Publications by authors named "Javier DeFelipe"

174 Publications

Dendritic spines are lost in clusters in Alzheimer's disease.

Sci Rep 2021 Jun 11;11(1):12350. Epub 2021 Jun 11.

Laboratorio Cajal de Circuitos Corticales (CTB), Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Campus Montegancedo S/N, Pozuelo de Alarcon, 28223, Madrid, Spain.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by a deterioration of neuronal connectivity. The pathological accumulation of tau in neurons is one of the hallmarks of AD and has been connected to the loss of dendritic spines of pyramidal cells, which are the major targets of cortical excitatory synapses and key elements in memory storage. However, the detailed mechanisms underlying the loss of dendritic spines in individuals with AD are still unclear. Here, we used graph-theory approaches to compare the distribution of dendritic spines from neurons with and without tau pathology of AD individuals. We found that the presence of tau pathology determines the loss of dendritic spines in clusters, ruling out alternative models where spine loss occurs at random locations. Since memory storage has been associated with synaptic clusters, the present results provide a new insight into the mechanisms by which tau drives synaptic damage in AD, paving the way to memory deficits through alterations of spine organization.
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http://dx.doi.org/10.1038/s41598-021-91726-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8196005PMC
June 2021

Neuronal excitation/inhibition imbalance: core element of a translational perspective on Alzheimer pathophysiology.

Ageing Res Rev 2021 Aug 21;69:101372. Epub 2021 May 21.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Pozuelo de Alarcón, 28223, Madrid, Spain; CIBERNED, Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas, 28031, Madrid, Spain; Instituto Cajal, Consejo Superior de Investigaciones Científicas, Arce 37 28002, Madrid, Spain.

Our incomplete understanding of the link between Alzheimer's Disease pathology and symptomatology is a crucial obstacle for therapeutic success. Recently, translational studies have begun to connect the dots between protein alterations and deposition, brain network dysfunction and cognitive deficits. Disturbance of neuronal activity, and in particular an imbalance in underlying excitation/inhibition (E/I), appears early in AD, and can be regarded as forming a central link between structural brain pathology and cognitive dysfunction. While there are emerging (non-)pharmacological options to influence this imbalance, the complexity of human brain dynamics has hindered identification of an optimal approach. We suggest that focusing on the integration of neurophysiological aspects of AD at the micro-, meso- and macroscale, with the support of computational network modeling, can unite fundamental and clinical knowledge, provide a general framework, and suggest rational therapeutic targets.
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http://dx.doi.org/10.1016/j.arr.2021.101372DOI Listing
August 2021

Three-Dimensional Synaptic Organization of Layer III of the Human Temporal Neocortex.

Cereb Cortex 2021 May 17. Epub 2021 May 17.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Madrid 28223, Spain.

In the present study, we have used focused ion beam/scanning electron microscopy (FIB/SEM) to perform a study of the synaptic organization of layer III of Brodmann's area 21 in human tissue samples obtained from autopsies and biopsies. We analyzed the synaptic density, 3D spatial distribution, and type (asymmetric/symmetric), as well as the size and shape of each synaptic junction of 4945 synapses that were fully reconstructed in 3D. Significant differences in the mean synaptic density between autopsy and biopsy samples were found (0.49 and 0.66 synapses/μm3, respectively). However, in both types of samples (autopsy and biopsy), the asymmetric:symmetric ratio was similar (93:7) and most asymmetric synapses were established on dendritic spines (75%), while most symmetric synapses were established on dendritic shafts (85%). We also compared several electron microscopy methods and analysis tools to estimate the synaptic density in the same brain tissue. We have shown that FIB/SEM is much more reliable and robust than the majority of the other commonly used EM techniques. The present work constitutes a detailed description of the synaptic organization of cortical layer III. Further studies on the rest of the cortical layers are necessary to better understand the functional organization of this temporal cortical region.
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http://dx.doi.org/10.1093/cercor/bhab120DOI Listing
May 2021

Variation in Pyramidal Cell Morphology Across the Human Anterior Temporal Lobe.

Cereb Cortex 2021 Mar 16. Epub 2021 Mar 16.

Instituto Cajal, Consejo Superior de Investigaciones Científicas, Madrid 28002, Spain.

Pyramidal neurons are the most abundant and characteristic neuronal type in the cerebral cortex and their dendritic spines are the main postsynaptic elements of cortical excitatory synapses. Previous studies have shown that pyramidal cell structure differs across layers, cortical areas, and species. However, within the human cortex, the pyramidal dendritic morphology has been quantified in detail in relatively few cortical areas. In the present work, we performed intracellular injections of Lucifer Yellow at several distances from the temporal pole. We found regional differences in pyramidal cell morphology, which showed large inter-individual variability in most of the morphological variables measured. However, some values remained similar in all cases. The smallest and least complex cells in the most posterior temporal region showed the greatest dendritic spine density. Neurons in the temporal pole showed the greatest sizes with the highest number of spines. Layer V cells were larger, more complex, and had a greater number of dendritic spines than those in layer III. The present results suggest that, while some aspects of pyramidal structure are conserved, there are specific variations across cortical regions, and species.
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http://dx.doi.org/10.1093/cercor/bhab034DOI Listing
March 2021

Metabolic Changes in Brain Slices over Time: a Multiplatform Metabolomics Approach.

