Morphology of neurons in the white matter of the adult human neocortex

Summary Neurons in the human cerebral cortical white matter below motor, visual, auditory and prefrontal orbital areas have been studied with the Golgi method, immunohistochemistry and diaphorase histochemistry. The majority of white matter neurons are pyramidal cells displaying the typical polarized, spiny dendritic system. The morphological variety includes stellate forms as well as bipolar pyramidal cells, and the expression of a certain morphological phenotype seems to depend on the position of the neuron. Spineless nonpyramidal neurons with multipolar to bitufted dendritic fields constitute less than 10% of the nuerons stained for microtubule associated protein (MAP-2). Only 3% of the MAP-2 immunoreactive neurons display nicotine adenine dinucleotide-diaphorase activity. The white matter pyramidal neurons are arranged in radial rows continuous with the columns of layer VI neurons. Neuron density is highest below layer VI, and decreases with increasing distance from the gray matter. White matter neurons are especially abundant below the primary motor cortex, and are least frequent below the visual cortex area 17. In contrast to other mammalian species, the white matter neurons in man are not only present during development, but persist throughout life.     

No change in total length of white matter fibers in Alzheimer's disease

White matter changes have been reported as part of Alzheimer dementia. To investigate this, the total subcortical myelinated nerve fiber length was estimated in postmortem brains from eight females (age 79-88 years) with severe Alzheimer's disease (AD) and compared with brains from 10 female control subjects (age 74-92 years). A stereological method for estimating myelinated brain fibers includes sampling systematically, randomly from the white matter, and counting fibers in unbiased counting frames using light microscopy at approximately 6000x magnification. The diameter of each counted fiber was measured to obtain the diameter distribution of myelinated fibers in both groups. The mean total myelinated fiber length was 81,554 km in the AD group and 78,896 km in the control group (P=0.63). All other measured parameters were also unaffected in the AD brains: The mean fiber length density was 248 km/cm3 in the AD group and 247 km/cm3 in the control group; the volume of white matter was 329 cm3 (AD) and 321 cm3 (control) and the volume density of myelinated fibers to white matter tissue volume was 0.30 in AD group and 0.31 in the control group. This is the first study of subcortical brain white matter fiber length using a stereological method on postmortem brains from AD patients and control subjects.     

Populations of subplate and interstitial neurons in fetal and adult human telencephalon

In the adult human telencephalon, subcortical (gyral) white matter contains a special population of interstitial neurons considered to be surviving descendants of fetal subplate neurons [Kostovic & Rakic (1980) Cytology and the time of origin of interstitial neurons in the white matter in infant and adult human and monkey telencephalon. J Neurocytol9, 219]. We designate this population of cells as superficial (gyral) interstitial neurons and describe their morphology and distribution in the postnatal and adult human cerebrum. Human fetal subplate neurons cannot be regarded as interstitial, because the subplate zone is an essential part of the fetal cortex, the major site of synaptogenesis and the 'waiting' compartment for growing cortical afferents, and contains both projection neurons and interneurons with distinct input-output connectivity. However, although the subplate zone is a transient fetal structure, many subplate neurons survive postnatally as superficial (gyral) interstitial neurons. The fetal white matter is represented by the intermediate zone and well-defined deep periventricular tracts of growing axons, such as the corpus callosum, anterior commissure, internal and external capsule, and the fountainhead of the corona radiata. These tracts gradually occupy the territory of transient fetal subventricular and ventricular zones.The human fetal white matter also contains distinct populations of deep fetal interstitial neurons, which, by virtue of their location, morphology, molecular phenotypes and advanced level of dendritic maturation, remain distinct from subplate neurons and neurons in adjacent structures (e.g. basal ganglia, basal forebrain). We describe the morphological, histochemical (nicotinamide-adenine dinucleotide phosphate-diaphorase) and immunocytochemical (neuron-specific nuclear protein, microtubule-associated protein-2, calbindin, calretinin, neuropeptide Y) features of both deep fetal interstitial neurons and deep (periventricular) interstitial neurons in the postnatal and adult deep cerebral white matter (i.e. corpus callosum, anterior commissure, internal and external capsule and the corona radiata/centrum semiovale). Although these deep interstitial neurons are poorly developed or absent in the brains of rodents, they represent a prominent feature of the significantly enlarged white matter of human and non-human primate brains.     

