Sublamina-Specific Dynamics and Ultrastructural Heterogeneity of Layer 6 Excitatory Synaptic Boutons in the Adult Human Temporal Lobe Neocortex.

Synapses "govern" the computational properties of any given network in the brain. However, their detailed quantitative morphology is still rather unknown, particularly in humans. Quantitative 3D-models of synaptic boutons (SBs) in layer (L)6a and L6b of the temporal lobe neocortex (TLN) were generated from biopsy samples after epilepsy surgery using fine-scale transmission electron microscopy, 3D-volume reconstructions and electron microscopic tomography. Beside the overall geometry of SBs, the size of active zones (AZs) and that of the three pools of synaptic vesicles (SVs) were quantified. SBs in L6 of the TLN were middle-sized (~5 µm2), the majority contained only a single but comparatively large AZ (~0.20 µm2). SBs had a total pool of ~1100 SVs with comparatively large readily releasable (RRP, ~10 SVs L6a), (RRP, ~15 SVs L6b), recycling (RP, ~150 SVs), and resting (~900 SVs) pools. All pools showed a remarkably large variability suggesting a strong modulation of short-term synaptic plasticity. In conclusion, L6 SBs are highly reliable in synaptic transmission within the L6 network in the TLN and may act as "amplifiers," "integrators" but also as "discriminators" for columnar specific, long-range extracortical and cortico-thalamic signals from the sensory periphery.

Sex dependent glial-specific changes in the chromatin accessibility landscape in late-onset Alzheimer's disease brains.

BACKGROUND: In the post-GWAS era, there is an unmet need to decode the underpinning genetic etiologies of late-onset Alzheimer's disease (LOAD) and translate the associations to causation. METHODS: We conducted ATAC-seq profiling using NeuN sorted-nuclei from 40 frozen brain tissues to determine LOAD-specific changes in chromatin accessibility landscape in a cell-type specific manner. RESULTS: We identified 211 LOAD-specific differential chromatin accessibility sites in neuronal-nuclei, four of which overlapped with LOAD-GWAS regions (±100 kb of SNP). While the non-neuronal nuclei did not show LOAD-specific differences, stratification by sex identified 842 LOAD-specific chromatin accessibility sites in females. Seven of these sex-dependent sites in the non-neuronal samples overlapped LOAD-GWAS regions including APOE. LOAD loci were functionally validated using single-nuclei RNA-seq datasets. CONCLUSIONS: Using brain sorted-nuclei enabled the identification of sex-dependent cell type-specific LOAD alterations in chromatin structure. These findings enhance the interpretation of LOAD-GWAS discoveries, provide potential pathomechanisms, and suggest novel LOAD-loci.

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

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.

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

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.

MRI-based brain structural changes following radiotherapy of Nasopharyngeal Carcinoma: A systematic review.

PURPOSE: Nasopharyngeal carcinoma (NPC) radiotherapy (RT) irradiates parts of the brain which may cause cerebral tissue changes. This study aimed to systematically review the brain microstructure changes using MRI-based measures, diffusion tensor imaging (DTI), diffusion kurtosis imaging (DKI) and voxel-based morphometry (VBM) and the impact of dose and latency following RT. METHODS: PubMed and Scopus databases were searched based on PRISMA guideline to determine studies focusing on changes following NPC RT. RESULTS: Eleven studies fulfilled the inclusion criteria. Microstructural changes occur most consistently in the temporal region. The changes were correlated with latency in seven studies; fractional anisotropy (FA) and gray matter (GM) volume remained low even after a longer period following RT and areas beyond irradiation site with reduced FA and GM measures. For dosage, only one study showed correlation, thus requiring further investigations. CONCLUSION: DTI, DKI and VBM may be used as a surveillance tool in detecting brain microstructural changes of NPC patients which correlates to latency and brain areas following RT.

Synaptic Organization of the Human Temporal Lobe Neocortex as Revealed by High-Resolution Transmission, Focused Ion Beam Scanning, and Electron Microscopic Tomography.

