Decreases In Synaptic Proteins Research Articles

The effects of neurosteroid allopregnanolone on synaptic dysfunction in the hippocampus in experimental parkinsonism rats: An electrophysiological and molecular study

The dopaminergic system is a powerful candidate targeted for changes of synaptic plasticity in the hippocampus. Higher incidence of Parkinson's disease (PD) in men than women indicates the influence of sex hormones on the PD development. Previous studies have shown that neurodegenerative diseases such as PD are related to the decline of Allopregnanolon (Allo), a metabolite of progesterone; it is also well known that learning and memory are influenced by oscillations in steroidal hormones. Although abnormalities in hippocampal plasticity have been observed in the toxic models of PD, effects of Allo on hippocampal LTP and hippocampal synaptic protein levels, which play an important role in maintaining the integrity of neural connections, have never been analyzed thus far. Experimental groups subjected to the long-term potentiation (LTP) were studied in the CA1 area of the hippocampus. In addition, the levels of hippocampal postsynaptic density protein 95 (PSD-95), neurexin-1 (Nrxn1) and neuroligin (Nlgn) as synaptic molecular components were determined by immunoblotting. Although dopamine denervation did not alter basal synaptic transmission and pair-pulse facilitation of field excitatory postsynaptic potentials (fEPSPs), the induction and maintenance of LTP were impaired in the CA1 region. In addition, the levels of PSD-95, Nrxn1 and Nlgn were significantly decreased in the hippocampus of 6-OHDA-treated animals. Such abnormalities in synaptic electrophysiological aspects and protein levels were abolished by the treatment with Allo. These findings showed that partial dopamine depletion led to unusual synaptic plasticity in the CA1 as well as the decrease in synaptic proteins in the hippocampus. Our results demonstrated that Allo ameliorated these deficits and preserved pre- and post-synaptic proteins. Therefore, Allo may be an effective factor in maintaining synaptic integrity in the mesolimbic pathway.

Blockage of AEP attenuates TBI-induced tau hyperphosphorylation and cognitive impairments in rats.

Traumatic brain injury (TBI) is regarded as a high-risk factor for Alzheimer's disease (AD). Asparaginyl endopeptidase (AEP), a lysosomal cysteine protease involved in AD pathogenesis, is normally activated under acidic conditions and also in TBI. However, both the molecular mechanism underlying AEP activation-mediated TBI-related AD pathologies, and the role of AEP as an AD therapeutic target, still remain unclear. Here, we report that TBI induces hippocampus dependent cognitive deficit and synaptic dysfunction, accompanied with AEP activation, I2PP2A (inhibitor 2 of PP2A, also called SET) mis-translocation from neuronal nucleus to cytoplasm, an obvious increase in AEP interaction with SET, and tau hyperphosphorylation in hippocampus of rats. Oxygen-glucose deprivation (OGD), mimicking an acidic condition, also leads to AEP activation, SET mis-translocation, PP2A inhibition, tau hyperphosphorylation, and a decrease in synaptic proteins, all of which are abrogated by AEP inhibitor AENK in primary neurons. Interestingly, AENK restores SET back to the nucleus, mitigates tau pathologies, rescuing TBI-induced cognitive deficit in rats. These findings highlight a novel etiopathogenic mechanism of TBI-related AD, which is initiated by AEP activation, accumulating SET in cytoplasm, and favoring tau pathology and cognitive impairments. Lowering AEP activity by AEP inhibitor would be beneficial to AD patients with TBI.

Advantages of Acute Brain Slices Prepared at Physiological Temperature in the Characterization of Synaptic Functions.

Acute brain slice preparation is a powerful experimental model for investigating the characteristics of synaptic function in the brain. Although brain tissue is usually cut at ice-cold temperature (CT) to facilitate slicing and avoid neuronal damage, exposure to CT causes molecular and architectural changes of synapses. To address these issues, we investigated ultrastructural and electrophysiological features of synapses in mouse acute cerebellar slices prepared at ice-cold and physiological temperature (PT). In the slices prepared at CT, we found significant spine loss and reconstruction, synaptic vesicle rearrangement and decrease in synaptic proteins, all of which were not detected in slices prepared at PT. Consistent with these structural findings, slices prepared at PT showed higher release probability. Furthermore, preparation at PT allows electrophysiological recording immediately after slicing resulting in higher detectability of long-term depression (LTD) after motor learning compared with that at CT. These results indicate substantial advantages of the slice preparation at PT for investigating synaptic functions in different physiological conditions.