Early history of subplate and interstitial neurons: from Theodor Meynert (1867) to the discovery of the subplate zone (1974)

In this historical review, we trace the early history of research on the fetal subplate zone, subplate neurons and interstitial neurons in the white matter of the adult nervous system. We arrive at several general conclusions. First, a century of research clearly testifies that interstitial neurons, subplate neurons and the subplate zone were first observed and variously described in the human brain - or, in more general terms, in large brains of gyrencephalic mammals, characterized by an abundant white matter and slow and protracted prenatal and postnatal development. Secondly, the subplate zone cannot be meaningfully defined using a single criterion - be it a specific population of cells, fibres or a specific molecular or genetic marker. The subplate zone is a highly dynamic architectonic compartment and its size and cellular composition do not remain constant during development. Thirdly, it is important to make a clear distinction between the subplate zone and the subplate (and interstitial) neurons. The transient existence of the subplate zone (as a specific architectonic compartment of the fetal telencephalic wall) should not be equated with the putative transient existence of subplate neurons. It is clear that in rodents, and to an even greater extent in humans and monkeys, a significant number of subplate cells survive and remain functional throughout life.     

Total number of cells in the human newborn telencephalic wall

The total cell numbers were estimated in the neocortical part of the human telencephalon in 10 normal brains of newborn babies within four major developmental zones: the cortical plate/marginal zone, the subplate, the intermediate zone and the ventricular/subventricular zone. Furthermore, the total number of neuron and glial cells was estimated in the cortical plate. The gestational ages ranged from 38 + 0-42 + 5 weeks + days of gestation. The mean total cell number was 32.6 x 10(9) (coefficient of error = 0.04) and the total number of neurons in the cortical plate 19.8 x 10(9) (coefficient of error = 0.06). This indicates that the total number of neocortical neurons equals the total number in the adults, which, however, is not the case for the glial cells.     

抑郁症模型大鼠大脑白质毛细血管的体视学

目的:探讨抑郁症大鼠大脑白质及白质内毛细血管的改变。方法:选取雄性SD大鼠,经筛选后随机分为对照组和模型组各10只。模型组给予5周的慢性不可预知性刺激,对照组不给予处理。造模成功后,分别从模型组和对照组中各随机选择5只SD大鼠,运用免疫组织化学和体视学方法对大脑白质及其内毛细血管进行定量研究。结果:与对照组相比,模型组的大脑白质内毛细血管体积密度、总体积、长度密度和总长度分别下降了19.6%、22.1%、27.0%和29.9%,差异具有统计学意义。白质体积、毛细血管总表面积密度和总表面积分别下降了5.1%、16.6%和11.3%,差异无统计学意义。结论:抑郁症模型大鼠大脑白质毛细血管存在改变,这一结果为抑郁症血管因素的机制研究提供了重要的依据,也为临床抗抑郁药物的开发提供了新的研究方向。     

Immunocytochemical detection of brain neurons using the selective marker NeuN

The aim of the present work was to develop optimal protocols for immunocytochemical reactions for nuclear protein NeuN for light and laser confocal microscopy which avoid the thermal antigen demasking procedure, which degrades the state of the tissue and requires use of expensive adhesive-coated slices. Maximal antigen retention was obtained after fixation in zinc-formalin and Bouin’s fluid (maximum one day). Two protocols are proposed allowing the thermal demasking procedure to be avoided during detection of neuron marker NeuN on paraffin sections examined by light and confocal microscopy. __________ Translated from Morfologiya, Vol. 128, No. 5, pp. 76–78, September–October, 2005. Corresponding Member of the Russian Academy of Medical Sciences     

The in vivo influence of white matter fiber orientation towards B0 on T2* in the human brain