Modern electron microscopy (EM) such as fine-scale transmission EM, focused ion beam scanning EM, and EM tomography have enormously improved our knowledge about the synaptic organization of the normal, developmental, and pathologically altered brain. In contrast to various animal species, comparably little is known about these structures in the human brain. Non-epileptic neocortical access tissue from epilepsy surgery was used to generate quantitative 3D models of synapses. Beside the overall geometry, the number, size, and shape of active zones and of the three functionally defined pools of synaptic vesicles representing morphological correlates for synaptic transmission and plasticity were quantified. EM tomography further allowed new insights in the morphological organization and size of the functionally defined readily releasable pool. Beside similarities, human synaptic boutons, although comparably small (approximately 5 µm), differed substantially in several structural parameters, such as the shape and size of active zones, which were on average 2 to 3-fold larger than in experimental animals. The total pool of synaptic vesicles exceeded that in experimental animals by approximately 2 to 3-fold, in particular the readily releasable and recycling pool by approximately 2 to 5-fold, although these pools seemed to be layer-specifically organized. Taken together, synaptic boutons in the human temporal lobe neocortex represent unique entities perfectly adapted to the "job" they have to fulfill in the circuitry in which they are embedded. Furthermore, the quantitative 3D models of synaptic boutons are useful to explain and even predict the functional properties of synaptic connections in the human neocortex.

Ultrastructural heterogeneity of layer 4 excitatory synaptic boutons in the adult human temporal lobe neocortex.

Synapses are fundamental building blocks controlling and modulating the 'behavior' of brain networks. How their structural composition, most notably their quantitative morphology underlie their computational properties remains rather unclear, particularly in humans. Here, excitatory synaptic boutons (SBs) in layer 4 (L4) of the temporal lobe neocortex (TLN) were quantitatively investigated. Biopsies from epilepsy surgery were used for fine-scale and tomographic electron microscopy (EM) to generate 3D-reconstructions of SBs. Particularly, the size of active zones (AZs) and that of the three functionally defined pools of synaptic vesicles (SVs) were quantified. SBs were comparatively small (~2.50 µm2), with a single AZ (~0.13 µm2); preferentially established on spines. SBs had a total pool of ~1800 SVs with strikingly large readily releasable (~20), recycling (~80) and resting pools (~850). Thus, human L4 SBs may act as 'amplifiers' of signals from the sensory periphery, integrate, synchronize and modulate intra- and extracortical synaptic activity.

Quantitative Three-Dimensional Reconstructions of Excitatory Synaptic Boutons in Layer 5 of the Adult Human Temporal Lobe Neocortex: A Fine-Scale Electron Microscopic Analysis.

Studies of synapses are available for different brain regions of several animal species including non-human primates, but comparatively little is known about their quantitative morphology in humans. Here, synaptic boutons in Layer 5 (L5) of the human temporal lobe (TL) neocortex were investigated in biopsy tissue, using fine-scale electron microscopy, and quantitative three-dimensional reconstructions. The size and organization of the presynaptic active zones (PreAZs), postsynaptic densities (PSDs), and that of the 3 distinct pools of synaptic vesicles (SVs) were particularly analyzed. L5 synaptic boutons were medium-sized (~6 µm2) with a single but relatively large PreAZ (~0.3 µm2). They contained a total of ~1500 SVs/bouton, ~20 constituting the putative readily releasable pool (RRP), ~180 the recycling pool (RP), and the remainder, the resting pool. The PreAZs, PSDs, and vesicle pools are ~3-fold larger than those of CNS synapses in other species. Astrocytic processes reached the synaptic cleft and may regulate the glutamate concentration. Profound differences exist between synapses in human TL neocortex and those described in various species, particularly in the size and geometry of PreAZs and PSDs, the large RRP/RP, and the astrocytic ensheathment suggesting high synaptic efficacy, strength, and modulation of synaptic transmission at human synapses.

Quantitative 3-D morphometric analysis of individual dendritic spines.

The observation and analysis of dendritic spines morphological changes poses a major challenge in neuroscience studies. The alterations of their density and/or morphology are indicators of the cellular processes involved in neural plasticity underlying learning and memory, and are symptomatic in neuropsychiatric disorders. Despite ongoing intense investigations in imaging approaches, the relationship between changes in spine morphology and synaptic function is still unknown. The existing quantitative analyses are difficult to perform and require extensive user intervention. Here, we propose a new method for (1) the three-dimensional (3-D) segmentation of dendritic spines using a multi-scale opening approach and (2) define 3-D morphological attributes of individual spines for the effective assessment of their structural plasticity. The method was validated using confocal light microscopy images of dendritic spines from dissociated hippocampal cultures and brain slices (1) to evaluate accuracy relative to manually labeled ground-truth annotations and relative to the state-of-the-art Imaris tool, (2) to analyze reproducibility of user-independence of the segmentation method, and (3) to quantitatively analyze morphological changes in individual spines before and after chemically induced long-term potentiation. The method was monitored and used to precisely describe the morphology of individual spines in real-time using consecutive images of the same dendritic fragment.