Mol Neurobiol 2021 Mar 2. Epub 2021 Mar 2.

Centro de Metabolómica y Bioanálisis (CEMBIO), Facultad de Farmacia, Universidad San Pablo-CEU, CEU Universities, Urbanización Montepríncipe, 28660, Boadilla del Monte, Spain.

Brain slice preparations are widely used for research in neuroscience. However, a high-quality preparation is essential and there is no consensus regarding stable parameters that can be used to define the status of the brain slice preparation after its collection at different time points. Thus, it is critical to fully characterize the experimental conditions for ex vivo studies using brain slices for electrophysiological recording. In this study, we used a multiplatform (LC-MS and GC-MS) untargeted metabolomics-based approach to shed light on the metabolome and lipidome changes taking place at different time intervals during the brain slice preparation process. We have found significant modifications in the levels of 300 compounds, including several lipid classes and their derivatives, as well as metabolites involved in the GABAergic pathway and the TCA cycle. All these preparation-dependent changes in the brain biochemistry related to the time interval should be taken into consideration for future studies to facilitate non-biased interpretations of the experimental results.
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http://dx.doi.org/10.1007/s12035-020-02264-yDOI Listing
March 2021

High levels of 27-hydroxycholesterol results in synaptic plasticity alterations in the hippocampus.

Sci Rep 2021 Feb 12;11(1):3736. Epub 2021 Feb 12.

Department of Neurobiology, Care Sciences and Society, Division of Neurogeriatrics, Center for Alzheimer Research, Karolinska Institutet, Stockholm, Sweden.

Alterations in brain cholesterol homeostasis in midlife are correlated with a higher risk of developing Alzheimer's disease (AD). However, global cholesterol-lowering therapies have yielded mixed results when it comes to slowing down or preventing cognitive decline in AD. We used the transgenic mouse model Cyp27Tg, with systemically high levels of 27-hydroxycholesterol (27-OH) to examine long-term potentiation (LTP) in the hippocampal CA1 region, combined with dendritic spine reconstruction of CA1 pyramidal neurons to detect morphological and functional synaptic alterations induced by 27-OH high levels. Our results show that elevated 27-OH levels lead to enhanced LTP in the Schaffer collateral-CA1 synapses. This increase is correlated with abnormally large dendritic spines in the stratum radiatum. Using immunohistochemistry for synaptopodin (actin-binding protein involved in the recruitment of the spine apparatus), we found a significantly higher density of synaptopodin-positive puncta in CA1 in Cyp27Tg mice. We hypothesize that high 27-OH levels alter synaptic potentiation and could lead to dysfunction of fine-tuned processing of information in hippocampal circuits resulting in cognitive impairment. We suggest that these alterations could be detrimental for synaptic function and cognition later in life, representing a potential mechanism by which hypercholesterolemia could lead to alterations in memory function in neurodegenerative diseases.
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http://dx.doi.org/10.1038/s41598-021-83008-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7881004PMC
February 2021

Three-dimensional analysis of synaptic organization in the hippocampal CA1 field in Alzheimer's disease.

Brain 2021 03;144(2):553-573

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, 28223 Pozuelo de Alarcón, Madrid, Spain.

Alzheimer's disease is the most common form of dementia, characterized by a persistent and progressive impairment of cognitive functions. Alzheimer's disease is typically associated with extracellular deposits of amyloid-β peptide and accumulation of abnormally phosphorylated tau protein inside neurons (amyloid-β and neurofibrillary pathologies). It has been proposed that these pathologies cause neuronal degeneration and synaptic alterations, which are thought to constitute the major neurobiological basis of cognitive dysfunction in Alzheimer's disease. The hippocampal formation is especially vulnerable in the early stages of Alzheimer's disease. However, the vast majority of electron microscopy studies have been performed in animal models. In the present study, we performed an extensive 3D study of the neuropil to investigate the synaptic organization in the stratum pyramidale and radiatum in the CA1 field of Alzheimer's disease cases with different stages of the disease, using focused ion beam/scanning electron microscopy (FIB/SEM). In cases with early stages of Alzheimer's disease, the synapse morphology looks normal and we observed no significant differences between control and Alzheimer's disease cases regarding the synaptic density, the ratio of excitatory and inhibitory synapses, or the spatial distribution of synapses. However, differences in the distribution of postsynaptic targets and synaptic shapes were found. Furthermore, a lower proportion of larger excitatory synapses in both strata were found in Alzheimer's disease cases. Individuals in late stages of the disease suffered the most severe synaptic alterations, including a decrease in synaptic density and morphological alterations of the remaining synapses. Since Alzheimer's disease cases show cortical atrophy, our data indicate a reduction in the total number (but not the density) of synapses at early stages of the disease, with this reduction being much more accentuated in subjects with late stages of Alzheimer's disease. The observed synaptic alterations may represent a structural basis for the progressive learning and memory dysfunctions seen in Alzheimer's disease cases.
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http://dx.doi.org/10.1093/brain/awaa406DOI Listing
March 2021

Neuronize v2: Bridging the Gap Between Existing Proprietary Tools to Optimize Neuroscientific Workflows.