It is well known that, at higher field strength, T2*‐weighted images show an extensive heterogeneity in white matter fiber bundles. The basis of this phenomenon is still under discussion, as many factors, such as iron concentration, myelination and tissue microstructure, could influence relaxation times. Furthermore, fiber direction in relation to the main magnetic field seems to influence T2* relaxation times. In this study, diffusion tensor imaging and T2* measurements were combined in seven subjects with the head in a normal and tilted position. It was shown that fiber orientation has a strong influence on T2* in the human brain, with the average T2* value changing from 49 ms for a perpendicular orientation to B₀ to 57 ms for a parallel orientation to B₀. Nevertheless, T2* times showed a wide variety of values at any orientation towards B₀. Copyright © 2010 John Wiley & Sons, Ltd.     

大脑白质有髓神经纤维髓鞘超微结构定量研究的体视学方法

目的:运用新的体视学方法研究大鼠白质有髓神经纤维髓鞘超微结构,以期为以后各种有关髓鞘超微结构的相关研究提供可靠的方法学依据.方法:6～8月龄雌性Long-Evans大鼠.运用电镜技术及相应的体视学方法计算白质内有髓神经纤维的总长度、有髓神经纤维髓鞘总体积、有髓神经纤维及轴突直径、髓鞘断面面积、髓鞘内外周长、髓鞘平均厚度.结果:获得了正常6～8月龄雌性Long-Evans大鼠有髓神经纤维总长度及髓鞘超微结构的各项数据,两种不同方法得到的髓鞘平均厚度并没有显著性差异.结论:把新的体视学方法和电子显微镜结合起来定量研究了大鼠大脑白质有髓神经纤维髓鞘超微结构,所描述的方法为以后进行髓鞘超微结构的定量研究提供了有用的工具.     

Grey and white matter GABA level differences in the human brain using two-dimensional, J-resolved spectroscopic imaging

A novel, two-dimensional, J-resolved chemical-shift imaging sequence was used to collect gamma-aminobutyric acid (GABA) spectroscopic imaging data on six healthy subjects at 4 T. Using image segmentation and a linear-regression analysis relating brain GABA level to tissue-type, a consistent and significant (n = 6, p < 0.01) elevation of mean GABA levels was measured in the cortical grey matter (0.96 +/- 0.24 mm) compared with white matter (0.44 +/- 0.16 mm) across all six subjects. The results suggest an approximately two-fold elevation of GABA levels in cortical grey matter compared with white matter in vivo. Our findings are consistent with ex vivo studies in the literature of both animal and human brain and demonstrate the significant potential of this technique for detecting and quantifying tissue-specific neurochemical pathology in vivo.     

Peculiar form of cerebral microdysgenesis characterized by white matter neurons with perineuronal and perivascular glial satellitosis: A study using a variety of human autopsied brains

Microdysgenesis (MD) is a neuropathological term that implies a variety of minor developmental abnormalities of the brain. Recently, MD has been used for pathological diagnosis of cerebral tissues surgically resected from epileptic patients. However, criteria or consensus on pathological diagnosis of MD is still vague and controversial because of the lack of control studies. Therefore, this study paid special attention to the presence of white matter neurons with perineuronal glial satellitosis (WMN-GS) and perivascular glial satellitosis (PVGS) in the white matter, which are occasionally observed in epileptic foci, in order to clarify whether they could be handled as definite findings of MD. The materials included 80 autopsied whole brains ranging from normal subjects to patients with cerebrovascular disorder, neurodegenerative diseases and malformations. In each case, the presence of WMN-GS and/or PVGS was searched in 10 gyri in all five lobes (rostral frontal lobe, caudal frontal lobe, parietal lobe, temporal lobe and oc-cipital lobe) and evaluated. Statistically significant, WMN-GS and/or PVGS preferentially appeared in a diseased group consisting of neuronal migration disorder and related conditions, such as polymicrogyria, nodular heterotopia or tuberous sclerosis, leading to a suggestive conclusion that the presence of WMN-GS and/or PVGS could be a peculiar form of MD possibly derived from neuronal migrational arrest or related events, even if they appear alone without any other gross abnormalities.     