Modified cuckoo search algorithm in microscopic image segmentation of hippocampus.

Microscopic image analysis is one of the challenging tasks due to the presence of weak correlation and different segments of interest that may lead to ambiguity. It is also valuable in foremost meadows of technology and medicine. Identification and counting of cells play a vital role in features extraction to diagnose particular diseases precisely. Different segments should be identified accurately in order to identify and to count cells in a microscope image. Consequently, in the current work, a novel method for cell segmentation and identification has been proposed that incorporated marking cells. Thus, a novel method based on cuckoo search after pre-processing step is employed. The method is developed and evaluated on light microscope images of rats' hippocampus which used as a sample for the brain cells. The proposed method can be applied on the color images directly. The proposed approach incorporates the McCulloch's method for lévy flight production in cuckoo search (CS) algorithm. Several objective functions, namely Otsu's method, Kapur entropy and Tsallis entropy are used for segmentation. In the cuckoo search process, the Otsu's between class variance, Kapur's entropy and Tsallis entropy are employed as the objective functions to be optimized. Experimental results are validated by different metrics, namely the peak signal to noise ratio (PSNR), mean square error, feature similarity index and CPU running time for all the test cases. The experimental results established that the Kapur's entropy segmentation method based on the modified CS required the least computational time compared to Otsu's between-class variance segmentation method and the Tsallis entropy segmentation method. Nevertheless, Tsallis entropy method with optimized multi-threshold levels achieved superior performance compared to the other two segmentation methods in terms of the PSNR.

Simple platform for chronic imaging of hippocampal activity during spontaneous behaviour in an awake mouse.

Chronic electrophysiological recordings of neuronal activity combined with two-photon Ca2+ imaging give access to high resolution and cellular specificity. In addition, awake drug-free experimentation is required for investigating the physiological mechanisms that operate in the brain. Here, we developed a simple head fixation platform, which allows simultaneous chronic imaging and electrophysiological recordings to be obtained from the hippocampus of awake mice. We performed quantitative analyses of spontaneous animal behaviour, the associated network states and the cellular activities in the dorsal hippocampus as well as estimated the brain stability limits to image dendritic processes and individual axonal boutons. Ca2+ imaging recordings revealed a relatively stereotyped hippocampal activity despite a high inter-animal and inter-day variability in the mouse behavior. In addition to quiet state and locomotion behavioural patterns, the platform allowed the reliable detection of walking steps and fine speed variations. The brain motion during locomotion was limited to ~1.8 µm, thus allowing for imaging of small sub-cellular structures to be performed in parallel with recordings of network and behavioural states. This simple device extends the drug-free experimentation in vivo, enabling high-stability optophysiological experiments with single-bouton resolution in the mouse awake brain.

Early developmental bisphenol-A exposure sex-independently impairs spatial memory by remodeling hippocampal dendritic architecture and synaptic transmission in rats.

Bisphenol-A (BPA, 4, 4'-isopropylidene-2-diphenol), a synthetic xenoestrogen that widely used in the production of polycarbonate plastics, has been reported to impair hippocampal development and function. Our previous study has shown that BPA exposure impairs Sprague-Dawley (SD) male hippocampal dendritic spine outgrowth. In this study, the sex-effect of chronic BPA exposure on spatial memory in SD male and female rats and the related synaptic mechanism were further investigated. We found that chronic BPA exposure impaired spatial memory in both SD male and female rats, suggesting a dysfunction of hippocampus without gender-specific effect. Further investigation indicated that BPA exposure causes significant impairment of dendrite and spine structure, manifested as decreased dendritic complexity, dendritic spine density and percentage of mushroom shaped spines in hippocampal CA1 and dentate gyrus (DG) neurons. Furthermore, a significant reduction in Arc expression was detected upon BPA exposure. Strikingly, BPA exposure significantly increased the mIPSC amplitude without altering the mEPSC amplitude or frequency, accompanied by increased GABAARß2/3 on postsynaptic membrane in cultured CA1 neurons. In summary, our study indicated that Arc, together with the increased surface GABAARß2/3, contributed to BPA induced spatial memory deficits, providing a novel molecular basis for BPA achieved brain impairment.

