Synaptic Compartments Research Articles

Subcellular TSC22D4 Localization in Cerebellum Granule Neurons of the Mouse Depends on Development and Differentiation

We previously demonstrated that TSC22D4, a protein encoded by the TGF-β1-activated gene Tsc22d4 (Thg-1pit) and highly expressed in postnatal and adult mouse cerebellum with multiple post-translationally modified protein forms, moves to nucleus when in vitro differentiated cerebellum granule neurons (CGNs) are committed to apoptosis by hyperpolarizing KCl concentrations in the culture medium. We have now studied TSC22D4 cytoplasmic/nuclear localization in CGNs and Purkinje cells: (1) during CGN differentiation/maturation in vivo, (2) during CGN differentiation in vitro, and (3) by in vitro culturing ex vivo cerebellum slices under conditions favoring/inhibiting CGN/Purkinje cell differentiation. We show that TSC22D4 displays both nuclear and cytoplasmic localizations in undifferentiated, early postnatal cerebellum CGNs, irrespectively of CGN proliferation/migration from external to internal granule cell layer, and that it specifically accumulates in the somatodendritic and synaptic compartments when CGNs mature, as indicated by TSC22D4 abundance at the level of adult cerebellum glomeruli and apparent lack in CGN nuclei. These features were also observed in cerebellum slices cultured in vitro under conditions favoring/inhibiting CGN/Purkinje cell differentiation. In vitro TSC22D4 silencing with siRNAs blocked CGN differentiation and inhibited neurite elongation in N1E-115 neuroblastoma cells, pinpointing the relevance of this protein to CGN differentiation.

Cognitive Deficits Associated with Alteration of Synaptic Metaplasticity Precede Plaque Deposition in AβPP23 Transgenic Mice

Synaptic dysfunction is an early event in the development of Alzheimer's disease (AD) and relates closely to the cognitive impairment characterizing this neurodegenerative process. A causative association has been proposed, largely on the basis of in vitro studies, between memory decline, soluble amyloid-β (Aβ) oligomers and alterations of glutamatergic neurotransmission. We aimed here to characterize in vivo N-methyl-D-aspartate receptor (NMDAR)-mediated signaling, at an early stage of AD, before extracellular amyloid plaques are deposited. We assessed the functional link between cognitive abilities and NMDAR-mediated pharmacological responses of six-month-old AβPP23 transgenic mice (AβPP23tg), overexpressing the human amyloid-β protein precursor carrying the Swedish double mutation. We found evidence of cognitive impairments in these mice, indicated by deficits in the delayed-non-matching-to-place task. Alterations of NMDAR-mediated signaling in this mouse model were confirmed by the reduced sensitivity of motor-activation and working memory to pharmacological inhibition of NMDAR activity. At the molecular level, AβPP23tg mice show hippocampal alterations in the trafficking of synaptic NMDAR subunits NR2A and NR2B and at an ultrastructural analysis show Aβ oligomers intracellularly localized in the synaptic compartments. Importantly, the behavioral and biochemical alterations of NMDAR signaling are associated with the inhibition of long-term synaptic potentiation and inversion of metaplasticity at CA1 synapses in hippocampal slices from AβPP23tg mice. These results indicate a general impairment of synaptic function and learning and memory in young AβPP23tg mice with Aβ oligomers but no amyloid plaques.

Gamma‐protocadherins are enriched and transported in specialized vesicles associated with the secretory pathway in neurons

