shown in brain sections as well as in neuronal cultures, and such complexes can either contain gephyrin or be devoid of gephyrin. Neuroplastin-65 specifically co-localizes with α1 or α2, but not with α3 subunits at GABAergic synapses. In addition, neuroplastin-65 also co-localizes with GABA(A) receptor α5 subunits at extra-synaptic sites. Down-regulation of neuroplastin-65 by shRNA causes a loss of GABA(A) receptor α2 subunits at GABAergic synapses.These results suggest that neuroplastin-65 can co-localize with a subset of GABA(A) receptor subtypes and might contribute to a novel anchoring mechanism confining GABA(A) receptors to particular synaptic or extra-synaptic sites, thus affecting receptor mobility and synaptic strength.
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3.42Impact points
Dopamine modulates memory consolidation of discrimination learning in the auditory cortex.
The European journal of neuroscience. 03/2012; 35(5):763-74.
In Mongolian gerbils, the auditory cortex is critical for discriminating rising vs. falling frequency-modulated tones. Based on our previous studies, we hypothesized that dopaminergic inputs to the auditory cortex during and shortly after acquisition of the discrimination strategy control long-term ... [more] In Mongolian gerbils, the auditory cortex is critical for discriminating rising vs. falling frequency-modulated tones. Based on our previous studies, we hypothesized that dopaminergic inputs to the auditory cortex during and shortly after acquisition of the discrimination strategy control long-term memory formation. To test this hypothesis, we studied frequency-modulated tone discrimination learning of gerbils in a shuttle box GO/NO-GO procedure following differential treatments. (i) Pre-exposure of gerbils to the frequency-modulated tones at 1 day before the first discrimination training session severely impaired the accuracy of the discrimination acquired in that session during the initial trials of a second training session, performed 1 day later. (ii) Local injection of the D1/D5 dopamine receptor antagonist SCH-23390 into the auditory cortex after task acquisition caused a discrimination deficit of similar extent and time course as with pre-exposure. This effect was dependent on the dose and time point of injection. (iii) Injection of the D1/D5 dopamine receptor agonist SKF-38393 into the auditory cortex after retraining caused a further discrimination improvement at the beginning of subsequent sessions. All three treatments, which supposedly interfered with dopamine signalling during conditioning and/or retraining, had a substantial impact on the dynamics of the discrimination performance particularly at the beginning of subsequent training sessions. These findings suggest that auditory-cortical dopamine activity after acquisition of a discrimination of complex sounds and after retrieval of weak frequency-modulated tone discrimination memory further improves memory consolidation, i.e. the correct association of two sounds with their respective GO/NO-GO meaning, in support of future memory recall.
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3.23Impact points
Accurate quantification of water-macromolecule exchange induced frequency shift: Effects of reference substance.
Magnetic resonance in medicine : official journal of the Society of Magnetic Resonance in Medicine / Society of Magnetic Resonance in Medicine. 02/2012;
Water-macromolecule exchange induces a bulk water frequency shift contributing to the contrast in phase imaging. For separating the effects of the water-macromolecule exchange and the macromolecule susceptibility, appropriate internal or external references are needed. In this study, two internal re... [more] Water-macromolecule exchange induces a bulk water frequency shift contributing to the contrast in phase imaging. For separating the effects of the water-macromolecule exchange and the macromolecule susceptibility, appropriate internal or external references are needed. In this study, two internal reference compounds, 2,2,3,3-tetradeuterio-3-trimethylsilyl-propionate (TMSP) and 1,4-dioxane, were used to study the macromolecule-dependent water frequency shift in a bovine serum albumin (BSA)-water system in detail. For TMSP, the water-macromolecule exchange shift depended on both the BSA and the reference concentration and stabilized to a value of 0.025 ppm/mM (298 K, TMSP concentrations > 30 mM). For dioxane, the dependency of the water-macromolecule exchange shift on the BSA concentration is independent of dioxane at low concentrations. The resulting shift was smaller (0.009 ppm/mM) when compared with using higher TMSP concentrations as reference. This discrepancy might be due to additional dioxane-water interactions. Measurements with an external chloroform reference in a coaxial geometry showed a shift of -0.013 ppm/mM resulting from the opposing effects of macromolecules in water exchange-induced shift and diamagnetic susceptibility shift. All these effects should be considered in the interpretation of tissue phase contrast. From the experimental data, the equilibrium binding constant between BSA and TMSP has been quantified to be K(d) = 1.3 ± 0.4, and the estimated number of interaction sites for BSA is 12.7 ± 2.6. Magn Reson Med, 2012. © 2012 Wiley Periodicals, Inc.