Frequent globular neuronal cytoplasmic inclusions in the medial temporal region as a possible characteristic feature in multiple system atrophy with dementia.

Multiple system atrophy (MSA) is an adult-onset neurodegenerative disease, which is characterized clinically by parkinsonism, cerebellar ataxia and/or autonomic dysfunction, and pathologically by alpha-synuclein-related multisystem neurodegeneration, so-called alpha-synucleinopathy, which particularly involves the striatonigral and olivopontocerebellar systems, with glial cytoplasmic inclusions and neuronal cytoplasmic/nuclear inclusions (NCIs/NNIs). In the recent consensus criteria for the diagnosis of MSA, dementia is described as one of the features not supporting a diagnosis of MSA. However, MSA with dementia has been reported, although the location of the lesion responsible for the dementia remains unclear. In the present study, we aimed to investigate where this lesion may be found, by analyzing 12 autopsy-proven MSA cases, with a particular focus on the medial temporal region. Three of 12 cases with MSA had dementia (MSA-D). Compared with MSA cases without dementia, MSA-D cases had frequent globular NCIs (G-NCIs) in the medial temporal region, especially in their subiculum. In addition, MSA-D cases could be divided into two types; MSA-D with distinct fronto-temporal lobar degeneration (FTLD type) and without distinct fronto-temporal lobar degeneration (non-FTLD type). There was no association between dementia and Alzheimer pathologies, such as neurofibrillary tangles and senile plaques. We suggest that frequent G-NCIs in the medial temporal region, and particularly the subiculum, is one of the important pathological findings of MSA-D, even when a case with MSA-D reveals no significant cerebral atrophy.

Neural substrate of quality of life in patients with schizophrenia: a magnetisation transfer imaging study.

The aim of this study was to investigate the neural substrate underlying quality of life (QoL) and to demonstrate the microstructural abnormalities associated with impaired QoL in a large sample of patients with schizophrenia, using magnetisation transfer imaging. A total of 81 right-handed men with a diagnosis of schizophrenia and 25 age- and sex-similar healthy controls were included and underwent a 3T MRI with magnetization transfer ratio (MTR) to detect microstructural abnormalities. Compared with healthy controls, patients with schizophrenia had grey matter (GM) decreased MTR values in the temporal lobe (BA21, BA37 and BA38), the bilateral insula, the occipital lobe (BA17, BA18 and BA19) and the cerebellum. Patients with impaired QoL had lower GM MTR values relative to patients with preserved QoL in the bilateral temporal pole (BA38), the bilateral insula, the secondary visual cortex (BA18), the vermis and the cerebellum. Significant correlations between MTR values and QoL scores (p < 0.005) were observed in the GM of patients in the right temporal pole (BA38), the bilateral insula, the vermis and the right cerebellum. Our study shows that QoL impairment in patients with schizophrenia is related to the microstructural changes in an extensive network, suggesting that QoL is a bio-psychosocial marker.

Lipofuscin Granules in the Epileptic Human Temporal Neocortex with Age.

Lipofuscin granules (LGs), the "age pigments", are autofluorescent cell products from lysosomes that diverge in number and size among brain regions. Human temporal cortex from 20- to 55-year-old epileptic subjects were studied with the fat soluble dye Sudan Black, under confocal and electron microscopy. Ultrastructural analysis showed that with age LGs increase in area, but not in number. Proportionally to the LGs area, the electron lucid portion increases and the electron dense reduces over time. The robust increase in lipid components is possibly due to modifications in the neuronal metabolism with age in physiological and pathological conditions.

Positive symptoms and water diffusivity of the prefrontal and temporal cortices in schizophrenia patients: a pilot study.

The development of diffusion tensor imaging (DTI) has provided information about microstructural changes in the brain. Most DTI studies have focused on white matter (WM). Few DTI studies have examined the gray matter (GM) in schizophrenia and, to date, there has been no attempt to identify the relationship between water diffusivity and symptom severity in schizophrenia. The present study aimed to examine microstructural deficits in the dorsal prefrontal cortex (DPFC) and temporal cortex in schizophrenia patients using fractional anisotropy (FA) and water diffusivity. This study also explored the relationship between DTI measurements and psychotic symptoms. Magnetic resonance imaging (MRI) and DTI were used to study 19 schizophrenia patients and 19 healthy controls. Fractional anisotropy, axial diffusivity, radial diffusivity, and regional volumes were measured in the prefrontal cortex and temporal cortex. On DTI measurements, patients showed increased axial and radial diffusivities in the prefrontal cortex and temporal cortex, but they did not demonstrate any difference in fractional anisotropy and regional volumes. Additionally, axial and radial diffusivities were significantly correlated with positive symptom scores in all regions of interest. These results indicate that water diffusivity measurements, including axial and radial diffusivities, can be used to identify microstructural changes in the gray matter in schizophrenia that may be related to symptom severity.