Gamma protocadherins (Pcdh-γs) resemble classical cadherins and have the potential to engage in cell-cell interactions with homophilic properties. Emerging evidence suggests non-conventional roles for some protocadherins in neural development. We sought to determine whether Pcdh-γ trafficking in neurons is consistent with an intracellular role for these molecules. Here we show that, in contrast to the largely surface localization of classical cadherins, endogenous Pcdh-γs are primarily intracellular in rat neurons in vivo and are equally distributed within organelles of subsynaptic dendritic and axonal compartments. A strikingly higher proportion of Pcdh-γ-containing organelles in synaptic compartments was observed at postnatal day 16. To determine the origin of Pcdh-γ-trafficking organelles, we isolated organelles with Pcdh-γ antibody-coupled magnetic beads from brain organelle suspensions. Vesicles with high levels of COPII and endoplasmic reticulum-Golgi intermediate compartment (ERGIC) components were isolated with the Pcdh-γ antibody but not with the classical cadherin antibody. In cultured hippocampal neurons, Pcdh-γ immunolabeling partially overlapped with calnexin- and COPII-positive puncta in dendrites. Mobile Pcdh-γ-GFP profiles dynamically codistributed with a DsRed construct coupled to ER retention signals by live imaging. Pcdh-γ expression correlated with accumulations of tubulovesicular and ER-like organelles in dendrites. Our results are consistent with the possibility that Pcdh-γs could have a unique function within the secretory pathway in addition to their documented surface roles.

Secreted factors as synaptic organizers

A critical step in synaptic development is the differentiation of presynaptic and postsynaptic compartments. This complex process is regulated by a variety of secreted factors that serve as synaptic organizers. Specifically, fibroblast growth factors, Wnts, neurotrophic factors and various other intercellular signaling molecules are proposed to regulate presynaptic and/or postsynaptic differentiation. Many of these factors appear to function at both the neuromuscular junction and in the central nervous system, although the specific function of the molecules differs between the two. Here we review secreted molecules that organize the synaptic compartments and discuss how these molecules shape synaptic development, focusing on mammalian in vivo systems. Their critical role in shaping a functional neural circuit is underscored by their possible link to a wide range of neurological and psychiatric disorders both in animal models and by mutations identified in human patients.

Reactive hypertrophy of synaptic varicosities within the hippocampus of prion-infected mice

Prion diseases are characteristically accompanied by extensive synaptic pathology that can occur during the preclinical phase of the disease and, in animal models, correlates with the first decline of hippocampus-dependent cognitive functions. This pathology is defined by abnormally shaped synapses in which the postsynaptic membrane modifies its curvature and potentially engulfs the juxtaposed presynaptic membrane. Using the intrahippocampally injected ME7 prion model, we further detailed the structural alterations of the population of ostensibly intact synaptic compartments within the hippocampus during this period of extensive synaptic loss. A disease stage-dependent increase in the average PSD (postsynaptic density) area, the average length of the active zone and the average number of synaptic vesicles indicated that the synapses that were visualized as the animal progressed to end-stage disease were undergoing hypertrophy. Similar findings in samples from AD (Alzheimer's disease) patients, aged and senile individuals, and animal models of neurodegenerative diseases suggest synaptic swelling as synaptic loss is initiated and/or compensatory reaction to counteract the synaptic loss.

Characterization of the brain 26S proteasome and its interacting proteins

Proteasome-mediated proteolysis is important for synaptic plasticity, neuronal development, protein quality control, and many other processes in neurons. To define proteasome composition in brain, we affinity purified 26S proteasomes from cytosolic and synaptic compartments of the rat cortex. Using tandem mass spectrometry, we identified the standard 26S subunits and a set of 28 proteasome-interacting proteins that associated substoichiometrically and may serve as regulators or cofactors. This set differed from those in other tissues and we also found several proteins that associated only with either the cytosolic or the synaptic proteasome. The latter included the ubiquitin-binding factor TAX1BP1 and synaptic vesicle protein SNAP-25. Native gel electrophoresis revealed a higher proportion of doubly-capped 26S proteasome (19S-20S-19S) in the cortex than in the liver or kidney. To investigate the interplay between proteasome regulation and synaptic plasticity, we exposed cultured neurons to glutamate receptor agonist NMDA. Within 4 h, this agent caused a prolonged decrease in the activity of the ubiquitin-proteasome system as shown by disassembly of 26S proteasomes, decrease in ubiquitin-protein conjugates, and dissociation of the ubiquitin ligases UBE3A (E6-AP) and HUWE1 from the proteasome. Surprisingly, the regulatory 19S particles were rapidly degraded by proteasomal, not lysosomal degradation, and the dissociated E3 enzymes also degraded. Thus the content of proteasomes and their set of associated proteins can be altered by neuronal activity, in a manner likely to influence synaptic plasticity and learning.

