Neuronal Mechanisms Recording the Stream of Consciousness-A Reappraisal of Wilder Penfield's (1891-1976) Concept of Experiential Phenomena Elicited by Electrical Stimulation of the Human Cortex.

Research on memory has been a major focus in the neurosciences over the past decades. An important advance was achieved by Wilder Penfield at the Montreal Neurological Institute, who reported from the 1930s to the 1950s about experiential phenomena induced by electrical brain stimulation in humans, implying neuronal causation of memory. Since then, neuroscientists have addressed the topic of memory from a range of subdisciplines; however, these reports by Penfield and his group as well as those on patient H. M. by Brenda Milner at the same institution continue to be referenced as groundbreaking. Further experimental work by Nobel laureates Eric Kandel and John O'Keefe, as well as by Edvard and May-Britt Moser related Penfield's patient documentation to experiential phenomena. However, our reassessment of Penfield's original patient documentation questions the stance that he had uncovered the "storehouse of memories." Human memory must be regarded more as context sensitive and as representative of an active reconstructive process, than as a simple recording of events. Hence, strategies aiming at naturalizing all phenomena of mind (including memory) to cellular and molecular mechanisms cannot convincingly refer to Penfield's electrophysiological studies alone as evidence that memories are solely caused by neuronal firing patterns.

Simultaneous quantitative susceptibility mapping and Flutemetamol-PET suggests local correlation of iron and ß-amyloid as an indicator of cognitive performance at high age.

The accumulation of ß-amyloid plaques is a hallmark of Alzheimer's disease (AD), and recently published data suggest that increased brain iron burden may reflect pathologies that synergistically contribute to the development of cognitive dysfunction. While preclinical disease stages are considered most promising for therapeutic intervention, the link between emerging AD-pathology and earliest clinical symptoms remains largely unclear. In the current study we therefore investigated local correlations between iron and ß-amyloid plaques, and their possible association with cognitive performance in healthy older adults. 116 older adults (mean age 75 ±â€¯7.4 years) received neuropsychological testing to calculate a composite cognitive score of performance in episodic memory, executive functioning, attention, language and communication. All participants were scanned on a combined PET-MRI instrument and were administered T1-sequences for anatomical mapping, quantitative susceptibility mapping (QSM) for assessing iron, and 18F-Flutemetamol-PET for estimating ß-amyloid plaque load. Biological parametric mapping (BPM) was used to generate masks indicating voxels with significant (p < 0.05) correlation between susceptibility and 18F-Flutemetamol-SUVR. We found a bilateral pattern of clusters characterized by a statistical relationship between magnetic susceptibility and 18F-Flutemetamol-SUVR, indicating local correlations between iron and ß-amyloid plaque deposition. For two bilateral clusters, located in the frontal and temporal cortex, significant relationships (p<0.05) between local ß-amyloid and the composite cognitive performance score could be observed. No relationship between whole-cortex ß-amyloid plaque load and cognitive performance was observable. Our data suggest that the local correlation of ß-amyloid plaque load and iron deposition may provide relevant information regarding cognitive performance of healthy older adults. Further studies are needed to clarify pathological correlates of the local interaction of ß-amyloid, iron and other causes of altered magnetic susceptibility.

Genetic ablation of the p66Shc adaptor protein reverses cognitive deficits and improves mitochondrial function in an APP transgenic mouse model of Alzheimer's disease.

The mammalian ShcA adaptor protein p66Shc is a key regulator of mitochondrial reactive oxygen species (ROS) production and has previously been shown to mediate amyloid ß (Aß)-peptide-induced cytotoxicity in vitro. Moreover, p66Shc is involved in mammalian longevity and lifespan determination as revealed in the p66Shc knockout mice, which are characterized by a 30% prolonged lifespan, lower ROS levels and protection from age-related impairment of physical and cognitive performance. In this study, we hypothesized a role for p66Shc in Aß-induced toxicity in vivo and investigated the effects of genetic p66Shc deletion in the PSAPP transgenic mice, an established Alzheimer's disease mouse model of ß-amyloidosis. p66Shc-ablated PSAPP mice were characterized by an improved survival and a complete rescue of Aß-induced cognitive deficits at the age of 15 months. Importantly, these beneficial effects on survival and cognitive performance were independent of Aß levels and amyloid plaque deposition, but were associated with improved brain mitochondrial respiration, a reversal of mitochondrial complex I dysfunction, restored adenosine triphosphate production and reduced ROS levels. The results of this study support a role for p66Shc in Aß-related mitochondrial dysfunction and oxidative damage in vivo, and suggest that p66Shc ablation may be a promising novel therapeutic strategy against Aß-induced toxicity and cognitive impairment.