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4.00Impact points
Homeostatic NMDA receptor down-regulation via brain derived neurotrophic factor and nitric oxide-dependent signalling in cortical but not in hippocampal neurons.
Journal of neurochemistry. 06/2011; 118(5):760-72.
Nitric oxide (NO) has been proposed to down-regulate NMDA receptors (NMDA-Rs) in a homeostatic manner. However, NMDA-R-dependent NO synthesis also can cause excitotoxic cell death. Using bicuculline-stimulated hippocampal and cortical cell cultures, we have addressed the role of the brain-derived ne... [more] Nitric oxide (NO) has been proposed to down-regulate NMDA receptors (NMDA-Rs) in a homeostatic manner. However, NMDA-R-dependent NO synthesis also can cause excitotoxic cell death. Using bicuculline-stimulated hippocampal and cortical cell cultures, we have addressed the role of the brain-derived neurotrophic factor-NO pathway in NMDA-R down-regulation. This pathway protected cortical cells from NMDA-induced death and led to NMDA-R inhibition. In contrast, no evidence was gained for the presence of this protective pathway in hippocampal neurons, in which NMDA-induced NO synthesis was confirmed to be toxic. Therefore, opposing effects of NO depended on the activation of different signalling pathways. The pathophysiological relevance of this observation was investigated in synaptosomes and post-synaptic densities isolated from rat hippocampi and cerebral cortices following kainic acid-induced status epilepticus. In cortical, but not in hippocampal synaptosomes, brain-derived neurotrophic factor induced NO synthesis and inhibited NMDA-R currents present in isolated post-synaptic densities. In conclusion, we identified a NO-dependent homeostatic response in the rat cerebral cortex induced by elevated activity. A low performance of this pathway in brain areas including the hippocampus may be related to their selective vulnerability in pathologies such as temporal lobe epilepsy.
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6.14Impact points
TRPV1 acts as a synaptic protein and regulates vesicle recycling.
Journal of cell science. 06/2010; 123(Pt 12):2045-57.
Electrophysiological studies demonstrate that transient receptor potential vanilloid subtype 1 (TRPV1) is involved in neuronal transmission. Although it is expressed in the peripheral as well as the central nervous system, the questions remain whether TRPV1 is present in synaptic structures and whet... [more] Electrophysiological studies demonstrate that transient receptor potential vanilloid subtype 1 (TRPV1) is involved in neuronal transmission. Although it is expressed in the peripheral as well as the central nervous system, the questions remain whether TRPV1 is present in synaptic structures and whether it is involved in synaptic processes. In the present study we gathered evidence that TRPV1 can be detected in spines of cortical neurons, that it colocalizes with both pre- and postsynaptic proteins, and that it regulates spine morphology. Moreover, TRPV1 is also present in biochemically prepared synaptosomes endogenously. In F11 cells, a cell line derived from dorsal-root-ganglion neurons, TRPV1 is enriched in the tips of elongated filopodia and also at sites of cell-cell contact. In addition, we also detected TRPV1 in synaptic transport vesicles, and in transport packets within filopodia and neurites. Using FM4-64 dye, we demonstrate that recycling and/or fusion of these vesicles can be rapidly modulated by TRPV1 activation, leading to rapid reorganization of filopodial structure. These data suggest that TRPV1 is involved in processes such as neuronal network formation, synapse modulation and release of synaptic transmitters.
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3.48Impact points
Congenital lack of nNOS impairs long-term social recognition memory and alters the olfactory bulb proteome.