Connectivity in the human fetal brain

Event Abstract Back to Event Connectivity in the human fetal brain Ivica Kostovic1* and Lana Vasung1 1 School of Medicine University of Zagreb, Croatian Institute for Brain Research, Netherlands It is generally accepted that complexity of neuronal connections in the human brain underlies complexity of cognitive and behavioral functions. In this paper we explain how developmental cellular events and connectivity patterns form special framework for early functions and establishment of adult circuitry. At the beginning of fetal period distributions of circuitry elements (axons, dendrites and synapses) is strictly bi-laminar outside of proliferative zones. The first quantifiable number of synapses occurs after 15 post conceptional weeks (PCW) in the transient subplate zone. This early synapses are substrate of endogenous, sensory non-driven activity. Distribution within the connectivity compartments is random and no modular units were observed. Important phenomenon in development of afferent input is "waiting" phenomenon: axons "wait" for several months until they gradually invade target cortical area. These waiting synaptic compartments may be visualized in both in vivo and postmortem images. The bipolar orientation of synapses, with predominance of deep circuitry is basis for cortical surface-positive electric response. The majority of described circuitry is transient. It is only after 23 PCW that sensory driven afferents establish connections in the corresponding cortical target area. At the end of gestation another phenomenon take place and that is a dramatic reorganization of fetal circuitry due to the retraction of axons and gradual disappearance of transient fetal layers. In conclusion, early fetal circuitry shows transient laminar distribution and transient endogenous activity. Sensory driven activity is late phenomenon and further development of connectivity proceeds after dramatic spatial and cellular reorganization of cortex. The knowledge on development of fetal circuitry is important for understanding of biological basis of motor sensory and cognitive functions as well as developmental origin of mental and cognitive disorders in men. Conference: Biomag 2010 - 17th International Conference on Biomagnetism , Dubrovnik, Croatia, 28 Mar - 1 Apr, 2010. Presentation Type: Oral Presentation Topic: Fetal and neonatal biomagnetism Citation: Kostovic I and Vasung L (2010). Connectivity in the human fetal brain. Front. Neurosci. Conference Abstract: Biomag 2010 - 17th International Conference on Biomagnetism . doi: 10.3389/conf.fnins.2010.06.00071 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 22 Mar 2010; Published Online: 22 Mar 2010. * Correspondence: Ivica Kostovic, School of Medicine University of Zagreb, Croatian Institute for Brain Research, Zagreb, Netherlands, ikostov@hiim.hr Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Ivica Kostovic Lana Vasung Google Ivica Kostovic Lana Vasung Google Scholar Ivica Kostovic Lana Vasung PubMed Ivica Kostovic Lana Vasung Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Expression of the neuroprotective slow Wallerian degeneration (Wld S ) gene in non-neuronal tissues

BackgroundThe slow Wallerian Degeneration (WldS) gene specifically protects axonal and synaptic compartments of neurons from a wide variety of degeneration-inducing stimuli, including; traumatic injury, Parkinson's disease, demyelinating neuropathies, some forms of motor neuron disease and global cerebral ischemia. The WldS gene encodes a novel Ube4b-Nmnat1 chimeric protein (WldS protein) that is responsible for conferring the neuroprotective phenotype. How the chimeric WldS protein confers neuroprotection remains controversial, but several studies have shown that expression in neurons in vivo and in vitro modifies key cellular pathways, including; NAD biosynthesis, ubiquitination, the mitochondrial proteome, cell cycle status and cell stress. Whether similar changes are induced in non-neuronal tissue and organs at a basal level in vivo remains to be determined. This may be of particular importance for the development and application of neuroprotective therapeutic strategies based around WldS-mediated pathways designed for use in human patients.ResultsWe have undertaken a detailed analysis of non-neuronal WldS expression in WldS mice, alongside gravimetric and histological analyses, to examine the influence of WldS expression in non-neuronal tissues. We show that expression of WldS RNA and protein are not restricted to neuronal tissue, but that the relative RNA and protein expression levels rarely correlate in these non-neuronal tissues. We show that WldS mice have normal body weight and growth characteristics as well as gravimetrically and histologically normal organs, regardless of WldS protein levels. Finally, we demonstrate that previously reported WldS-induced changes in cell cycle and cell stress status are neuronal-specific, not recapitulated in non-neuronal tissues at a basal level.ConclusionsWe conclude that expression of WldS protein has no adverse effects on non-neuronal tissue at a basal level in vivo, supporting the possibility of its safe use in future therapeutic strategies targeting axonal and/or synaptic compartments in patients with neurodegenerative disease. Future experiments determining whether WldS protein can modify responses to injury in non-neuronal tissue are now required.