Proteomic alterations of brain subcellular organelles caused by low-dose copper exposure: implication for Alzheimer's disease.

Excessive copper intake can lead to neurotoxicity, but there is a lack of comprehensive understanding on the potential impact of copper exposure especially at a low-dose on brain. We used 3xTg-AD mice to explore the potential neurotoxicity of chronic, low-dose copper treatment (0.13ppm copper chloride in drinking water) on behavior and the brain hippocampal mitochondrial and nuclear proteome. Low-dose copper increased the spatial memory impairment of these animals, increased accumulation of intracellular amyloid 1-42 (Aβ1-42), decreased ATP content, increased the positive staining of 8-hydroxyguanosine (8-OHdG), a marker of DNA oxidative damage, and caused apoptosis and a decrease in synaptic proteins. Mitochondrial proteomic analysis by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) revealed modulation of 24 hippocampal mitochondrial proteins (14 increased and 10 decreased) in copper-treated vs. untreated 3xTg-AD mice. Nuclear proteomic analysis revealed 43 modulated hippocampal nuclear proteins (25 increased and 18 decreased) in copper-treated 3xTg-AD vs. untreated mice. Classification of modulated mitochondrial and nuclear proteins included functional categories such as energy metabolism, synaptic-related proteins, DNA damage and apoptosis-related proteins, and oxidative stress-related proteins. Among these differentially expressed mitochondrial and nuclear proteins, nine proteins were abnormally expressed in both hippocampus mitochondria and nuclei, including electron transport chain-related proteins NADH dehydrogenase 1 alpha subcomplex subunit 10 (NDUAA), cytochrome b-c1 complex subunit Rieske (UCRI), cytochrome c oxidase subunit 5B (COX5B), and ATP synthase subunit d (ATP5H), glycolytic-related pyruvate kinase PKM (KPYM) and pyruvate dehydrogenase E1 component subunit alpha (ODPA). Furthermore, we found coenzyme Q10 (CoQ10), an endogenous mitochondrial protective factor/antioxidant, modulated the expression of 12 differentially expressed hippocampal proteins (4 increased and 8 decreased), which could be classified in functional categories such as glycolysis and synaptic-related proteins, oxidative stress-related proteins, implying that CoQ10 improved synaptic function, suppress oxidative stress, and regulate glycolysis. For the proteomics study, we validated the expression of several proteins related to synapses, DNA and apoptosis. The data confirmed that synapsin-2, a synaptic-related protein, was significantly decreased in both mitochondria and nuclei of copper-exposed 3xTg-AD mice. In mitochondria, dynamin-1 (DYN1), an apoptosis-related proteins, was significantly decreased. In the cellular nuclei, paraspeckle protein 1 (PSPC1) and purin-rich element-binding protein alpha (Purα), two DNA damage-related proteins, were significantly decreased and increased, respectively. We conclude that low-dose copper exposure exacerbates the spatial memory impairment of 3xTg-AD mice and perturbs multiple biological/pathogenic processes by dysregulating the mitochondrial and nuclear proteome. Exposure to copper might therefore contribute to the evolution of AD.

P120-catenin is necessary for neuroprotection induced by CDK5 silencing in models of Alzheimer's disease.