大鼠大脑白质及白质内有髓神经纤维的性别与年龄差异

目的:研究正常年轻和中老年组雌雄性大鼠大脑白质及白质内有髓神经纤维之间的性别差异,并探讨大脑发育过程中性别差异随年龄增加的改变。方法:运用透射电子显微镜和体视学方法对6~8月龄的年轻Long-Evans大鼠及18月龄同种中老年大鼠大脑白质及其有髓神经纤维进行定量研究。结果:年轻组雄性大鼠大脑白质、有髓神经纤维及其髓鞘的总体积均显著大于雌性,而中老年组雌性大鼠大脑白质、有髓神经纤维体积密度、有髓神经纤维及其髓鞘的总体积均显著大于雄性。结论:年轻组及中老年组大鼠大脑白质、白质内有髓神经纤维及其髓鞘总体积均存在性别差异,随着年龄的增长,雄性大鼠大脑白质及白质内有髓神经纤维体积的减少较雌性更为明显。     

应用DTI技术对人脑白质纤维束的初步研究

目的建立扩散张量纤维束成像对人脑白质纤维的显示方法,并应用中国数字化可视人体数据进行对照观察,验证扩散张量成像(DTI)方法的可靠性。方法选择5名健康志愿者进行DTI成像,采用DtiStudio软件进行分析处理,重建出部分各向异性(FA)图、容积比(VR)图、相对各向异性(RA)图、表面扩散系数(ADC)图以及二维彩色张量图。应用中国数字化可视人体数据集断面图像、FA图及彩色FA图进行对照观察,利用fibertracking纤维跟踪软件及3DMRI软件进行三维重建显示脑内主要白质纤维束,辨认脑内白质纤维束的位置、形态。结果应用DTI纤维束成像可以清晰准确地描绘脑白质内主要神经纤维束的解剖图谱,包括联络纤维如弓形纤维、钩束、扣带束、上纵束和下纵束,连合纤维如胼胝体、前连合和穹隆,投射纤维如锥体束、视放射、内侧丘系等。DTI纤维束成像结果与已知解剖知识、中国可视化人体断面图像具有很好的一致性。结论应用DTI纤维束成像可以清晰准确地描绘脑白质内主要神经纤维束的解剖图谱,其结果与中国可视化人体断面图像、已知解剖知识是一致的,应用DTI纤维束成像研究脑内纤维连通性是可靠的。     

Sublaminar organization of the human subplate: developmental changes in the distribution of neurons,glia, growing axons and extracellular matrix