Front Neuroanat 2020 6;14:585793. Epub 2020 Oct 6.

Department of Computer Science, Universidad Rey Juan Carlos, Madrid, Spain.

Knowledge about neuron morphology is key to understanding brain structure and function. There are a variety of software tools that are used to segment and trace the neuron morphology. However, these tools usually utilize proprietary formats. This causes interoperability problems since the information extracted with one tool cannot be used in other tools. This article aims to improve neuronal reconstruction workflows by facilitating the interoperability between two of the most commonly used software tools-Neurolucida (NL) and Imaris (Filament Tracer). The new functionality has been included in an existing tool-Neuronize-giving rise to its second version. Neuronize v2 makes it possible to automatically use the data extracted with Imaris Filament Tracer to generate a tracing with dendritic spine information that can be read directly by NL. It also includes some other new features, such as the ability to unify and/or correct inaccurately-formed meshes (i.e., dendritic spines) and to calculate new metrics. This tool greatly facilitates the process of neuronal reconstruction, bridging the gap between existing proprietary tools to optimize neuroscientific workflows.
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http://dx.doi.org/10.3389/fnana.2020.585793DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7646287PMC
October 2020

Comparing basal dendrite branches in human and mouse hippocampal CA1 pyramidal neurons with Bayesian networks.

Sci Rep 2020 10 29;10(1):18592. Epub 2020 Oct 29.

Computational Intelligence Group, Departamento de Inteligencia Artificial, Universidad Politécnica de Madrid, Boadilla del Monte, 28660, Madrid, Spain.

Pyramidal neurons are the most common cell type in the cerebral cortex. Understanding how they differ between species is a key challenge in neuroscience. A recent study provided a unique set of human and mouse pyramidal neurons of the CA1 region of the hippocampus, and used it to compare the morphology of apical and basal dendritic branches of the two species. The study found inter-species differences in the magnitude of the morphometrics and similarities regarding their variation with respect to morphological determinants such as branch type and branch order. We use the same data set to perform additional comparisons of basal dendrites. In order to isolate the heterogeneity due to intrinsic differences between species from the heterogeneity due to differences in morphological determinants, we fit multivariate models over the morphometrics and the determinants. In particular, we use conditional linear Gaussian Bayesian networks, which provide a concise graphical representation of the independencies and correlations among the variables. We also extend the previous study by considering additional morphometrics and by formally testing whether a morphometric increases or decreases with the distance from the soma. This study introduces a multivariate methodology for inter-species comparison of morphology.
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http://dx.doi.org/10.1038/s41598-020-73617-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7596062PMC
October 2020

A community-based transcriptomics classification and nomenclature of neocortical cell types.

Nat Neurosci 2020 12;23(12):1456-1468

Vanderbilt University, Nashville, TN, USA.

To understand the function of cortical circuits, it is necessary to catalog their cellular diversity. Past attempts to do so using anatomical, physiological or molecular features of cortical cells have not resulted in a unified taxonomy of neuronal or glial cell types, partly due to limited data. Single-cell transcriptomics is enabling, for the first time, systematic high-throughput measurements of cortical cells and generation of datasets that hold the promise of being complete, accurate and permanent. Statistical analyses of these data reveal clusters that often correspond to cell types previously defined by morphological or physiological criteria and that appear conserved across cortical areas and species. To capitalize on these new methods, we propose the adoption of a transcriptome-based taxonomy of cell types for mammalian neocortex. This classification should be hierarchical and use a standardized nomenclature. It should be based on a probabilistic definition of a cell type and incorporate data from different approaches, developmental stages and species. A community-based classification and data aggregation model, such as a knowledge graph, could provide a common foundation for the study of cortical circuits. This community-based classification, nomenclature and data aggregation could serve as an example for cell type atlases in other parts of the body.
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http://dx.doi.org/10.1038/s41593-020-0685-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7683348PMC
December 2020

Estimation of the number of synapses in the hippocampus and brain-wide by volume electron microscopy and genetic labeling.

Sci Rep 2020 08 19;10(1):14014. Epub 2020 Aug 19.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, 28223, Pozuelo de Alarcón, Madrid, Spain.