Neurobiology of learning and memory. 07/2009;
The gaseous neurotransmitter nitric oxide (NO), synthesized by the enzyme neuronal nitric oxide synthase (nNOS), is thought to play a major role in the modulation of memory. We tested adult nNOS-deficient and wild-type mice for their recognition memory abilities in the social discrimination paradigm... [more] The gaseous neurotransmitter nitric oxide (NO), synthesized by the enzyme neuronal nitric oxide synthase (nNOS), is thought to play a major role in the modulation of memory. We tested adult nNOS-deficient and wild-type mice for their recognition memory abilities in the social discrimination paradigm, which is based on olfactory cues. Subsequently, proteomic investigation of the olfactory bulbs of both genotypes were performed under basal conditions and 6 h after learning, i.e., during the consolidation of long-term memory. Short-term and intermediate-term recognition memory was normal in nNOS-deficient mice. However, unlike wild-type mice, nNOS-deficient mice failed to consolidate an olfactory cued long-term recognition memory. Proteomic analysis revealed changes in glycolytic enzymes (e.g., fructose-bisphosphate aldolase C, glyceraldehyde-3-phosphate dehydrogenase), voltage-dependent anion selective channels 1 and 2, alpha-synuclein, F-actin-interacting proteins (e.g., neuronal protein 25/transgelin 3), proteins of the ubiquitin proteasome system, and heterogeneous nuclear ribonucleoproteins implicated in the regulation of messenger RNA trafficking, stability and translation. Our data suggest that, in the mouse, NO of nNOS origin is critically involved in the regulation of protein synthesis-dependent olfactory long-term memory consolidation within relevant brain structures including the olfactory bulb.
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9.43Impact points
Calneurons provide a calcium threshold for trans-Golgi network to plasma membrane trafficking.
Proceedings of the National Academy of Sciences of the United States of America. 06/2009;
Phosphatidylinositol 4-OH kinase IIIbeta (PI-4Kbeta) is involved in the regulated local synthesis of phospholipids that are crucial for trans-Golgi network (TGN)-to-plasma membrane trafficking. In this study, we show that the calcium sensor proteins calneuron-1 and calneuron-2 physically associate w... [more] Phosphatidylinositol 4-OH kinase IIIbeta (PI-4Kbeta) is involved in the regulated local synthesis of phospholipids that are crucial for trans-Golgi network (TGN)-to-plasma membrane trafficking. In this study, we show that the calcium sensor proteins calneuron-1 and calneuron-2 physically associate with PI-4Kbeta, inhibit the enzyme profoundly at resting and low calcium levels, and negatively interfere with Golgi-to-plasma membrane trafficking. At high calcium levels this inhibition is released and PI-4Kbeta is activated via a preferential association with neuronal calcium sensor-1 (NCS-1). In accord to its supposed function as a filter for subthreshold Golgi calcium transients, neuronal overexpression of calneuron-1 enlarges the size of the TGN caused by a build-up of vesicle proteins and reduces the number of axonal Piccolo-Bassoon transport vesicles, large dense core vesicles that carry a set of essential proteins for the formation of the presynaptic active zone during development. A corresponding protein knockdown has the opposite effect. The opposing roles of calneurons and NCS-1 provide a molecular switch to decode local calcium transients at the Golgi and impose a calcium threshold for PI-4Kbeta activity and vesicle trafficking.
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2.72Impact points
Altered Postsynaptic-Density-Levels of Caldendrin in the Para-Chloroamphetamine-Induced Serotonin Syndrome but not in the Rat Ketamine Model of Psychosis.
Neurochemical research. 03/2009;
Caldendrin is a synaptic calcium sensor protein that is tightly associated with the postsynaptic density (PSD). Previous work has shown that the association of the protein with the synapse is highly dynamic and is increased in an activity-dependent manner. In the present study the caldendrin-associa... [more] Caldendrin is a synaptic calcium sensor protein that is tightly associated with the postsynaptic density (PSD). Previous work has shown that the association of the protein with the synapse is highly dynamic and is increased in an activity-dependent manner. In the present study the caldendrin-association with the postsynaptic cytomatrix was analyzed in animal models of psychosis and drug abuse induced neurotoxicity. Subchronic administration of the N-methyl-D: -aspartate (NMDA)-receptor antagonist ketamine, serving as a model of NMDA-receptor hypofunction and schizophrenia showed no significant effect on the PSD-levels of caldendrin, indicating that NMDA-receptor activity is not required to keep caldendrin at the synapse. However, administration of high doses of the serotonergic neurotoxin p-chloroamphetamine (PCA) lead to significant changes in the association of caldendrin with the PSD. These results underscore the dynamic association of caldendrin with the PSD and suggest a role of this synaptic calcium sensor in the PCA-induced serotonin syndrome.
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1.81Impact points
Scaffolding proteins in highly purified rat olfactory cilia membranes.
Neuroreport. 08/2008; 19(11):1123-1126.