T-cell brain infiltration and immature antigen-presenting cells in transgenic models of Alzheimer's disease-like cerebral amyloidosis.

Cerebral beta-amyloidosis, one of the pathological hallmarks of Alzheimer's disease (AD), elicits a well-characterised, microglia-mediated local innate immune response. In contrast, it is not clear whether cells of the adaptive immune system, in particular T-cells, react to cerebral amyloidosis in AD. Even though parenchymal T-cells have been described in post-mortem brains of AD patients, it is not known whether infiltrating T-cells are specifically recruited to the extracellular deposits of beta-amyloid, and whether they are locally activated into proliferating, effector cells upon interaction with antigen-presenting cells (APCs). To address these issues we have analysed by confocal microscopy and flow-cytometry the localisation and activation status of both T-cells and APCs in transgenic (tg) mice models of AD-like cerebral amyloidosis. Increased numbers of infiltrating T-cells were found in amyloid-burdened brain regions of tg mice, with concomitant up-regulation of endothelial adhesion molecules ICAM-1 and VCAM-1, compared to non-tg littermates. The infiltrating T-cells in tg brains did not co-localise with amyloid plaques, produced less interferon-gamma than those in controls and did not proliferate locally. Bona-fide dendritic cells were virtually absent from the brain parenchyma of both non-tg and tg mice, and APCs from tg brains showed an immature phenotype, with accumulation of MHC-II in intracellular compartments. These results indicate that cerebral amyloidosis promotes T-cell infiltration but interferes with local antigen presentation and T-cell activation. The inability of the brain immune surveillance to orchestrate a protective immune response to amyloid-beta peptide might contribute to the accumulation of amyloid in the progression of the disease.

Post-ischaemic silencing of p66Shc reduces ischaemia/reperfusion brain injury and its expression correlates to clinical outcome in stroke.

AIM: Constitutive genetic deletion of the adaptor protein p66(Shc) was shown to protect from ischaemia/reperfusion injury. Here, we aimed at understanding the molecular mechanisms underlying this effect in stroke and studied p66(Shc) gene regulation in human ischaemic stroke. METHODS AND RESULTS: Ischaemia/reperfusion brain injury was induced by performing a transient middle cerebral artery occlusion surgery on wild-type mice. After the ischaemic episode and upon reperfusion, small interfering RNA targeting p66(Shc) was injected intravenously. We observed that post-ischaemic p66(Shc) knockdown preserved blood-brain barrier integrity that resulted in improved stroke outcome, as identified by smaller lesion volumes, decreased neurological deficits, and increased survival. Experiments on primary human brain microvascular endothelial cells demonstrated that silencing of the adaptor protein p66(Shc) preserves claudin-5 protein levels during hypoxia/reoxygenation by reducing nicotinamide adenine dinucleotide phosphate oxidase activity and reactive oxygen species production. Further, we found that in peripheral blood monocytes of acute ischaemic stroke patients p66(Shc) gene expression is transiently increased and that this increase correlates with short-term neurological outcome. CONCLUSION: Post-ischaemic silencing of p66(Shc) upon reperfusion improves stroke outcome in mice while the expression of p66(Shc) gene correlates with short-term outcome in patients with ischaemic stroke.

Active vaccination with ankyrin G reduces ß-amyloid pathology in APP transgenic mice.

Serum antibodies against amyloid-ß peptide (Aß) in humans with or without diagnosis of Alzheimer's disease (AD) indicate the possibility of immune responses against brain antigens. In an unbiased screening for antibodies directed against brain proteins, we found in AD patients high serum levels of antibodies against the neuronal cytoskeletal protein ankyrin G (ankG); these correlated with slower rates of cognitive decline. Neuronal expression of ankG was higher in AD brains than in nondemented age-matched healthy control subjects. AnkG was present in exosomal vesicles, and it accumulated in ß-amyloid plaques. Active immunization with ankG of arcAß transgenic mice reduced brain ß-amyloid pathology and increased brain levels of soluble Aß(42). AnkG immunization induced a reduction in ß-amyloid pathology, also in Swedish transgenic mice(.) Anti-ankG monoclonal antibodies reduced Aß-induced loss of dendritic spines in hippocampal ArcAß organotypic cultures. Together, these data established a role for ankG in the human adaptive immune response against resident brain proteins, and they show that ankG immunization reduces brain ß-amyloid and its related neuropathology.