Determining the number of synapses that are present in different brain regions is crucial to understand brain connectivity as a whole. Membrane-associated guanylate kinases (MAGUKs) are a family of scaffolding proteins that are expressed in excitatory glutamatergic synapses. We used genetic labeling of two of these proteins (PSD95 and SAP102), and Spinning Disc confocal Microscopy (SDM), to estimate the number of fluorescent puncta in the CA1 area of the hippocampus. We also used FIB-SEM, a three-dimensional electron microscopy technique, to calculate the actual numbers of synapses in the same area. We then estimated the ratio between the three-dimensional densities obtained with FIB-SEM (synapses/µm) and the bi-dimensional densities obtained with SDM (puncta/100 µm). Given that it is impractical to use FIB-SEM brain-wide, we used previously available SDM data from other brain regions and we applied this ratio as a conversion factor to estimate the minimum density of synapses in those regions. We found the highest densities of synapses in the isocortex, olfactory areas, hippocampal formation and cortical subplate. Low densities were found in the pallidum, hypothalamus, brainstem and cerebellum. Finally, the striatum and thalamus showed a wide range of synapse densities.
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http://dx.doi.org/10.1038/s41598-020-70859-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7438319PMC
August 2020

Three-dimensional synaptic organization of the human hippocampal CA1 field.

Elife 2020 07 21;9. Epub 2020 Jul 21.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Madrid, Spain.

The hippocampal CA1 field integrates a wide variety of subcortical and cortical inputs, but its synaptic organization in humans is still unknown due to the difficulties involved studying the human brain via electron microscope techniques. However, we have shown that the 3D reconstruction method using Focused Ion Beam/Scanning Electron Microscopy (FIB/SEM) can be applied to study in detail the synaptic organization of the human brain obtained from autopsies, yielding excellent results. Using this technology, 24,752 synapses were fully reconstructed in CA1, revealing that most of them were excitatory, targeting dendritic spines and displaying a macular shape, regardless of the layer examined. However, remarkable differences were observed between layers. These data constitute the first extensive description of the synaptic organization of the neuropil of the human CA1 region.
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http://dx.doi.org/10.7554/eLife.57013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7375818PMC
July 2020

Area-Specific Synapse Structure in Branched Posterior Nucleus Axons Reveals a New Level of Complexity in Thalamocortical Networks.

J Neurosci 2020 03 13;40(13):2663-2679. Epub 2020 Feb 13.

Department of Anatomy and Neuroscience, School of Medicine, Autónoma de Madrid University, 28029 Madrid, Spain,

Thalamocortical posterior nucleus (Po) axons innervating the vibrissal somatosensory (S1) and motor (MC) cortices are key links in the brain neuronal network that allows rodents to explore the environment whisking with their motile snout vibrissae. Here, using fine-scale high-end 3D electron microscopy, we demonstrate in adult male C57BL/6 wild-type mice marked differences between MC versus S1 Po synapses in (1) bouton and active zone size, (2) neurotransmitter vesicle pool size, (3) distribution of mitochondria around synapses, and (4) proportion of synapses established on dendritic spines and dendritic shafts. These differences are as large, or even more pronounced, than those between Po and ventro-posterior thalamic nucleus synapses in S1. Moreover, using single-axon transfection labeling, we demonstrate that the above differences actually occur on the MC versus the S1 branches of individual Po cell axons that innervate both areas. Along with recently-discovered divergences in efficacy and plasticity, the synaptic structure differences reported here thus reveal a new subcellular level of complexity. This is a finding that upends current models of thalamocortical circuitry, and that might as well illuminate the functional logic of other branched projection axon systems. Many long-distance brain connections depend on neurons whose branched axons target separate regions. Using 3D electron microscopy and single-cell transfection, we investigated the mouse Posterior thalamic nucleus (Po) cell axons that simultaneously innervate motor and sensory areas of the cerebral cortex involved in whisker movement control. We demonstrate significant differences in the size of the boutons made in each area by individual Po axons, as well as in functionally-relevant parameters in the composition of their synapses. In addition, we found similarly large differences between the synapses of Po versus ventral posteromedial thalamic nucleus axons in the whisker sensory cortex. Area-specific synapse structure in individual axons implies a new, unsuspected level of complexity in long-distance brain connections.
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http://dx.doi.org/10.1523/JNEUROSCI.2886-19.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7096142PMC
March 2020

Volume Electron Microscopy Study of the Relationship Between Synapses and Astrocytes in the Developing Rat Somatosensory Cortex.

Cereb Cortex 2020 05;30(6):3800-3819

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, 28223 Pozuelo de Alarcón, Madrid, Spain.