Postsynaptic regulation of synaptic plasticity by synaptotagmin 4 requires both C2 domains

Ca(2+) influx into synaptic compartments during activity is a key mediator of neuronal plasticity. Although the role of presynaptic Ca(2+) in triggering vesicle fusion though the Ca(2+) sensor synaptotagmin 1 (Syt 1) is established, molecular mechanisms that underlie responses to postsynaptic Ca(2+) influx remain unclear. In this study, we demonstrate that fusion-competent Syt 4 vesicles localize postsynaptically at both neuromuscular junctions (NMJs) and central nervous system synapses in Drosophila melanogaster. Syt 4 messenger RNA and protein expression are strongly regulated by neuronal activity, whereas altered levels of postsynaptic Syt 4 modify synaptic growth and presynaptic release properties. Syt 4 is required for known forms of activity-dependent structural plasticity at NMJs. Synaptic proliferation and retrograde signaling mediated by Syt 4 requires functional C2A and C2B Ca(2+)-binding sites, as well as serine 284, an evolutionarily conserved substitution for a key Ca(2+)-binding aspartic acid found in other synaptotagmins. These data suggest that Syt 4 regulates activity-dependent release of postsynaptic retrograde signals that promote synaptic plasticity, similar to the role of Syt 1 as a Ca(2+) sensor for presynaptic vesicle fusion.

Comprehensive Mapping of Post-Translational Modifications on Synaptic, Nuclear, and Histone Proteins in the Adult Mouse Brain

Post-translational modifications (PTMs) of proteins in the adult brain are known to mark activity-dependent processes for complex brain functions such as learning and memory. Multiple PTMs occur in nerve cells, and are able to modulate proteins in different subcellular compartments. In synaptic terminals, protein phosphorylation is the primary PTM that contributes to the control of the activity and localization of synaptic proteins. In the nucleus, it can modulate histones and proteins involved with the transcriptional machinery and, in combination with other PTMs such as acetylation, methylation and ubiquitination, acts to regulate chromatin remodelling and gene expression. The combination of histone PTMs is highly complex and is known to be unique to each gene. The ensemble of PTMs in the adult brain, however, remains unknown. Here, we describe a novel proteomic approach that allows the isolation and identification of PTMs on synaptic and nuclear proteins, in particular on histones. Using subcellular fractionation, we identified 2082 unique phosphopeptides from 1062 phosphoproteins, and 196 unique PTM sites on histones H1, H2A, H2B, H3 and H4. A comparison of phosphorylation sites in synaptic and nuclear compartments, and on histones, suggests that different kinases and kinase motifs are involved. Overall, our data demonstrates the complexity of PTMs in the brain and the prevalence of histone PTMs, and reveals potentially important regulatory sites on proteins involved in synaptic plasticity and brain functions.