Regulation of microglial expression of integrins by poly(ADP-ribose) polymerase-1.

Excitotoxic brain lesions initially result in the primary destruction of brain parenchyma, after which microglial cells migrate towards the sites of injury. At these sites, the cells produce large quantities of oxygen radicals and cause secondary damage that accounts for most of the loss of brain function. Here we show that this microglial migration is strongly controlled in living brain tissue by expression of the integrin CD11a, regulated by the nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) through the formation of a nuclear PARP-NF-kappaB-protein complex. Downregulation of PARP or CD11a by transfection with antisense DNA abrogated microglial migration almost completely and prevented neurons from secondary damage.

Formation of neurofibrillary tangles in P301l tau transgenic mice induced by Abeta 42 fibrils.

beta-Amyloid plaques and neurofibrillary tangles (NFTs) are the defining neuropathological hallmarks of Alzheimer's disease, but their pathophysiological relation is unclear. Injection of beta-amyloid Abeta42 fibrils into the brains of P301L mutant tau transgenic mice caused fivefold increases in the numbers of NFTs in cell bodies within the amygdala from where neurons project to the injection sites. Gallyas silver impregnation identified NFTs that contained tau phosphorylated at serine 212/threonine 214 and serine 422. NFTs were composed of twisted filaments and occurred in 6-month-old mice as early as 18 days after Abeta42 injections. Our data support the hypothesis that Abeta42 fibrils can accelerate NFT formation in vivo.

Release of Alzheimer amyloid precursor derivatives stimulated by activation of muscarinic acetylcholine receptors.

Altered processing of the amyloid precursor protein (APP) is a central event in the formation of amyloid deposits in the brains of individuals with Alzheimer's disease. To investigate whether cellular APP processing is controlled by cell-surface neurotransmitter receptors, human embryonic kidney (293) cell lines were transfected with the genes for human brain muscarinic acetylcholine receptors. Stimulation of m1 and m3 receptor subtypes with carbachol increased the basal release of APP derivatives within minutes of treatment, indicating that preexisting APP is released in response to receptor activation. Receptor-activated APP release was blocked by staurosporine, suggesting that protein kinases mediate neurotransmitter receptor-controlled APP processing.

GABA and glutamate moderate beta-amyloid related functional connectivity in cognitively unimpaired old-aged adults.

BACKGROUND: Effects of beta-amyloid accumulation on neuronal function precede the clinical manifestation of Alzheimer's disease (AD) by years and affect distinct cognitive brain networks. As previous studies suggest a link between beta-amyloid and dysregulation of excitatory and inhibitory neurotransmitters, we aimed to investigate the impact of GABA and glutamate on beta-amyloid related functional connectivity. METHODS: 29 cognitively unimpaired old-aged adults (age = 70.03 ±â€¯5.77 years) were administered 11C-Pittsburgh Compound B (PiB) positron-emission tomography (PET), and MRI at 7 Tesla (7T) including blood oxygen level dependent (BOLD) functional MRI (fMRI) at rest for measuring static and dynamic functional connectivity. An advanced 7T MR spectroscopic imaging (MRSI) sequence based on the free induction decay acquisition localized by outer volume suppression' (FIDLOVS) technology was used for gray matter specific measures of GABA and glutamate in the posterior cingulate and precuneus (PCP) region. RESULTS: GABA and glutamate MR-spectra indicated significantly higher levels in gray matter than in white matter. A global effect of beta-amyloid on functional connectivity in the frontal, occipital and inferior temporal lobes was observable. Interactive effects of beta-amyloid with gray matter GABA displayed positive PCP connectivity to the frontomedial regions, and the interaction of beta-amyloid with gray matter glutamate indicated positive PCP connectivity to frontal and cerebellar regions. Furthermore, decreased whole-brain but increased fronto-occipital and temporo-parietal dynamic connectivity was found, when GABA interacted with regional beta-amyloid deposits in the amygdala, frontal lobe, hippocampus, insula and striatum. CONCLUSIONS: GABA, and less so glutamate, may moderate beta-amyloid related functional connectivity. Additional research is needed to better characterize their interaction and potential impact on AD.