Odour-mediated signal transduction is a complex process that occurs in the cilia of olfactory sensory neurons. To gain insight in to the molecular organization of the odour transduction machinery, we developed a procedure to purify olfactory cilia membranes by differential centrifugation of rat olfa... [more] Odour-mediated signal transduction is a complex process that occurs in the cilia of olfactory sensory neurons. To gain insight in to the molecular organization of the odour transduction machinery, we developed a procedure to purify olfactory cilia membranes by differential centrifugation of rat olfactory epithelium extracts. We tested whether known scaffolding proteins that might participate in the assembly of the complex chemotransduction apparatus are present in the purified membrane fraction. Utilizing immunoblotting and immunohistochemistry, we show that the multidomain scaffolding proteins ProSAP/Shanks and calcium/calmodulin-dependent serine protein kinase CASK are present in the olfactory cilia. Ion channels involved in chemotransduction could be reconstituted into planar lipid bilayers for electrophysiological recordings. Our procedure should allow the identification of further chemotransduction-related proteins.
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15.05Impact points
A comparison of the synaptic proteome in human chronic schizophrenia and rat ketamine psychosis suggest that prohibitin is involved in the synaptic pathology of schizophrenia.
Molecular psychiatry. 06/2008;
Many studies in recent years suggest that schizophrenia is a synaptic disease that crucially involves a hypofunction of N-methyl-D-aspartate receptor-mediated signaling. However, at present it is unclear how these pathological processes are reflected in the protein content of the synapse. We have em... [more] Many studies in recent years suggest that schizophrenia is a synaptic disease that crucially involves a hypofunction of N-methyl-D-aspartate receptor-mediated signaling. However, at present it is unclear how these pathological processes are reflected in the protein content of the synapse. We have employed two-dimensional gel electrophoresis in conjunction with mass spectrometry to characterize and compare the synaptic proteomes of the human left dorsolateral prefrontal cortex in chronic schizophrenia and of the cerebral cortex of rats treated subchronically with ketamine. We found consistent changes in the synaptic proteomes of human schizophrenics and in rats with induced ketamine psychosis compared to controls. However, commonly regulated proteins between both groups were very limited and only prohibitin was found upregulated in both chronic schizophrenia and the rat ketamine model. Prohibitin, however, could be a new potential marker for the synaptic pathology of schizophrenia and might be causally involved in the disease process.Molecular Psychiatry advance online publication, 27 May 2008; doi:10.1038/mp.2008.60.
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6.98Impact points
Dopaminergic Modulation of Auditory Cortex-Dependent Memory Consolidation through mTOR.
Cerebral cortex (New York, N.Y. : 1991). 04/2008;
Previous studies in the auditory cortex of Mongolian gerbils on discrimination learning of the direction of frequency-modulated tones (FMs) revealed that long-term memory formation involves activation of the dopaminergic system, activity of the protein kinase mammalian target of rapamycin (mTOR), an... [more] Previous studies in the auditory cortex of Mongolian gerbils on discrimination learning of the direction of frequency-modulated tones (FMs) revealed that long-term memory formation involves activation of the dopaminergic system, activity of the protein kinase mammalian target of rapamycin (mTOR), and protein synthesis. This led to the hypothesis that the dopaminergic system might modulate memory formation via regulation of mTOR, which is implicated in translational control. Here, we report that the D1/D5 dopamine receptor agonist SKF-38393 substantially improved gerbils' FM discrimination learning when administered systemically or locally into the auditory cortex shortly before, shortly after, or 1 day before conditioning. Although acquisition performance during initial training was normal, the discrimination of FMs was enhanced during retraining performed hours or days after agonist injection compared with vehicle-injected controls. The D1/D5 receptor antagonist SCH-23390, the mTOR inhibitor rapamycin, and the protein synthesis blocker anisomycin suppressed this effect. By immunohistochemistry, D1 dopamine receptors were identified in the gerbil auditory cortex predominantly in the infragranular layers. Together, these findings suggest that in the gerbil auditory cortex dopaminergic inputs regulate mTOR-mediated, protein synthesis-dependent mechanisms, thus controlling for hours or days the consolidation of memory required for the discrimination of complex auditory stimuli.
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12.92Impact points
Caldendrin-Jacob: a protein liaison that couples NMDA receptor signalling to the nucleus.