Synaptic Proteins Linked to HIV-1 Infection and Immunoproteasome Induction: Proteomic Analysis of Human Synaptosomes

Infection of the central nervous system with human immunodeficiency virus type 1 (HIV-1) can produce morphological changes in the neocortical synaptodendritic arbor that are correlated with neurocognitive impairment. To determine whether HIV-1 infection influences the protein composition of human synapses, a proteomic study of isolated nerve endings was undertaken. Synaptosomes from frontal neocortex were isolated using isopyknic centrifugation from 19 human brain specimens. Purity and enrichment were assessed by measuring pre- and postsynaptic protein markers. Two-dimensional polyacrylamide gel electrophoresis and matrix-assisted laser desorption ionization time-of-flight mass spectrometry was used to screen for proteins differentially expressed in HIV/AIDS. The concentrations of 31 candidate protein spots were potentially abnormal in HIV-infected decedents with HIV encephalitis and/or increased expression of immunoproteasome subunits. Immunoblots showed that the concentration of some of them was related to HIV-1 infection of the brain and immunoproteasome (IPS) induction. Synapsin 1b and stathmin were inversely related to brain HIV-1 load; 14-3-3ζ and 14-4-4ε proteins were higher in subjects with HIV-1 loads. Perturbed synaptosome proteins were linked with IPS subunit composition, and 14-3-3ζ was histologically colocalized with IPS subunits in stained neocortical neurons. Proteomics illustrates that certain human proteins within the synaptic compartment are involved with changes in the synaptodendritic arbor and neurocognitive impairment in HIV-1-infected people.

Organisation structurale, moléculaire, formation et maturation de la jonction neuromusculaire

The neuromuscular junction is made up of the apposition of highly differentiated domains of three types of cell: the motor neuronal ending, the terminal Schwann cell and the muscle postsynaptic membrane. These three components are surrounded by a basal lamina, dedicated to molecular signal exchanges controlling neuromuscular formation, maturation and maintenance. This functional and structural differentiated complex conducts synaptic neurotransmission to the skeletal muscle fiber. Nerve and muscle have distinct roles in synaptic compartment differentiation. The initial steps of this differentiation and the motor endplate formation require several postsynaptic molecular agents including agrin, the tyrosine kinase receptor MuSK. Neuregulin is essentially involved in Schwann cell survival and guidance for axonal growth.

Formation et régénération synaptique

Abstract Synapse formation is probably the key process in neural development allowing signal transmission between nerve cells. As an interesting model of synapse maturation, we considered first the neuromuscular junction (NMJ), whose development is particularly dependent on intercellular interactions between the motor nerve and the skeletal muscle. Nerve and muscle have distinct roles in synaptic compartment differentiation. The initial steps of this differentiation and motor endplate formation require several postsynaptic molecular agents including agrin, the tyrosine kinase receptor MuSK and rapsyn. The agrin or motoneuron dependence of this process continues to be debated while the following steps of axonal growth and postsynaptic apparatus maintenance essentially depend on neuronal agrin and a neuron-specific signal dispersing ectopic AChR aggregate remainders, possibly mediated by acetylcholine itself. Neuregulin is essentially involved in Schwann's cell survival and guidance for axonal growth. In this paper, we will discuss the similarities between Central Nervous System (CNS) synaptic formation and Motor innervation. The limited ability of the CNS to create new synapses after nervous system injury will be then discussed with a final consideration of some new strategies elaborated to circumvent the limitations of lesion extension processes.

Presynaptic translation: stepping out of the postsynaptic shadow

The ability of the nervous system to convert transient experiences into long-lasting structural changes at the synapse relies upon protein synthesis. It has become increasingly clear that a critical subset of this synthesis occurs within the synaptic compartment. While this process has been extensively characterized in the postsynaptic compartment, the contribution of local translation to presynaptic function remains largely unexplored. However, recent evidence highlights the potential importance of translation within the presynaptic compartment. Work in cultured neurons has shown that presynaptic translation occurs specifically at synapses undergoing long-term plasticity and may contribute to the maintenance of nascent synapses. Studies from our laboratory have demonstrated that Fragile X proteins, which regulate mRNA localization and translation, are expressed at the presynaptic apparatus. Further, mRNAs encoding presynaptic proteins traffic into axons. Here we discuss recent advances in the study of presynaptic translation as well as the challenges confronting the field. Understanding the regulation of presynaptic function by local protein synthesis promises to shed new light on activity-dependent modification of synaptic architecture.