The objective of this paper was to collect normative data essential for analyzing the subplate (SP) role in pathogenesis of developmental disorders, characterized by abnormal circuitry, such as hypoxic‐ischemic lesions, autism and schizophrenia. The main cytological features of the SP, such as low cell density, early differentiation of neurons and glia, plexiform arrangement of axons and dendrites, presence of synapses and a large amount of extracellular matrix (ECM) distinguish this compartment from the cell‐dense cortical plate (CP; towards pia) and large fiber bundles of external axonal strata of fetal white matter (towards ventricle). For SP delineation from these adjacent layers based on combined cytological criteria, we analyzed the sublaminar distribution of different microstructural elements and the associated maturational gradients throughout development, using immunocytochemical and histological techniques on postmortem brain material (Zagreb Neuroembryological Collection). The analysis revealed that the SP compartment of the lateral neocortex shows changes in laminar organization throughout fetal development: the monolayer in the early fetal period (presubplate) undergoes dramatic bilaminar transformation between 13 and 15 postconceptional weeks (PCW), followed by subtle sublamination in three ‘floors’ (deep, intermediate, superficial) of midgestation (15–21 PCW). During the stationary phase (22–28 PCW), SP persists as a trilaminar compartment, gradually losing its sublaminar organization towards the end of gestation and remains as a single layer of SP remnant in the newborn brain. Based on these sublaminar transformations, we have documented developmental changes in the distribution, maturational gradients and expression of molecular markers in SP synapses, transitional forms of astroglia, neurons and ECM, which occur concomitantly with the ingrowth of thalamo‐cortical, basal forebrain and cortico‐cortical axons in a deep to superficial fashion. The deep SP is the zone of ingrowing axons – ‘entrance (ingrowth) zone’. The process of axonal ingrowth begins with thalamo‐cortical fibers and basal forebrain afferents, indicating an oblique geometry. During the later fetal period, deep SP receives long cortico‐cortical axons exhibiting a tangential geometry. Intermediate SP (‘proper’) is the navigation and ‘nexus’ sublamina consisting of a plexiform arrangement of cellular elements providing guidance and substrate for axonal growth, and also containing transient connectivity of dendrites and axons in a tangential plane without radial boundaries immersed in an ECM‐rich continuum. Superficial SP is the axonal accumulation (‘waiting compartment’) and target selection zone, indicating a dense distribution of synaptic markers, accumulation of thalamo‐cortical axons (around 20 PCW), overlapping with dendrites from layer VI neurons. In the late preterm brain period, superficial SP contains a chondroitin sulfate non‐immunoreactive band. The developmental dynamics for the distribution of neuronal, glial and ECM markers comply with sequential ingrowth of afferents in three levels of SP: ECM and synaptic markers shift from deep to superficial SP, with transient forms of glia following this arrangement, and calretinin neurons are concentrated in the SP during the formation phase. These results indicate developmental and morphogenetic roles in the SP cellular (transient glia, neurons and synapses) and ECM framework, enabling the spatial accommodation, navigation and establishment of numerous connections of cortical pathways in the expanded human brain. The original findings of early developmental dynamics of transitional subtypes of astroglia, calretinin neurons, ECM and synaptic markers presented in the SP are interesting in the light of recent concepts concerning its functional and morphogenetic role and an increasing interest in SP as a prospective substrate of abnormalities in cortical circuitry, leading to a cognitive deficit in different neurodevelopmental disorders.     

Periventricular white matter damage in the hypoxic neonatal brain: Role of microglial cells

Periventricular white matter damage (PWMD) also known as periventricular white matter injury, is one of the major causes of neurological impairment in premature newborns. The etiology of white matter injury is multifaceted with hypoxia-ischemia being an important underlying factor. The developing oligodendrocytes are susceptible to damage resulting in myelination deficits. Excess release of glutamate, free radical production, release of cytokines and iron accumulation are factors thought to mediate damage to the developing white matter. Recent studies have also suggested a role for vascular endothelial growth factor and nitric oxide in the pathogenesis of PWMD. Although the role of microglial cells in the development of PWMD is still debatable, our recent investigations have shed some light on their involvement in the pathogenesis of PWMD. Challenges for the future include in-depth investigation of crosstalk between microglia and immature oligodendrocytes as well as other glial cells and vascular endothelial cells.     

Stereological estimation of the total number of myelinated callosal fibers in human subjects

Using the fractionator principle, the total number, density and diameter size of myelinated callosal fibers were estimated in the corpus callosum (CC) of 10 Danish males between 39 and 60years of age. All sampled brains had been used in previous quantitative studies, for example, studies of neocortical neuron number, and were selected to determine whether the variability in the neocortical neuron number correlated with the total number of myelinated callosal fibers. Middle-aged males had an average of 138×10(6) (coefficient of variance; CV=0.19) myelinated fibers, but did not show any correlation with the neocortical neuron number (r=0.25; P=0.49). The mean area of the CC was estimated to be 7.2cm(2) (CV=0.17), and showed a significant correlation with the number of callosal fibers (r=0.69, P=0.041). Additionally, an overall density decline from the anterior to the posterior region of the CC was observed, with an inverse relationship between the distribution of large and small fibers along the callosal axis. This study suggests that many mechanisms are involved in the development and determination of axonal projections across the CC that cannot simply be explained by the neocortical neuron number. Further, a positive correlation between callosal fibers and the CC area verifies that callosal fibers are the factor responsible for CC size. Finally, the number of callosal fibers and their diameters are distributed along the CC in a specific pattern that reflects interactions with different brain regions.     