Impaired spatial reference memory and increased exploratory behavior in P301L tau transgenic mice.

The neuropathological hallmark shared between Alzheimer's disease (AD) and familial frontotemporal dementia (FTDP-17) are neurofibrillary tangles (NFT) which are composed of filamentous aggregates of the microtubule-associated protein tau. Their formation has been reproduced in transgenic mice, which express the FTDP-17-associated mutation P301L of tau. In these mice, tau aggregates are found in many brain areas including the hippocampus and the amygdala, both of which are characterized by NFT formation in AD. Previous studies using an amygdala-specific test battery revealed an increase in exploratory behavior and an accelerated extinction of conditioned taste aversion in these mice. Here, we assessed P301L mice in behavioral tests known to depend on an intact hippocampus. Morris water maze and Y-maze revealed intact spatial working memory but impairment in spatial reference memory at 6 and 11 months of age. In addition, a modest disinhibition of exploratory behavior at 6 months of age was confirmed in the open field and the elevated O-maze and was more pronounced during aging.

New molecules for hippocampal development.

Pathfinding by developing axons towards their proper targets is an essential step in establishing appropriate neuronal connections. Recent work involving cell culture assays and molecular biology strategies, including knockout animals, strongly indicates that a complex network of guidance signals regulates the formation of hippocampal connections during development. Outgrowing axons are routed towards the hippocampal formation by specific expression of long-range cues, which include secreted class 3 semaphorins, netrin 1 and Slit proteins. Local membrane- or substrate-anchored molecules, such as ligands of the ephrin A subclass, provide layer-specific positional information. Understanding the molecular mechanisms that underlie axonal guidance during hippocampal development might be of importance in making therapeutic use of sprouting fibers, which are produced following the loss of afferents in CNS lesion.

Morphological features of the entorhinal-hippocampal connection.

The goal of this review in an overview of the structural elements of the entorhinal-hippocampal connection. The development of the dendrites of hippocampal neurons will be outlined in relation to afferent pathway specificity and the mature dendritic structure compared. Interneurons will be contrasted to pyramidal cells in terms of processing of physiological signals and convergence and divergence in control of hippocampal circuits. Mechanisms of axonal guidance and target recognition, target structures, the involvement of receptor distribution on hippocampal dendrites and the involvement of non-neuronal cellular elements in the establishment of specific connections will be presented. Mechanisms relevant for the maintenance of shape and morphological specializations of hippocampal dendrites will be reviewed. One of the significant contexts in which to view these structural elements is the degree of plasticity in which they participate, during development and origination of dendrites, mature synaptic plasticity and after lesions, when the cells must continue to maintain and reconstitute function, to remain part of the circuitry in the hippocampus. This review will be presented in four main sections: (1) interneurons-development, role in synchronizing influence and hippocampal network functioning; (2) principal cells in CA1, CA3 and dentate gyrus regions-their development, function in terms of synaptic integration, differentiating structure and alterations with lesions; (3) glia and glia/neuronal interactions-response to lesions and developmental guidance mechanisms; and (4) network and circuit aspects of hippocampal morphology and functioning. Finally, the interwoven role of these various elements participating in hippocampal network function will be discussed.

First one in, last one out: the role of gabaergic transmission in generation and degeneration.

This paper is the result of discussions between scientists working in widely separated areas, united by an interest in the hippocampus. The discussions focused on the possible role of GABA in the development and maturation of the hippocampus and in neurodegeneration in Alzheimer's disease (AD). GABA neurons are among the first to differentiate in the hippocampus and the properties of GABA neurotransmission in the developing hippocampus are distinct from those in the adult. GABAergic transmission may play a role in the clustering and maturation of GABA receptors, as well as of receptors for other neurotransmitters. The development and maturation of synaptic connections involves changes in the organization of the cytoskeleton, and mechanical force generation is probably required to establish appropriate points of contact. This generation of force may require coupling of specific receptors to the cytoskeleton through specialized proteins. In AD, much of the developmental process is progressively unraveled in the hippocampus, as afferent fibers, most notably from entorhinal excitatory neurons and from basal forebrain cholinergic cells, degenerate. This denervation undoubtedly has consequences for receptor systems, dendritic morphology and the underlying cytoskeleton. GABA neurons remain in the AD hippocampus, and may actually contribute to abnormal firing and degeneration of remaining pyramidal neurons. This attempt to bring together data from different areas of research has allowed the development of a scheme which identifies significant specific gaps in our knowledge, which could be readily filled by focused experimental work.