In recent years, numerous studies have shown that astrocytes play an important role in neuronal processing of information. One of the most interesting findings is the existence of bidirectional interactions between neurons and astrocytes at synapses, which has given rise to the concept of "tripartite synapses" from a functional point of view. We used focused ion beam milling and scanning electron microscopy (FIB/SEM) to examine in 3D the relationship of synapses with astrocytes that were previously labeled by intracellular injections in the rat somatosensory cortex. We observed that a large number of synapses (32%) had no contact with astrocytic processes. The remaining synapses (68%) were in contact with astrocytic processes, either at the level of the synaptic cleft (44%) or with the pre- and/or post-synaptic elements (24%). Regarding synaptic morphology, larger synapses with more complex shapes were most frequently found within the population that had the synaptic cleft in contact with astrocytic processes. Furthermore, we observed that although synapses were randomly distributed in space, synapses that were free of astrocytic processes tended to form clusters. Overall, at least in the developing rat neocortex, the concept of tripartite synapse only seems to be applicable to a subset of synapses.
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http://dx.doi.org/10.1093/cercor/bhz343DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7233003PMC
May 2020

Calbindin immunostaining in the CA1 hippocampal pyramidal cell layer of the human and mouse: A comparative study.

J Chem Neuroanat 2020 Jan 13;104:101745. Epub 2020 Jan 13.

Departamento de Neurobiología Funcional y de Sistemas, Instituto Cajal, CSIC, Madrid, Spain; Laboratorio Cajal de Circuitos Corticales (CTB), Universidad Politécnica de Madrid, Madrid, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), ISCIII, Madrid, Spain. Electronic address:

Immunostaining for calbindin (CB) is commonly used to label particular populations of neurons. Recently, it has been shown that the CA1 pyramidal cells in the mouse can be subdivided along the radial axis into superficial and deep pyramidal cells and that this segregation in the radial axis may represent a general principle of structural and functional organization of the hippocampus. One of the most widely used markers of the superficial pyramidal cells is CB. However, this laminar segregation of pyramidal cells has not been reported in the human CA1 using CB immunostaining. The problem is that the different pattern of CB immunostaining observed in the mouse compared to the human could be explained by technical features, of which one of the most important is the postmortem time (PT) delay typical of the brain tissue obtained from humans. In the present study, we have studied the influences of PT delays and fixation procedures and we found that the clear differences found between the CA1 of the human and mouse do not depend on the fixation, but represent actual species-specific differences. These remarkable differences between species should be taken into consideration when making interpretations in translational studies from mouse to human brains.
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http://dx.doi.org/10.1016/j.jchemneu.2020.101745DOI Listing
January 2020

Subregional Density of Neurons, Neurofibrillary Tangles and Amyloid Plaques in the Hippocampus of Patients With Alzheimer's Disease.

Front Neuroanat 2019 19;13:99. Epub 2019 Dec 19.

Cajal Laboratory of Cortical Circuits, Centre for Biomedical Technology (CTB), Universidad Politécnica de Madrid, Madrid, Spain.

A variety of anatomical alterations have been reported in the hippocampal formation of patients with Alzheimer's Disease (AD) and these alterations have been correlated with cognitive symptoms in the early stages of the disease. Major hallmarks in AD are the presence of paired helical filaments of tau protein (PHF) within neurons, also known as neurofibrillary tangles (NFTs), and aggregates of amyloid-β protein (Aβ) which form plaques in the extracellular space. Nevertheless, how the density of plaques and NFTs relate to the severity of cell loss and cognitive decline is not yet clear. The aim of the present study was to further examine the possible relationship of both Aβ plaques and NFTs with neuronal loss in several hippocampal fields (DG, CA3, CA1, and subiculum) of 11 demented AD patients. For this purpose, using stereological techniques, we compared neuronal densities (Nissl-stained, and immunoreactive neurons for NeuN) with: (i) numbers of neurons immunostained for two isoforms of PHF (PHF and PHF); and (ii) number of Aβ plaques. We found that CA1 showed the highest number of NFTs and Aβ plaques, whereas DG and CA3 displayed the lowest number of these markers. Furthermore, AD patients showed a variable neuronal loss in CA1 due to tangle-related cell death, which seems to correlate with the presence of extracellular tangles.
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http://dx.doi.org/10.3389/fnana.2019.00099DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6930895PMC
December 2019

The Golgi Apparatus of Neocortical Glial Cells During Hibernation in the Syrian Hamster.

Front Neuroanat 2019 19;13:92. Epub 2019 Nov 19.

Laboratorio Cajal de Circuitos Corticales (CTB), Universidad Politécnica de Madrid, Madrid, Spain.

Hibernating mammals undergo torpor periods characterized by a general decrease in body temperature, metabolic rate, and brain activity accompanied by complex adaptive brain changes that appear to protect the brain from extreme conditions of hypoxia and low temperatures. These processes are accompanied by morphological and neurochemical changes in the brain including those in cortical neurons such as the fragmentation and reduction of the Golgi apparatus (GA), which both reverse a few hours after arousal from the torpor state. In the present study, we characterized - by immunofluorescence and confocal microscopy - the GA of cortical astrocytes, oligodendrocytes, and microglial cells in the Syrian hamster, which is a facultative hibernator. We also show that after artificial induction of hibernation, in addition to neurons, the GA of glia in the Syrian hamster undergoes important structural changes, as well as modifications in the intensity of immunostaining and distribution patterns of Golgi structural proteins at different stages of the hibernation cycle.
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http://dx.doi.org/10.3389/fnana.2019.00092DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6882278PMC
November 2019

Classification of GABAergic interneurons by leading neuroscientists.