Amyloid Beta Annular Protofibrils in Cell Processes and Synapses Accumulate with Aging and Alzheimer-Associated Genetic Modification

Amyloid β (Aβ) annular protofibrils (APFs) have been described where the structure is related to that of β barrel pore-forming bacterial toxins and exhibits cellular toxicity. To investigate the relationship of Aβ APFs to disease and their ultrastructural localization in brain tissue, we conducted a pre-embedding immunoelectron microscopic study using anti-annular protofibril antiserum. We examined brain tissues of young- and old-aged amyloid precursor protein transgenic mice (APP23), neprilysin knockout APP23 mice, and nontransgenic littermates. αAPF-immunoreactions tended to be found (1) on plasma membranes and vesicles inside of cell processes, but not on amyloid fibrils, (2) with higher density due to aging, APP transgene, and neprilysin deficiency, and (3) with higher positive rate at synaptic compartments in aged APP23, especially in neprilysin knockout APP23 mice. These findings imply that APFs are distinct from amyloid fibrils, interact with biological membranes, and might be related to synaptic dysfunction in Alzheimer model mouse brains.

Cellular transport and membrane dynamics of the glycine receptor

Regulation of synaptic transmission is essential to tune individual-to-network neuronal activity. One way to modulate synaptic strength is to regulate neurotransmitter receptor numbers at postsynaptic sites. This can be achieved either through plasma membrane insertion of receptors derived from intracellular vesicle pools, a process depending on active cytoskeleton transport, or through surface membrane removal via endocytosis. In parallel, lateral diffusion events along the plasma membrane allow the exchange of receptor molecules between synaptic and extrasynaptic compartments, contributing to synaptic strength regulation. In recent years, results obtained from several groups studying glycine receptor (GlyR) trafficking and dynamics shed light on the regulation of synaptic GlyR density. Here, we review (i) proteins and mechanisms involved in GlyR cytoskeletal transport, (ii) the diffusion dynamics of GlyR and of its scaffolding protein gephyrin that control receptor numbers, and its relationship with synaptic plasticity, and (iii) adaptative changes in GlyR diffusion in response to global activity modifications, as a homeostatic mechanism.

The JIP3 scaffold protein UNC‐16 regulates RAB‐5 dependent membrane trafficking at C. elegans synapses

How endosomes contribute to the maintenance of vesicular structures at presynaptic terminals remains controversial and poorly understood. Here, we have investigated synaptic endosomal compartments in the presynaptic terminals of C. elegans GABAergic motor neurons. Using RAB reporters, we find that several subsynaptic compartments reside in, or near, presynaptic regions. Loss of function in the C. elegans JIP3 protein, UNC-16, causes a RAB-5-containing compartment to accumulate abnormally at presynaptic terminals. Ultrastructural analysis shows that synapses in unc-16 mutants contain reduced number of synaptic vesicles, accompanied by an increase in the size and number of cisternae. FRAP analysis revealed a slow recovery of RAB-5 in unc-16 mutants, suggestive of an impairment of RAB-5 activity state and local vesicular trafficking. Overexpression of RAB-5:GDP partially suppresses, whereas overexpression of RAB-5:GTP enhances, the synaptic defects of unc-16 mutants. Our data demonstrate a novel function of UNC-16 in the regulation of synaptic membrane trafficking and suggest that the synaptic RAB-5 compartment contributes to synaptic vesicle biogenesis or maintenance.