Colocalization of cerebral iron with Amyloid beta in Mild Cognitive Impairment.

Quantitative Susceptibility Mapping (QSM) MRI at 7 Tesla and 11-Carbon Pittsburgh-Compound-B PET were used for investigating the relationship between brain iron and Amyloid beta (Aß) plaque-load in a context of increased risk for Alzheimer's disease (AD), as reflected by the Apolipoprotein E ε4 (APOE-e4) allele and mild cognitive impairment (MCI) in elderly subjects. Carriers of APOE-e4 with normal cognition had higher cortical Aß-plaque-load than non-carriers. In MCI an association between APOE-e4 and higher Aß-plaque-load was observable both for cortical and subcortical brain-regions. APOE-e4 and MCI was also associated with higher cortical iron. Moreover, cerebral iron significantly affected functional coupling, and was furthermore associated with increased Aß-plaque-load (R2-adjusted = 0.80, p < 0.001) and APOE-e4 carrier status (p < 0.001) in MCI. This study confirms earlier reports on an association between increased brain iron-burden and risk for neurocognitive dysfunction due to AD, and indicates that disease-progression is conferred by spatial colocalization of brain iron deposits with Aß-plaques.

A role for the Eph ligand ephrin-A3 in entorhino-hippocampal axon targeting.

Neurons of layers II and III of the entorhinal cortex constitute the major afferent connection of the hippocampus. The molecular mechanisms that target the entorhinal axons to specific layers in the hippocampus are not known. EphA5, a member of the Eph receptor family, which has been shown to play critical roles in axon guidance, is expressed in the entorhinal cortex, the origin of the perforant pathway. In addition, ligands that interact with EphA5 are expressed in distinct hippocampal regions during development of the entorhino-hippocampal projection. Of these ligands, ephrin-A3 mRNA is localized both in the granular cell layer of the dentate gyrus and in the pyramidal cell layer of the cornu ammonis, whereas ephrin-A5 mRNA is only expressed in the pyramidal cell layer of the cornu ammonis. In the dentate gyrus, the ligand protein is not present in the termination zone of the entorhinal efferents (the outer molecular layer of the dentate gyrus) but is concentrated in the inner molecular layer into which entorhinal efferents do not grow. We used outgrowth and stripe assays to test the effects of ephrin-A3 and ephrin-A5 on the outgrowth behavior of entorhinal axons. This functional analysis revealed that entorhinal neurites were repelled by ephrin-A3 but not by ephrin-A5. These observations suggest that ephrin-A3 plays an important role in the layer-specific termination of the perforant pathway and that this ligand may interact with the EphA5 receptor to restrict entorhinal axon terminals in the outer molecular layer of the dentate gyrus.

The human DIMINUTO/DWARF1 homolog seladin-1 confers resistance to Alzheimer's disease-associated neurodegeneration and oxidative stress.

In Alzheimer's disease (AD) brains, selected populations of neurons degenerate heavily, whereas others are frequently spared from degeneration. To address the cellular basis for this selective vulnerability of neurons in distinct brain regions, we compared gene expression between the severely affected inferior temporal lobes and the mostly unaffected fronto-parietal cortices by using an mRNA differential display. We identified seladin-1, a novel gene, which was downregulated in large pyramidal neurons in vulnerable regions in AD but not control brains. Seladin-1 is a human homolog of the DIMINUTO/DWARF1 gene described in plants and Caenorhabditis elegans. Its sequence shares similarities with flavin-adenin-dinucleotide (FAD)-dependent oxidoreductases. In human control brain, seladin-1 was highly expressed in almost all neurons. In PC12 cell clones that were selected for resistance against AD-associated amyloid-beta peptide (Abeta)-induced toxicity, both mRNA and protein levels of seladin-1 were approximately threefold higher as compared with the non-resistant wild-type cells. Functional expression of seladin-1 in human neuroglioma H4 cells resulted in the inhibition of caspase 3 activation after either Abeta-mediated toxicity or oxidative stress and protected the cells from apoptotic cell death. In apoptotic cells, however, endogenous seladin-1 was cleaved to a 40 kDa derivative in a caspase-dependent manner. These results establish that seladin-1 is an important factor for the protection of cells against Abeta toxicity and oxidative stress, and they suggest that seladin-1 may be involved in the regulation of cell survival and death. Decreased expression of seladin-1 in specific neurons may be a cause for selective vulnerability in AD.