A high-resolution study of hippocampal and medial temporal lobe correlates of spatial context and prospective overlapping route memory.

When navigating our world we often first plan or retrieve an ideal route to our goal, avoiding alternative paths that lead to other destinations. The medial temporal lobe (MTL) has been implicated in processing contextual information, sequence memory, and uniquely retrieving routes that overlap or "cross paths." However, the identity of subregions of the hippocampus and neighboring cortex that support these functions in humans remains unclear. The present study used high-resolution functional magnetic resonance imaging (hr-fMRI) in humans to test whether the CA3/DG hippocampal subfield and parahippocampal cortex are important for processing spatial context and route retrieval, and whether the CA1 subfield facilitates prospective planning of mazes that must be distinguished from alternative overlapping routes. During hr-fMRI scanning, participants navigated virtual mazes that were well-learned from prior training while also learning new mazes. Some routes learned during scanning shared hallways with those learned during pre-scan training, requiring participants to select between alternative paths. Critically, each maze began with a distinct spatial contextual Cue period. Our analysis targeted activity from the Cue period, during which participants identified the current navigational episode, facilitating retrieval of upcoming route components and distinguishing mazes that overlap. Results demonstrated that multiple MTL regions were predominantly active for the contextual Cue period of the task, with specific regions of CA3/DG, parahippocampal cortex, and perirhinal cortex being consistently recruited across trials for Cue periods of both novel and familiar mazes. During early trials of the task, both CA3/DG and CA1 were more active for overlapping than non-overlapping Cue periods. Trial-by-trial Cue period responses in CA1 tracked subsequent overlapping maze performance across runs. Together, our findings provide novel insight into the contributions of MTL subfields to processing spatial context and route retrieval, and support a prominent role for CA1 in distinguishing overlapping episodes during navigational "look-ahead" periods.

Crush cytologic findings of a cerebral granular cell astrocytoma.

BACKGROUND: Granular cell astrocytoma (GCA) is a rare variant of astrocytoma, characterized by an aggressive prognosis compared to conventional astrocytomas of the same World Health Organization grade. Intraoperative smears provide useful clues in diagnosing neuropathology, especially in rarely encountered central nervous system tumors. CASE: The patient was a 53-year-old man who presented with a huge mass at the left temporal lobe with peritumoral edema on MRI. The crush smears revealed singly-scattered, large eosinophilic cells with eccentrically located nuclei as well as plump, finely-granular cytoplasm with distinct borders. Mild cellular atypia and absence of mitotic activity were noted. These cells were admixed with small mature lymphoid cells. Histology showed scattered large granular cells which were positive for glial fibrillary acidic protein. CONCLUSION: The most helpful imprint cytologic findings of GCA were as follows: (1) large cells containing eosinophilic granular cytoplasm rather than the foamy or bubbly cytoplasm associated with macrophages or renal cell carcinomas; (2) distinct granular cell borders in contrast to the ruffled membrane of macrophages, and (3) markedly large-sized granular cells, ranging from 60 to 100 µm in diameter.

Age-related modulation of γ-secretase activity in non-human primate brains.

Age-dependent accumulation of the amyloid-ß peptide (Aß) in the brain is a pre-condition for development of Alzheimer's disease. A relative increase in the generation of longer Aß species such as Aß42 and Aß43 is critical for Aß deposition, but the underlying mechanism remains unresolved. Here, we performed a cell-free assay using microsome fractions of temporal cortex tissues from 42 cynomolgus monkeys and found that Aß40-generating γ-secretase activity (γ40) decreased with age, whereas Aß42-generating γ-secretase activity (γ42) was unaltered. In ELISAs, more than 80% of monkeys over 20-years old showed evidence of Aß accumulation in the temporal cortex. The ratio of γ42 to γ40 increased with age and correlated with the level of accumulated Aß. These results suggest that γ-secretase activity undergoes age-related, non-genetic modulation and that this modulation may cause Aß accumulation in aging brains. Similar modulation may predispose aged human brains to Alzheimer's disease.