Loss of translation elongation factor (eEF1A2) expression in vivo differentiates between Wallerian degeneration and dying‐back neuronal pathology

Wallerian degeneration and dying-back pathology are two well-known cellular pathways capable of regulating the breakdown and loss of axonal and synaptic compartments of neurons in vivo. However, the underlying mechanisms and molecular triggers of these pathways remain elusive. Here, we show that loss of translation elongation factor eEF1A2 expression in lower motor neurons and skeletal muscle fibres in homozygous Wasted mice triggered a dying-back neuropathy. Synaptic loss at the neuromuscular junction occurred in advance of axonal pathology and by a mechanism morphologically distinct from Wallerian degeneration. Dying-back pathology in Wasted mice was accompanied by reduced expression levels of the zinc finger protein ZPR1, as found in other dying-back neuropathies such as spinal muscular atrophy. Surprisingly, experimental nerve lesion revealed that Wallerian degeneration was significantly delayed in homozygous Wasted mice; morphological assessment revealed that approximately 80% of neuromuscular junctions in deep lumbrical muscles at 24 h and approximately 50% at 48 h had retained motor nerve terminals following tibial nerve lesion. This was in contrast to wild-type and heterozygous Wasted mice where < 5% of neuromuscular junctions had retained motor nerve terminals at 24 h post-lesion. These data show that eEF1A2 expression is required to prevent the initiation of dying-back pathology at the neuromuscular junction in vivo. In contrast, loss of eEF1A2 expression significantly inhibited the initiation and progression of Wallerian degeneration in vivo. We conclude that loss of eEF1A2 expression distinguishes mechanisms underlying dying-back pathology from those responsible for Wallerian degeneration in vivo and suggest that eEF1A2-dependent cascades may provide novel molecular targets to manipulate neurodegenerative pathways in lower motor neurons.

Caspase-3 Is Enriched in Postsynaptic Densities and Increased in Alzheimer's Disease

Progressive synaptic degeneration and neuron loss are major structural correlates of cognitive impairment in Alzheimer's disease (AD). The mechanisms by which synaptic degeneration in AD occurs have not been established. The activation of proteins within the caspase family has been implicated in AD-associated neurodegeneration, and synaptically localized caspase activity could play a role in the synaptic degeneration and loss found in AD. We used synaptosomal fractionation with Western blotting and immunohistochemistry to examine the anatomical, subcellular, and subsynaptic expression patterns of caspase 3 in both the anterior cingulate cortex and hippocampus of control and AD patients. In both control and AD cases, there was a selective enrichment of caspase- 3 at synapses, particularly in the postsynaptic density (PSD) fractions. Compared with controls, AD patients exhibited significant increases in synaptic procaspase- 3 and active caspase-3 expression levels that were most evident in the PSD fractions. These data demonstrate for the first time the preferential localization and increase of caspase-3 in the PSD fractions in AD and suggest an important role for caspase 3 in synapse degeneration during disease progression.

Contralateral cortical response of a subpopulation of interneurons and the glial glutamate transporter GLT1 to an ischemic core.

Introduction: Cerebral ischemia is an important cause of brain lesion in humans. The target in research has been the ischemic core or the penumbra zones; little attention has been given to areas outside the core or the penumbra but connected with the primary site of injury. Objective: Evaluate the laminar response of a subpopulation of gabaergic cells, those that are parvalbumin (PV) positive and the astrocytes through the expression of the glial transporter GLT1 on the contralateral cortex to an ischemic core. Methodology: For this purpose we used the medial cerebral artery occlusion model in rats. The artery was occluded for 90 minutes and the animals were sacrificed at 24 and 72 hours post-ischemia. The brains were removed, cut in a vibratome at 50 microns and incubated with the primary antibodies against PV or GLT1. Sections were developed using the vectastain Kit. In control tissue the primary antibody was omitted. Results: When compared with control animals, treated ones show a decrease in the expression of GLT1, especially in layers III and IV of the contralateral cortex to the ischemic core. PV positive cells increases in layers II and V. Conclusion: Increases in the expression of PV cells could correspond to an adaptation associated with glutamate increases in the synaptic compartment. These increases may be due to decreases in the expression of GLT1 transporter, that could not remove the glutamate present in the synaptic cleft, generating hyperactivity in the contralateral cortex. These changes could represent an example of neuronal and glial plasticity in remote areas to an ischemic core but connected to the primary site of injury.