Cyclin-dependent kinase 5 (CDK5) plays important roles in synaptic function. Its unregulated over-activation has been, however, associated with neurodegeneration in Alzheimer's disease. Our previous studies revealed that CDK5 silencing ameliorates tauopathy and spatial memory impairment in the 3xTgAD mouse model. However, how CDK5 targeting affects synaptic adhesion proteins, such as those involved in the cadherin/catenin system, during learning and memory processes is not completely understood. In this study, we detected reduced expression of p120 catenin (p120 ctn), N-cadherin, and β-catenin in the brain of human Alzheimer's disease patients, in addition to a reduced PSD95 and GluN2B protein levels in a 3xTgAD mouse model. Such decrease in synaptic proteins was recovered by CDK5 silencing in mice leading to a better learning and memory performance. Additionally, CDK5 inhibition or knockout increased p120 ctn levels. Moreover, in a glutamate-induced excitotoxicity model, CDK5 silencing-induced neuroprotection depended on p120 ctn. Together, those findings suggest that p120 ctn plays an important role in the neuronal dysfunction of Alzheimer's disease models and contributes to CDK5 silencing-induced neuroprotection and improvement of memory function. p120ctn is part of the synaptic adhesion molecular complex N-cadh/p120ctn/B-ctn/PSD95, and it has a pivotal role in cell adhesion stabilization and dendritic spine modulation. Our data show that synaptic adhesion complex is affected in AD human brains and in AD models. This complex is recovered by the silencing of CDK5, preventing memory dysfunction in an AD mice model and contributing to the neuroprotection in a depend-mode of p120ctn.

Decreased astrocytic thrombospondin-1 secretion after chronic ammonia treatment reduces the level of synaptic proteins: in vitro and in vivo studies.

Chronic hepatic encephalopathy (CHE) is a major complication in patients with severe liver disease. Elevated blood and brain ammonia levels have been implicated in its pathogenesis, and astrocytes are the principal neural cells involved in this disorder. Since defective synthesis and release of astrocytic factors have been shown to impair synaptic integrity in other neurological conditions, we examined whether thrombospondin-1 (TSP-1), an astrocytic factor involved in the maintenance of synaptic integrity, is also altered in CHE. Cultured astrocytes were exposed to ammonia (NH₄Cl, 0.5-2.5 mM) for 1-10 days, and TSP-1 content was measured in cell extracts and culture media. Astrocytes exposed to ammonia exhibited a reduction in intra- and extracellular TSP-1 levels. Exposure of cultured neurons to conditioned media from ammonia-treated astrocytes showed a decrease in synaptophysin, PSD95, and synaptotagmin levels. Conditioned media from TSP-1 over-expressing astrocytes that were treated with ammonia, when added to cultured neurons, reversed the decline in synaptic proteins. Recombinant TSP-1 similarly reversed the decrease in synaptic proteins. Metformin, an agent known to increase TSP-1 synthesis in other cell types, also reversed the ammonia-induced TSP-1 reduction. Likewise, we found a significant decline in TSP-1 level in cortical astrocytes, as well as a reduction in synaptophysin content in vivo in a rat model of CHE. These findings suggest that TSP-1 may represent an important therapeutic target for CHE. Defective release of astrocytic factors may impair synaptic integrity in chronic hepatic encephalopathy. We found a reduction in the release of the astrocytic matricellular proteins thrombospondin-1 (TSP-1) in ammonia-treated astrocytes; such reduction was associated with a decrease in synaptic proteins caused by conditioned media from ammonia-treated astrocytes. Exposure of neurons to CM from ammonia-treated astrocytes, in which TSP-1 is over-expressed, reversed (by approx 75%) the reduction in synaptic proteins. NF-kB = nuclear factor kappa B; PSD95 = post-synaptic density protein 95; ONS = oxidative/nitrative stress.

Extinction of an instrumental response: a cognitive behavioral assay in Fmr1 knockout mice