Sci Data 2019 10 22;6(1):221. Epub 2019 Oct 22.

Laboratorio Cajal de Circuitos Corticales, Universidad Politécnica de Madrid and Instituto Cajal (CSIC), Pozuelo de Alarcón, 28223, Spain.

There is currently no unique catalog of cortical GABAergic interneuron types. In 2013, we asked 48 leading neuroscientists to classify 320 interneurons by inspecting images of their morphology. That study was the first to quantify the degree of agreement among neuroscientists in morphology-based interneuron classification, showing high agreement for the chandelier and Martinotti types, yet low agreement for most of the remaining types considered. Here we present the dataset containing the classification choices by the neuroscientists according to interneuron type as well as to five prominent morphological features. These data can be used as crisp or soft training labels for learning supervised machine learning interneuron classifiers, while further analyses can try to pinpoint anatomical characteristics that make an interneuron especially difficult or especially easy to classify.
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http://dx.doi.org/10.1038/s41597-019-0246-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6805952PMC
October 2019

Slow-Wave Activity in the S1HL Cortex Is Contributed by Different Layer-Specific Field Potential Sources during Development.

J Neurosci 2019 11 23;39(45):8900-8915. Epub 2019 Sep 23.

Departamentos de Neurociencia Traslacional, y

Spontaneous correlated activity in cortical columns is critical for postnatal circuit refinement. We used spatial discrimination techniques to explore the late maturation of synaptic pathways through the laminar distribution of the field potential (FP) generators underlying spontaneous and evoked activities of the S1HL cortex in juvenile (P14-P16) and adult anesthetized rats. Juveniles exhibit an intermittent FP pattern resembling Up/Down states in adults, but with much reduced power and different laminar distribution. Whereas FPs in active periods are dominated by a layer VI generator in juveniles, in adults a developing multipart generator takes over, displaying current sinks in middle layers (III-V). The blockade of excitatory transmission in upper and middle layers of adults recovered the juvenile-like FP profiles. In addition to the layer VI generator, a gamma-specific generator in supragranular layers was the same in both age groups. While searching for dynamical coupling among generators in juveniles we found significant cross-correlation in ∼one-half of the tested pairs, whereas excessive coherence hindered their efficient separation in adults. Also, potentials evoked by tactile and electrical stimuli showed different short-latency dipoles between the two age groups, and the juveniles lacked the characteristic long latency UP state currents in middle layers. In addition, the mean firing rate of neurons was lower in juveniles. Thus, cortical FPs originate from different intra-columnar segments as they become active postnatally. We suggest that although some cortical segments are active early postnatally, a functional sensory-motor control relies on a delayed maturation and network integration of synaptic connections in middle layers. Early postnatal activity in the rodent cortex is mostly endogenous, whereas it becomes driven by peripheral input at later stages. The precise schedule for the maturation of synaptic pathways is largely unknown. We explored this in the somatosensory hindlimb cortex at an age when animals begin to use their limbs by uncovering the laminar distribution of the field potential generators underlying the dominant delta waves in juveniles and adults. Our results suggest that field potentials are mostly generated by a pathway in deep layers, whereas other pathways mature later in middle layers and take over in adults. We suggest that a functional sensory-motor control relies on a delayed maturation and network integration of synaptic connections in middle layers.
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http://dx.doi.org/10.1523/JNEUROSCI.1212-19.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6832678PMC
November 2019

Differential expression of secretagogin immunostaining in the hippocampal formation and the entorhinal and perirhinal cortices of humans, rats, and mice.

J Comp Neurol 2020 03 10;528(4):523-541. Epub 2019 Oct 10.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Madrid, Spain.

Secretagogin (SCGN) is a recently discovered calcium-binding protein belonging to the group of EF-hand calcium-binding proteins. SCGN immunostaining has been described in various regions of the human, rat and mouse brain. In these studies, it has been reported that, in general, the patterns of SCGN staining differ between rodents and human brains. These differences have been interpreted as uncovering phylogenetic differences in SCGN expression. Nevertheless, an important aspect that is not usually taken into account is that different methods are used for obtaining and processing brain tissue coming from humans and experimental animals. This is a critical issue since it has been shown that post-mortem time delay and the method of fixation (i.e., perfused vs. nonperfused brains) may influence the results of the immunostaining. Thus, it is not clear whether differences found in comparative studies with the human brain are simply due to technical factors or species-specific differences. In the present study, we analyzed the pattern of SCGN immunostaining in the adult human hippocampal formation (DG, CA1, CA2, CA3, subiculum, presubiculum, and parasubiculum) as well as in the entorhinal and perirhinal cortices. This pattern of immunostaining was compared with rat and mouse that were fixed either by perfusion or immersion and with different post-mortem time delays (up to 5 hr) to mimic the way the human brain tissue is usually processed. We found a number of clear similarities and differences in the pattern of labeling among the human, rat, and mouse in these brain regions as well as between the different brain regions examined within each species. These differences were not due to the fixation.
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http://dx.doi.org/10.1002/cne.24773DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6972606PMC
March 2020