PLoS biology. 03/2008; 6(2):e34.
NMDA (N-methyl-D-aspartate) receptors and calcium can exert multiple and very divergent effects within neuronal cells, thereby impacting opposing occurrences such as synaptic plasticity and neuronal degeneration. The neuronal Ca2+ sensor Caldendrin is a postsynaptic density component with high simil... [more] NMDA (N-methyl-D-aspartate) receptors and calcium can exert multiple and very divergent effects within neuronal cells, thereby impacting opposing occurrences such as synaptic plasticity and neuronal degeneration. The neuronal Ca2+ sensor Caldendrin is a postsynaptic density component with high similarity to calmodulin. Jacob, a recently identified Caldendrin binding partner, is a novel protein abundantly expressed in limbic brain and cerebral cortex. Strictly depending upon activation of NMDA-type glutamate receptors, Jacob is recruited to neuronal nuclei, resulting in a rapid stripping of synaptic contacts and in a drastically altered morphology of the dendritic tree. Jacob's nuclear trafficking from distal dendrites crucially requires the classical Importin pathway. Caldendrin binds to Jacob's nuclear localization signal in a Ca2+-dependent manner, thereby controlling Jacob's extranuclear localization by competing with the binding of Importin-alpha to Jacob's nuclear localization signal. This competition requires sustained synapto-dendritic Ca2+ levels, which presumably cannot be achieved by activation of extrasynaptic NMDA receptors, but are confined to Ca2+ microdomains such as postsynaptic spines. Extrasynaptic NMDA receptors, as opposed to their synaptic counterparts, trigger the cAMP response element-binding protein (CREB) shut-off pathway, and cell death. We found that nuclear knockdown of Jacob prevents CREB shut-off after extrasynaptic NMDA receptor activation, whereas its nuclear overexpression induces CREB shut-off without NMDA receptor stimulation. Importantly, nuclear knockdown of Jacob attenuates NMDA-induced loss of synaptic contacts, and neuronal degeneration. This defines a novel mechanism of synapse-to-nucleus communication via a synaptic Ca2+-sensor protein, which links the activity of NMDA receptors to nuclear signalling events involved in modelling synapto-dendritic input and NMDA receptor-induced cellular degeneration.
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4.46Impact points
A reduced number of cortical neurons show increased Caldendrin protein levels in chronic schizophrenia.
Schizophrenia research. 12/2007; 96(1-3):246-56.
Caldendrin is a neuronal calcium sensor protein that is tightly associated with the postsynaptic density (PSD) of excitatory synapses. It has an established role in synapto-dendritic Ca(2+)-signaling as a multifunctional regulator of intracellular Ca(2+) levels. Previous work has shown that expressi... [more] Caldendrin is a neuronal calcium sensor protein that is tightly associated with the postsynaptic density (PSD) of excitatory synapses. It has an established role in synapto-dendritic Ca(2+)-signaling as a multifunctional regulator of intracellular Ca(2+) levels. Previous work has shown that expression levels of protein components involved in signaling processes at excitatory synapses are significantly altered in the brains of schizophrenic patients. Furthermore, it is widely accepted that synaptic pathology associated with the glutamatergic N-methyl-d-aspartate (NMDA) receptor is a feature of the disease. Here we report that in postmortem brains of chronic schizophrenics (N: 12) as compared to age-and sex-matched controls (N: 12) the number of Caldendrin-immunoreactive neurons are significantly reduced in the left dorsolateral prefrontal cortex, a brain region prominently associated with schizophrenia. Less dramatic changes were observed in other cortical regions. However, despite the reduced number of immunoreactive neurons, absolute Caldendrin protein levels were elevated and no change in Caldendrin PSD-levels were observed as compared to the left dorsolateral prefrontal cortex in the normal human brain. Thus, synapto-dendritic Ca(2+)-signaling via Caldendrin is altered in schizophrenic patients by a redistribution of the protein into a lower number of pyramidal neurons, which express higher Caldendrin levels. Since Caldendrin is a multivalent regulator of voltage dependent Ca(2+)-channels and Ca(2+)-release channels the loss of Caldendrin mediated synapto-dendritic Ca(2+)-signaling processes in some neurons together with its concurrent upregulation in others should profoundly change their synapto-dendritic Ca(2+)-signaling. These observations add to existing evidence for a de-regulation of neuronal Ca(2+)-signaling in schizophrenia.