Brain white matter abnormalities associated with copy number variants

White matter (WM) signal abnormalities are demonstrated in various neurodevelopmental disorders on brain magnetic resonance imaging (MRI). The pattern of WM abnormalities can aid in the diagnostic process. This study aims to characterize the WM changes found in microdeletion/microduplication syndromes. Thirteen patients with neurodevelopmental disorders due to copy number variations were collected from a cohort of children with evidence of WM abnormalities on brain MRI, in two medical centers. A pediatric neuroradiologist blindly interpreted the MRI scans. Clinical and genetic findings were retrospectively extracted from the medical records. WM changes included: multifocal (10/13) periventricular (12/13) and subcortical (5/13) signal abnormalities and WM volume loss (6/13). Dysgenesis of the corpus callosum was depicted in 12/13. The main clinical features were: global developmental delay (13/13), hypotonia (11/13), epilepsy (10/13), dysmorphic features (9/13), microcephaly (6/13), short stature (6/13), and systemic involvement (6/13). We showed that different chromosomal micro‐rearrangement syndromes share similar MRI patterns of nonspecific multifocal predominantly periventricular WM changes associated with corpus callosum dysgenesis with or without WM and gray matter loss. Hence, the association of these features in a patient evaluated for global developmental delay/intellectual disability suggests a chromosomal micro‐rearrangement syndrome, and a chromosomal microarray analysis should be performed.     

Spatiotemporal changes in diffusion,T2 and susceptibility of white matter following mild traumatic brain injury

Impaired white matter integrity in traumatic brain injury (TBI) can lead to deficits in various neurological functions. The differentiation of the underlying pathological processes, e.g. edema, demyelination, axonal damage, to name a few, is of key clinical interest for the assessment of white matter injury. In this study, a combination of T₂, diffusion and susceptibility MRI was used to study the spatiotemporal changes in white matter at 1 h, 3 h, and 1, 2, 7 and 14 days following TBI, using a rat controlled cortical impact (CCI) model. Based on radial diffusivity (RD), the rats were divided into two groups: group 1 showed widespread increases in RD along the corpus callosum of the ipsilesional hemisphere at day 2, and group 2 showed normal RD. Based on this group separation, group 1 also showed similar widespread changes in fractional anisotropy (FA) and T₂ at day 2, and group 2 showed normal FA and T₂. The widespread changes in RD and T₂ in group 1 on day 2 were apparently dominated by edema, which obscured possible myelin and axonal damage. In contrast, the susceptibility of group 1 showed more localized increases near the impact site on day 2, and otherwise similar contrast to the contralesional hemisphere. The localized susceptibility increase is probably a result of demyelination and axonal injury. The extent of brain damage between the two groups revealed by MRI was consistent with behavioral results, with the first group showing significantly increased forelimb asymmetry and increased forelimb foot fault deficits. Our results suggest that the combination of T₂, diffusion and susceptibility MRI may provide an opportunity for the differential assessment of edema and axonal damage in TBI. Copyright © 2016 John Wiley & Sons, Ltd.     

Enhanced delineation of white matter structures of the fixed mouse brain using Gd‐DTPA in microscopic MRI

The purpose of this study was to investigate the effect of gadolinium (III) diethyltriaminepenta‐acetic acid (Gd‐DTPA) mixed with a fixative on the image contrast between the white and gray matter of the perfusion‐fixed mouse brain. A series of microscopic MRI (µMRI) studies using different concentrations of Gd‐DTPA were performed at multiple time points to determine the optimal Gd‐DTPA concentration and fixation time necessary to maximize the contrast‐to‐noise ratio between the white and gray matter with relatively short scan time using a three‐dimensional gradient‐echo pulse sequence. On the basis of the experimental results, high‐resolution (39 µm isotropic) images with excellent contrast‐to‐noise ratio (～50) were acquired in less than 2 h of scan time after the specimen had been soaked in 10 mM Gd‐DTPA for 4 days. Excellent correlation was noted between µMRI and histology in that the µMRI clearly depicted brain regions that were also observed by the Kluver–Barrera stain. The enhanced contrast between the white and gray matter obtained by the proposed µMRI method may facilitate the development of µMRI‐based morphological phenotyping methods for mouse models of neurological disorders. Copyright © 2008 John Wiley & Sons, Ltd.