Fragile X (FX) is the most common genetic cause of intellectual disability and autism. Previous studies have shown that partial inhibition of metabotropic glutamate receptor signaling is sufficient to correct behavioral phenotypes in a mouse model of FX, including audiogenic seizures, open-field hyperactivity and social behavior. These phenotypes model well the epilepsy (15%), hyperactivity (20%) and autism (30%) that are comorbid with FX in human patients. Identifying reliable and robust mouse phenotypes to model cognitive impairments is critical considering the 90% comorbidity of FX and intellectual disability. Recent work characterized a five-choice visuospatial discrimination assay testing cognitive flexibility, in which FX model mice show impairments associated with decreases in synaptic proteins in prefrontal cortex (PFC). In this study, we sought to determine whether instrumental extinction, another process requiring PFC, is altered in FX model mice, and whether downregulation of metabotropic glutamate receptor signaling pathways is sufficient to correct both visuospatial discrimination and extinction phenotypes. We report that instrumental extinction is consistently exaggerated in FX model mice. However, neither the extinction phenotype nor the visuospatial discrimination phenotype is corrected by approaches targeting metabotropic glutamate receptor signaling. This work describes a novel behavioral extinction assay to model impaired cognition in mouse models of neurodevelopmental disorders, provides evidence that extinction is exaggerated in the FX mouse model and suggests possible limitations of metabotropic glutamate receptor-based pharmacotherapy.

Transmission of alpha-synuclein in Parkinson’s disease

The accumulation of misfolded proteins is a fundamental pathogenic process in neurodegenerative diseases. However, the factors that trigger aggregation and spreading of α-synuclein (α-syn), the principal component of the intraneuronal inclusions known as Lewy bodies (LBs) and Lewy neurites (LNs) that characterize Parkinson’s disease (PD) and dementia with LBs (DLB) in brain are poorly understood. We demonstrate that pre-formed fibrils (pffs) generated from recombinant α-syn enter cultured primary neurons generated from non-transgenic wildtype mice, promote recruitment of soluble endogenous α-syn into insoluble PD-like LBs and LNs. Accumulation of pathologic α-syn led to selective decreases in synaptic proteins, progressive impairments in neuronal excitability and connectivity, and eventually, neuron death. Moreover, we further show that in young asymptomatic α-syn transgenic (Tg) mice, intracerebral injections of synthetic α-syn pffs accelerate both the formation of intracellular LB/LN-like inclusions and the onset of neurological symptoms in recipient animals. Pathologic α-syn propagated along major central nervous system (CNS) pathways to regions far beyond injection sites and dramatically reduced survival with a highly reproducible interval from injection to death in inoculated animals. Thus, synthetic α-Syn pffs are wholly sufficient to initiate PD-like LBs/LNs and to transmit disease in primary neurons in vitro and Tg mice in vivo. Thus, these data support a prion-like cascade in synucleinopathies whereby cell-to-cell transmission and propagation of misfolded α-syn underlie the CNS proliferation of LBs/LNs. These findings open up new avenues for understanding the progression of PD and for developing novel therapeutics.

Exogenous α-Synuclein Fibrils Induce Lewy Body Pathology Leading to Synaptic Dysfunction and Neuron Death

Inclusions composed of α-synuclein (α-syn), i.e., Lewy bodies (LBs) and Lewy neurites (LNs), define synucleinopathies including Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Here, we demonstrate that preformed fibrils generated from full-length and truncated recombinant α-syn enter primary neurons, probably by adsorptive-mediated endocytosis, and promote recruitment of soluble endogenous α-syn into insoluble PD-like LBs and LNs. Remarkably, endogenous α-syn was sufficient for formation of these aggregates, and overexpression of wild-type or mutant α-syn was not required. LN-like pathology first developed in axons and propagated to form LB-like inclusions in perikarya. Accumulation of pathologic α-syn led to selective decreases in synaptic proteins, progressive impairments in neuronal excitability and connectivity, and, eventually, neuron death. Thus, our data contribute important insights into the etiology and pathogenesis of PD-like α-syn inclusions and their impact on neuronal functions, and they provide a model for discovering therapeutics targeting pathologic α-syn-mediated neurodegeneration.