Distinct role of protein phosphatase 2A subunit Calpha in the regulation of E-cadherin and beta-catenin during development.

Protein phosphatase 2A (PP2A) plays central roles in development, cell growth and transformation. Inactivation of the gene encoding the PP2A catalytic subunit Calpha by gene targeting generates a lethal embryonic phenotype. No mesoderm is formed in Calpha(-/-) embryos. Here, we found that during normal early embryonic development Calpha was predominantly present at the plasma membrane whereas the highly homologous isoform Cbeta was localized to the cytoplasm and nuclei, suggesting the inability of Cbeta to compensate for vital functions of Calpha in Calpha(-/-) embryos. In addition, PP2A was found in a complex containing the PP2A substrates E-cadherin and beta-catenin. In Calpha(-/-) embryos, E-cadherin and beta-catenin were redistributed from the plasma membrane to the cytosol. Cytosolic concentrations of beta-catenin were low. Our results suggest that Calpha is required for stabilization of E-cadherin/beta-catenin complexes at the plasma membrane.

Gene expression screening in Xenopus identifies molecular pathways, predicts gene function and provides a global view of embryonic patterning.

In a large-scale gene expression screen 1765 randomly picked cDNAs were analyzed by whole-mount in situ hybridization in Xenopus embryos. Two hundred and seventy three unique, differentially expressed genes were identified, 204 of which are novel in Xenopus. Partial DNA sequences and expression patterns were documented and assembled into a database, 'AXelDB'. Approximately 30% of cDNAs analyzed represent differentially expressed genes and about 5% show highly regionalized expression. Novel marker genes and potential developmental regulators were found. Differential expression of mitochondrial genes was observed. Marker genes were used to study regionalization of the entire gastrula as well as the tail forming region and the epidermis of the tailbud embryo. Four 'synexpression' groups representing genes with shared, complex expression pattern that predict molecular pathways involved in patterning and differentiation were identified. According to their probable functional significance these groups are designated as Delta1, Bmp4, ER-import and Chromatin group. Within synexpression groups, a likely function of genes without sequence similarity can be predicted. The results indicate that synexpression groups have strong prognostic value. A cluster analysis was made by comparing gene expression patterns to derive a novel parameter, 'tissue relatedness'. In conclusion, this study describes a semi-functional approach to investigate genes expressed during early development and provides global insight into embryonic patterning.

Molecular and functional analysis of hyperpolarization-activated pacemaker channels in the hippocampus after entorhinal cortex lesion.

Differential display of hippocampal tissue after entorhinal cortex lesion (ECL) revealed decreases in mRNA encoding the neuronal hyperpolarization-activated, cyclic nucleotide-gated channel HCN1. In situ hybridization confirmed that hippocampal transcripts of HCN1, but not HCN2/3/4, are down-regulated after ECL. Expression recovered at approximately 21 days after lesion (dal). Immunohistochemistry demonstrated a corresponding regulation of HCN1 protein expression in CA1-CA3 dendrites, hilar mossy cells and interneurons, and granule cells. Patch-clamp recordings in the early phase after lesion from mossy cells and hilar interneurons revealed an increase in the fast time constant of current activation and a profound negative shift in voltage activation of Ih. Whereas current activation recovered at 30 dal, the voltage activation remained hyperpolarized in mossy cells and hilar interneurons. Granule cells, however, were devoid of any detectable somatic Ih currents. Hence, denervation of the hippocampus decreases HCN1 and concomitantly the Ih activity in hilar neurons, and the recovery of h-current activation kinetics occurs parallel to postlesion sprouting.