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4.00Impact points
Antagonistic effects of TrkB and p75(NTR) on NMDA receptor currents in post-synaptic densities transplanted into Xenopus oocytes.
Journal of neurochemistry. 06/2007; 101(6):1672-84.
Brain-derived neurotrophic factor (BDNF) and its receptor TrkB are essential regulators of synaptic function in the adult CNS. A TrkB-mediated effect at excitatory synapses is enhancement of NMDA receptor (NMDA-R)-mediated currents. Recently, opposing effects of TrkB and the pan-neurotrophin recepto... [more] Brain-derived neurotrophic factor (BDNF) and its receptor TrkB are essential regulators of synaptic function in the adult CNS. A TrkB-mediated effect at excitatory synapses is enhancement of NMDA receptor (NMDA-R)-mediated currents. Recently, opposing effects of TrkB and the pan-neurotrophin receptor p75(NTR) on long-term synaptic depression and long-term potentiation have been reported in the hippocampus. To further study the regulation of NMDA-Rs by neurotrophin receptors in their native protein environment, we micro-transplanted rat forebrain post-synaptic densities (PSDs) into Xenopus oocytes. One-minute incubations of oocytes with BDNF led to dual effects on NMDA-R currents: either TrkB-dependent potentiation or TrkB-independent inhibition were observed. Pro-nerve growth factor, a ligand for p75(NTR) but not for TrkB, produced a reversible, dose-dependent, TrkB-independent and p75(NTR)-dependent inhibition of NMDA-Rs. Fractionation experiments showed that p75(NTR) is highly enriched in the PSD protein fraction. Immunoprecipitation and pull-down experiments further revealed that p75(NTR) is a core component of the PSD, where it interacts with the PDZ3 domain of the scaffolding protein SAP90/PSD-95. Our data provide striking evidence for a rapid inhibitory effect of p75(NTR) on NMDA-R currents that antagonizes TrkB-mediated NMDA-R potentiation. These opposing mechanisms might be present in a large proportion of forebrain synapses and may contribute importantly to synaptic plasticity.
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2.73Impact points
Splice-isoform specific immunolocalization of neuronal nitric oxide synthase in mouse and rat brain reveals that the PDZ-complex-building nNOSalpha beta-finger is largely exposed to antibodies.
Developmental neurobiology. 04/2007; 67(4):422-37.
Knock out mice deficient for the splice-isoform alphaalpha of neuronal nitric oxide synthase (nNOSalphaalpha) display residual nitric oxide synthase activity and immunosignal. To attribute this signal to the two minor neuronal nitric oxide synthase splice variants, betabeta and gammagamma, we genera... [more] Knock out mice deficient for the splice-isoform alphaalpha of neuronal nitric oxide synthase (nNOSalphaalpha) display residual nitric oxide synthase activity and immunosignal. To attribute this signal to the two minor neuronal nitric oxide synthase splice variants, betabeta and gammagamma, we generated isoform-specific anti-peptide antibodies against the nNOSalphaalpha specific betabeta-finger motif involved in PDZ domain scaffolding and the nNOSbetabeta specific N-terminus. The nNOSalphaalpha betabeta-finger-specific antibody clearly recognized the 160-kDa band of recombinant nNOSalphaalpha on Western blots. Using immunocytochemistry, this antibody displayed, in rats and wild-type mice, a labeling pattern similar to but not identical with that obtained using a commercial pan-nNOS antibody. This similarity indicates that the majority of immunocytochemically detectable nNOS is not likely to be complexed with PDZ-domain proteins via the betabeta-finger motif. This conclusion was confirmed by the inhibition of PSD-95/nNOS interaction by the nNOSalphaalpha betabeta-finger antibody in pull-down assays. By contrast, nNOSalphaalpha betabeta-finger labeling was clearly reduced in hippocampal and cortical neuropil areas enriched in NMDA receptor complex containing spine synapses. In nNOSalphaalpha knock out mice, nNOSalphaalpha was not detectable, whereas the pan-nNOS antibody showed a distinct labeling of cell bodies throughout the brain, most likely reflecting betabeta/gammagamma-isoforms in these cells. The nNOSbetabeta antibody clearly detected bacterial expressed nNOSbetabeta fusion protein and nNOSbetabeta in overexpressing HEK cells by Western blotting. Immunocytochemically, individual cell bodies in striatum, cerebral cortex, and in some brain stem nuclei were labeled in knock out but not in wild-type mice, indicating an upregulation of nNOSbetabeta in nNOSalphaalpha deficient animals.