Tetrahydrohyperforin prevents cognitive deficit, Aβ deposition, tau phosphorylation and synaptotoxicity in the APPswe/PSEN1ΔE9 model of Alzheimer's disease: a possible effect on APP processing

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by a progressive deterioration of cognitive abilities, amyloid-β peptide (Aβ) accumulation and synaptic alterations. Previous studies indicated that hyperforin, a component of the St John's Wort, prevents Aβ neurotoxicity and some behavioral impairments in a rat model of AD. In this study we examined the ability of tetrahydrohyperforin (IDN5607), a stable hyperforin derivative, to prevent the cognitive deficit and synaptic impairment in an in vivo model of AD. In double transgenic APPswe/PSEN1ΔE9 mice, IDN5706 improves memory and prevents the impairment of synaptic plasticity in a dose-dependent manner, inducing a recovery of long-term potentiation. In agreement with these findings, IDN5706 prevented the decrease in synaptic proteins in hippocampus and cortex. In addition, decreased levels of tau hyperphosphorylation, astrogliosis, and total fibrillar and oligomeric forms of Aβ were determined in double transgenic mice treated with IDN5706. In cultured cells, IDN5706 decreased the proteolytic processing of the amyloid precursor protein that leads to Aβ peptide generation. These findings indicate that IDN5706 ameliorates AD neuropathology and could be considered of therapeutic relevance in AD treatment.

Phenylbutyric acid prevents rats from electroconvulsion-induced memory deficit with alterations of memory-related proteins and tau hyperphosphorylation

Electroconvulsive therapy has been commonly applied in the treatment of refractory depression, but its cognitive side effects are noticed and restrict its application. The molecular mechanisms underlying the side effects remain elusive, and there is no efficient prevention. By employing a recognized electroconvulsive shock (ECS) rat model, we found in the present study that ECS induced spatial memory deficits with simultaneous decreases in synaptic proteins of N-methyl-D-aspartate receptor 2A/B (NR2A/B) and postsynaptic density 95 (PSD95), the immediate early gene c-Fos and cAMP response element binding (CREB) proteins, all of which are memory-related proteins. ECS also caused tau hyperphosphorylation at multiple Alzheimer-related phosphorylation sites with activation of glycogen synthase kinase-3β (GSK-3β), Akt and phospho-PKR-like endoreticulum (PERK), and inhibition of protein phosphatase-2A (PP)-2A. Intraperitoneal injection of phenylbutyric acid (PBA), an aromatic short chain fatty acid with the functions of molecule chaperon, prevented rats from the ECS-induced memory deficits, alterations of the memory-associated proteins, and tau hyperphosphorylation. Our data suggest that PBA may be potentially used for attenuating the side effects caused by electroconvulsive therapy.

The importance of interneurons in schizophrenic and affective disorders

In brains of patients with schizophrenic and affective disorders pathomorphological changes have been shown focussing in frontal and temporal cortex. The volume reduction in prefrontal cortex of schizophrenic patients is hypothesized to be based on a reduction of neuropil. A decrease of synaptic proteins and a decrease of dendritic spines of pyramidal cells can additionally be the origin of disconnections of neurons. Affection of the glutamatergic, GABA-ergic and dopaminergic system and reduction of interneurons could be the correlate of a deficient neuronal network which might be combined with exogen factors generate psychotic symptoms. Reelin and associated proteins are candidate molecules. Their dysregulation might explain essential features of the dysfunctional network of schizophrenia.

Deltamethrin differentially affects neuronal subtypes in hippocampal primary culture

The effects of deltamethrin on neuronal development and survival were studied using primary mouse hippocampal neurons in culture. Repeated applications of deltamethrin (between 2 nM and 2000 nM) decreased the number of neurons by 16–40%, respectively. Neuronal death was accompanied by an overall decrease of synaptic proteins. Deltamethrin treatment increased the K +-stimulated release of amino acid transmitters, GABA and glutamate. The release of the latter might also contribute to neuronal damage. A considerable number of neurons survived treatment with high concentrations of deltamethrin (200–2000 nM) and still displayed characteristics of mature neurons such as synaptic contacts or the expression of members of the Kv1 channel family. When analyzing subtypes of neurons calbindin- as well as somatostatin-positive neurons decreased by 50% after repeated treatment with 2 nM deltamethrin. Under the same conditions neuropeptide Y-positive neurons were up-regulated by 250%. Taken together these data show that deltamethrin at concentrations relevant in human toxicology differentially affects survival of neuronal subtypes by exerting either deleterious or supportive effects. We conclude that deltamethrin disturbs fine-tuning of neuronal efficiency in neuronal networks and might also interfere with the correct wiring during development.