Differential Structure of Hippocampal CA1 Pyramidal Neurons in the Human and Mouse.

Cereb Cortex 2020 03;30(2):730-752

Instituto Cajal, Consejo Superior de Investigaciones Científicas (CSIC), Madrid 28002, Spain.

Pyramidal neurons are the most common cell type and are considered the main output neuron in most mammalian forebrain structures. In terms of function, differences in the structure of the dendrites of these neurons appear to be crucial in determining how neurons integrate information. To further shed light on the structure of the human pyramidal neurons we investigated the geometry of pyramidal cells in the human and mouse CA1 region-one of the most evolutionary conserved archicortical regions, which is critically involved in the formation, consolidation, and retrieval of memory. We aimed to assess to what extent neurons corresponding to a homologous region in different species have parallel morphologies. Over 100 intracellularly injected and 3D-reconstructed cells across both species revealed that dendritic and axonal morphologies of human cells are not only larger but also have structural differences, when compared to mouse. The results show that human CA1 pyramidal cells are not a stretched version of mouse CA1 cells. These results indicate that there are some morphological parameters of the pyramidal cells that are conserved, whereas others are species-specific.
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http://dx.doi.org/10.1093/cercor/bhz122DOI Listing
March 2020

Phospho-Tau Changes in the Human CA1 During Alzheimer's Disease Progression.

J Alzheimers Dis 2019 ;69(1):277-288

Laboratorio Cajal de Circuitos Corticales (CTB), Universidad Politécnica de Madrid, Madrid, Spain.

Despite extensive studies regarding tau phosphorylation progression in both human Alzheimer's disease cases and animal models, the molecular and structural changes responsible for neurofibrillary tangle development are still not well understood. Here, by using the antibodies AT100 (recognizes tau protein phosphorylated at Thr212 and Ser214 in the proline-rich region) and pS396 (recognizes tau protein phosphorylated at serine residue 396 in the C-terminal region), we examined phospho-tau immunostaining in neurons from the hippocampal CA1 region of 21 human cases with tau pathology ranging from Braak stage I to VI. Our results indicate that the AT100/pS396 ratio decreases in CA1 in accordance with the severity of the disease, along with its colocalization. We therefore propose the AT100/pS396 ratio as a new tool to analyze the tau pathology progression. Our findings also suggest a conformational modification in tau protein that may cause the disappearance of the AT100 epitope in the late stages of tau pathology, which may play a role in the toxic tangle aggregation. Thus, this study provides new insights underlying the stages for the formation of neurofibrillary tangles in Alzheimer's disease.
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http://dx.doi.org/10.3233/JAD-181263DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6598029PMC
September 2020

InTool Explorer: An Interactive Exploratory Analysis Tool for Versatile Visualizations of Neuroscientific Data.

Front Neuroanat 2019 11;13:28. Epub 2019 Mar 11.

Laboratorio Cajal de Circuitos Corticales (CTB), Universidad Politécnica de Madrid (UPM), Madrid, Spain.

The bottleneck for progress in many research areas within neuroscience has shifted from the data acquisition to the data analysis stages. In the present article, we propose a method named InTool Explorer that we have developed to perform interactive exploratory data analysis, focusing on neuroanatomy as an example of its utility. This tool is freely-available software that has been designed to facilitate the study of complex neuroscience data. InTool Explorer requires no more than an internet connection and a web browser. The main contribution of this tool is to provide a user-designed canvas for data visualization and interaction, to perform specific exploratory tasks according to the user needs. Moreover, InTool Explorer permits visualization of the datasets in a very dynamic and versatile way using a linked-card approach. For this purpose, the tool allows the user to select among different predefined card types. Each card type offers an abstract data representation, a filtering tool or a set of statistical analysis methods. Additionally, InTool Explorer makes it possible linking raw images to the data. These images can be used by InTool Explorer to define new customized filtering cards. Another significant contribution of this tool is that it allows fast visualization of the data, error finding, and re-evaluation to establish new hypotheses or new lines of research. Thus, regarding its practical application in the laboratory, InTool Explorer provides a new opportunity to study and analyze neuroscience data prior to any statistical analysis being carried out.
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http://dx.doi.org/10.3389/fnana.2019.00028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6421977PMC
March 2019

Patterns of Dendritic Basal Field Orientation of Pyramidal Neurons in the Rat Somatosensory Cortex.

eNeuro 2018 Nov-Dec;5(6). Epub 2019 Jan 17.