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4.73Impact points
The effects of p-chloroamphetamine, methamphetamine and 3,4-methylenedioxymethamphetamine (ecstasy) on the gene expression of cytoskeletal proteins in the rat brain.
Addiction biology. 03/2007; 12(1):69-80.
Repeated administration of beta-phenylalkylamines is known to produce neuronal changes in the central and peripheral nervous systems of mammals. It is suggested that various components of the cytoskeleton undergo profound alterations after amphetamine use and misuse, contributing to behavioral chang... [more] Repeated administration of beta-phenylalkylamines is known to produce neuronal changes in the central and peripheral nervous systems of mammals. It is suggested that various components of the cytoskeleton undergo profound alterations after amphetamine use and misuse, contributing to behavioral changes and neurotoxicity. Here we studied the expression of microtubule-associated protein 2 (MAP2) and beta-actin after repeated intraperitoneal applications with equimolar doses of p-chloroamphetamine (PCA), methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA) in the brain of male Wistar rats. Effective (molecular) pharmacological doses (ED) were derived and used for the calculation of (molecular) pharmacological indices (PI). Besides clear but different dose-response curves on the toxicity of the drugs, in situ hybridization and Western blot analysis revealed that repeated administration of these compounds resulted in different substance- and dose-dependent changes in MAP2 gene expression, e.g. in the frontoparietal somatosensoric cortex. In contrast, the expression of beta-actin was not influenced by any of the compounds at the dose levels tested. Lethal doses were determined with 2.1 (PCA), >5.1 (METH) and 8.4 mg/kg/day (MDMA). Linear and non-linear repeat-dose lethality was observed for MDMA and PCA, respectively, whereas METH was non-lethal in the dose range used. Values for ED(MAP2) were 0.3, 0.52 and >16.8 mg/kg/day, and therefore those for PI(MAP2) were 20, 4, and 0.5 for METH, PCA and MDMA, respectively. Although the results on mortality did not reflect changes in MAP2 gene expression, they suggest a remarkable difference for those amphetamines without substituents or with a halogen atom at the paraposition of the benzene ring, such as METH or PCA, when compared with MDMA-like substances.
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2.46Impact points
The immunolocalization of the synaptic glycoprotein neuroplastin differs substantially between the human and the rodent brain.
Brain research. 03/2007; 1134(1):107-12.
Neuroplastin is a cell adhesion molecule of the immunoglobulin superfamily that exists in two splice isoforms, np65/np55, and that was reported to play a prominent role in synaptic plasticity processes. The splice isoform np65 associates with synapses in an activity-dependent manner and has been sho... [more] Neuroplastin is a cell adhesion molecule of the immunoglobulin superfamily that exists in two splice isoforms, np65/np55, and that was reported to play a prominent role in synaptic plasticity processes. The splice isoform np65 associates with synapses in an activity-dependent manner and has been shown to play a role for the induction of hippocampal long-term potentiation in rodents. We have therefore analyzed the distribution of neuroplastins in human brain. Neuroplastin is present in many neuronal cell types of the forebrain and cerebellum and immunoreactive label covers the cell soma, neurites and also puncta in the neuropil were visible. Interestingly, we found some remarkable species differences in the expression patterns of neuroplastins between the human and the rodent brain. In human brain np65 is prominently present in cerebellum while np55 is the predominant isoform in mouse and rat cerebellum. Moreover, the parasagittal stripe-type of staining seen with np55 in mouse cerebellum is not found in human brain. In addition we found no segregation of np65 immunolabel in hippocampal subregions like it was reported previously for the rat. These results might indicate different cellular functions of the molecule in different species.
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4.66Impact points
Neuronal Ca2+ signaling via caldendrin and calneurons.
Biochimica et biophysica acta. 12/2006; 1763(11):1229-37.
The calcium sensor protein caldendrin is abundantly expressed in neurons and is thought to play an important role in different aspects of synapto-dendritic Ca2+ signaling. Caldendrin is highly abundant in the postsynaptic density of a subset of excitatory synapses in brain and its distinct localizat... [more] The calcium sensor protein caldendrin is abundantly expressed in neurons and is thought to play an important role in different aspects of synapto-dendritic Ca2+ signaling. Caldendrin is highly abundant in the postsynaptic density of a subset of excitatory synapses in brain and its distinct localization raises several decisive questions about its function. Previous work suggests that caldendrin is tightly associated with Ca2+ - and Ca2+ release channels and might be involved in different aspects of the organization of the postsynaptic scaffold as well as with synapse-to-nucleus communication. In this report we introduce two new EF-hand calcium sensor proteins termed calneurons that apart from calmodulin represent the closest homologues of caldendrin in brain. Calneurons have a different EF-hand organization than other calcium sensor proteins, are prominently expressed in neurons and will presumably bind Ca2+ with higher affinity than caldendrin. Despite some significant structural differences it is conceivable that they are involved in similar Ca2+ regulated processes like caldendrin and neuronal calcium sensor proteins.
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5.33Impact points
ProSAP-interacting protein 1 (ProSAPiP1), a novel protein of the postsynaptic density that links the spine-associated Rap-Gap (SPAR) to the scaffolding protein ProSAP2/Shank3.
The Journal of biological chemistry. 06/2006; 281(19):13805-16.
ProSAPs/Shanks are a family of proteins that have a major scaffolding function for components of the postsynaptic density (PSD) of excitatory brain synapses. Members of the family harbor a variety of domains for protein-protein interactions, one of which is a unique PDZ domain that differs significa... [more] ProSAPs/Shanks are a family of proteins that have a major scaffolding function for components of the postsynaptic density (PSD) of excitatory brain synapses. Members of the family harbor a variety of domains for protein-protein interactions, one of which is a unique PDZ domain that differs significantly from those of other proteins. We have identified a novel binding partner for this PDZ domain, termed ProSAPiP1, that is highly enriched in the PSD and shares significant sequence homology with the PSD protein PSD-Zip70. Both molecules code for a Fez1 domain that can be found in a total of four related proteins. ProSAPiP1 is widely expressed in rat brain and co-localizes with ProSAP2/Shank3 in excitatory spines and synapses. ProSAP2/Shank3 co-immunoprecipitates with ProSAPiP1 but not with PSD-Zip70. Both proteins, however, bind and recruit SPAR to synapses with a central coiled-coil region that harbors a leucine zipper motif. This region is also responsible for homo- and heteromultimerization of ProSAPiP1 and PSD-Zip70. Thus, ProSAPiP1 and PSD-Zip70 are founders of a novel family of scaffolding proteins, the "Fezzins," which adds further complexity to the organization of the PSD protein network.
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3.57Impact points
Brevican-containing perineuronal nets of extracellular matrix in dissociated hippocampal primary cultures.
Molecular and cellular neurosciences. 05/2006; 31(4):774-84.
Perineuronal nets (PNN) are specialized extracellular matrix structures enwrapping CNS neurons, which are important regulators for neuronal and synaptic functions. Brevican, a chondroitin sulfate proteoglycan, is an integral component of PNN. Here, we have investigated the appearance of these struct... [more] Perineuronal nets (PNN) are specialized extracellular matrix structures enwrapping CNS neurons, which are important regulators for neuronal and synaptic functions. Brevican, a chondroitin sulfate proteoglycan, is an integral component of PNN. Here, we have investigated the appearance of these structures in hippocampal primary cultures. The expression profile of brevican in mixed cultures resembles the in vivo pattern with a strong upregulation of all isoforms during the second and 3rd weeks in culture. Brevican is primarily synthesized by co-cultured glial fibrillary acidic protein (GFAP-)-positive astrocytes and co-assembles with its interaction partners in PNN-like structures on neuronal somata and neurites as identified by counterstaining with the PNN marker Vicia villosa lectin. Both excitatory and inhibitory synapses are embedded into PNN. Furthermore, axon initial segments are strongly covered by a dense brevican coat. Altogether, we show that mature primary cultures can form PNN, and that basic features of these extracellular matrix structures may be studied in vitro.
Following (5)
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Christina Spilker
Leibniz Institute for Neuroscience -
Ka Wan Li
Vrije Universiteit Amsterdam -
Susanne tom Dieck
Max-Planck-Institut für Hirnforschung -
Karl-Heinz Braunewell
Southern Research Institute -
Bernhard Bogerts
Otto-von-Guericke-Universität Magdeburg