Laboratorio Cajal de Circuitos Corticales, Centro de Tecnología Biomédica, Universidad Politécnica de Madrid, Pozuelo de Alarcón 28223, Spain.

The study of neuronal dendritic orientation is of interest because it is related to how neurons grow dendrites to establish the synaptic input that neurons receive. The dendritic orientations of neurons in the nervous system vary, ranging from rather heterogeneously distributed (asymmetric) to homogeneously distributed (symmetric) dendritic arbors. Here, we analyze the dendritic orientation of the basal dendrites of intracellularly labeled pyramidal neurons from horizontal sections of Layers II-VI of the hindlimb somatosensory (S1HL) cortex of 14-d-old (P14) rats. We used circular statistics and proposed two new graphical descriptive representations of the neuron. We found that the dendritic pattern of most neurons was asymmetric. Furthermore, we found that there is a mixture of different types of orientations within any given group of neurons in any cortical layer. In addition, we investigated whether dendritic orientation was related to the physical location within the brain with respect to the anterior, dorsal, posterior and ventral directions. Generally, there was a preference towards the anterior orientation. A comparison between layers revealed that the preference for the anterior orientation was more pronounced in neurons located in Layers II, III, IV, and Va than for the neurons located in Layers Vb and VI. The dorsal orientation was the least preferred orientation in all layers, except for Layers IV and Va, where the ventral orientation had the lowest preference. Therefore, the orientation of basal dendritic arbors of pyramidal cells is variable and asymmetric, although a majority has a single orientation with a preference for the anterior direction in P14 rats.
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http://dx.doi.org/10.1523/ENEURO.0142-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6335082PMC
March 2019

Metabolomic Study of Hibernating Syrian Hamster Brains: In Search of Neuroprotective Agents.

J Proteome Res 2019 03 22;18(3):1175-1190. Epub 2019 Jan 22.

Laboratorio Cajal de Circuitos Corticales (CTB) , Universidad Politécnica de Madrid , Campus Montegancedo , 28223 Pozuelo de Alarcón , Madrid , Spain.

Syrian hamsters undergo a reversible hyperphosphorylation of protein τ during hibernation, providing a unique natural model that may unveil the physiological mechanisms behind this critical process involved in the development of Alzheimer's disease and other tauopathies. The hibernation cycle of these animals fluctuates between a pair of stages: 3-4 days of torpor bouts interspersed with periods of euthermia called arousals that last several hours. In this study, we investigated for the first time the metabolic changes in brain tissue during hibernation. A total of 337 metabolites showed statistically significant differences during hibernation. Based on these metabolites, several pathways were found to be significantly regulated and, therefore, play a key role in the regulation of hibernation processes. The increase in the levels of ceramides containing more than 20 C atoms was found in torpor animals, reflecting a higher activity of CerS2 during hibernation, linked to neurofibrillary tangle generation and structural changes in the Golgi apparatus. Our results open up the debate about the possible significance of some metabolites during hibernation, which may possibly be related to τ phosphorylation and dephosphorylation events. In general, this study may provide insights into novel neuroprotective agents because the alterations described throughout the hibernation process are reversible.
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http://dx.doi.org/10.1021/acs.jproteome.8b00816DOI Listing
March 2019

A Method for the Symbolic Representation of Neurons.

Front Neuroanat 2018 18;12:106. Epub 2018 Dec 18.

Department of Computer Engineering, Universidad Rey Juan Carlos, Madrid, Spain.

The field of neuroanatomy has progressed considerably in recent decades, thanks to the emergence of novel methods which provide new insights into the organization of the nervous system. These new methods have produced a wealth of data that needs to be analyzed, shifting the bottleneck from the acquisition to the analysis of data. In other disciplines, such as in many engineering areas, scientists and engineers are dealing with increasingly complex systems, using hierarchical decompositions, graphical models and simplified schematic diagrams for analysis and design processes. This approach makes it possible for users to simultaneously combine global system views and very detailed representations of specific areas of interest, by selecting appropriate representations for each of these views. In this way, users can concentrate on specific details while also maintaining a general system overview - a capability that is essential for understanding structure and function whenever complexity is an issue. Following this approach, this paper focuses on a graphical tool designed to help neuroanatomists to better understand and detect morphological characteristics of neuronal cells. The method presented here, based on a symbolic representation that can be tailored to enhance a particular range of features of a neuron or neuron set, has proven to be useful for highlighting particular geometries that may be hidden due to the complexity of the analysis tasks and the richness of neuronal morphologies. A software tool has been developed to generate graphical representations of neurons from 3D computer-aided reconstruction files.
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http://dx.doi.org/10.3389/fnana.2018.00106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6305400PMC
December 2018