Relief of hypoxia by angiogenesis promotes neural stem cell differentiation by targeting glycolysis

Blood vessels are part of the stem cell niche in the developing cerebral cortex, but their in vivo role in controlling the expansion and differentiation of neural stem cells (NSCs) in development has not been studied. Here, we report that relief of hypoxia in the developing cerebral cortex by ingrowth of blood vessels temporo‐spatially coincided with NSC differentiation. Selective perturbation of brain angiogenesis in vessel‐specific Gpr124 null embryos, which prevented the relief from hypoxia, increased NSC expansion at the expense of differentiation. Conversely, exposure to increased oxygen levels rescued NSC differentiation in Gpr124 null embryos and increased it further in WT embryos, suggesting that niche blood vessels regulate NSC differentiation at least in part by providing oxygen. Consistent herewith, hypoxia‐inducible factor (HIF)‐1α levels controlled the switch of NSC expansion to differentiation. Finally, we provide evidence that high glycolytic activity of NSCs is required to prevent their precocious differentiation in vivo. Thus, blood vessel function is required for efficient NSC differentiation in the developing cerebral cortex by providing oxygen and possibly regulating NSC metabolism.     

Implantation of embryonic neocortex and spinal cord into injured peripheral nerve of adult rats

Spinal cord and cerebral cortex of 14-day-old embryos of Wistar rats were implanted into the sciatic nerve of mature rats in order to study dynamics of the development of neuronal and neuroglial elements in ectopic sites. By means of light and electron microscopy it has been stated that the implanted nerve cells of the cortex and spinal cord survive during 5 month and differentiate from neuroepithelial cells and neuroblasts up to young and mature neurons. It was found that thirty days after operation the spinal cord implants contained myelinated nerve fibers and numerous synapses. The data obtained suggest that the implants of fetal spinal cord are more favorable for regeneration of the injured nervous stems than the cerebral cortex.     

Early synaptogenesis in the spinal cord of human embryos

Synaptogenesis in the spinal cord was studied by electron microscopy in human embryos ranging from four to six weeks of intrauterine life. The first synapses were found in the motor nucleus of 6-mm human embryos. A hypothesis is suggested about the role of these early synapses in neuronal development.     

Synapsoarchitechtonics of the parietal and acoustic regions of the cerebral cortex of cats]

The synapse architecture of the simcipital and auditory cortex of the cat (fields 7 and 22 after M. O. Gurevich and oth., 1929) was studied electron microscopically. In the both areas of the cortex there are much more axo-dendritic synapses that axo-somatic ones. In the upper layers the synapses are more often formed on small dendrites and thorns, while in layers IV-VI they often occur on the main trunks of large dendrites. The synapses on small branches and thorns of dendrites contain spherical vesicles, and the synapses on on large dendrites are formed by the terminals of two kinds-with flattened and spherical vesicles. The amount of axo-somatic synapses increases towards the lower layers of the cerebral cortes. The synapses on the soma and apical dendrites of the pyramid neurons always contain flattened vesicles; on the stellate neurons there occur perisynaptic terminals with sperical vesicles as well.     

Morphology of superficial postnatal cerebral cortex with special reference to synapses

Summary The ultrastructure of layers I and II of the motor cerebral cortex of rat brain has been studied at birth, 4, 7 and 14 days postnatal and in the adult.Compared with the adult, neonatal rat motor cortex exhibited a large extracellular space which decreases with increasing age. At all stages studied the neurons were seen to contain the organelles usually found in adult neurons. Growth cones were present in decreasing numbers up to 14 days old.Synapses were detectable at birth and there was an obvious increase in their number throughout the postnatal development. At the earliest stages studied there was a lack of specialization characteristic of the adult. Many synapses were either avesicular or relatively so and lacked the high degree of modification of adult pre- and postsynaptic membranes. By 7 days after birth many synapses existed which in all morphological respects resembled those of the adult, and by 14 days, the majority were of the adult type.These findings, particularly with reference to the postnatal development of synapses, have been discussed in relation to the known electrophysiological findings.The authors wish to express their thanks to Mr. R. Birchenough and Mr. J. Manston for their technical assistance.     

Comparison of Proteins Involved with Cyclic AMP Metabolism Between Synaptic Membrane and Postsynaptic Density Preparations Isolated from Canine Cerebral Cortex and Cerebellum

Synaptic membrane and postsynaptic density (PSD) fractions isolated from canine cerebral cortex and cerebellum were assayed for the following proteins: adenylate cyclase and phosphodiesterase (PDE) activities against cyclic AMP and cyclic GMP, the regulatory subunit of the cyclic AMP-dependent protein kinase, and the substrate proteins for this kinase. The results were expressed on the basis of both the protein content of the fractions and the number of synapses in the synaptic membrane fractions. The number of synapses on a constant protein content basis was about three times higher in the cerebral cortex synaptic membrane fraction than in the comparable cerebellar fraction. Adenylate cyclase activity was from 3.4 to 5.6 times higher in the cerebral cortex membrane fraction than in the cerebellar membrane fraction based on protein content but only slightly higher based on synapse counts. PSD fractions had no adenylate cyclase activity. The cyclic AMP-PDE activity was from 17 to 27 times higher in the cerebral cortex membrane fraction than in the cerebellar membrane fraction based on protein content, and about five times higher based on synapse counts. By doing PDE histochemistry at the electron microscopy level it was found that all the cerebral cortex PSDs in the isolated fraction contained PDE activity, none being found associated with the broken-up material in the fraction. The amount of the regulatory subunit of the cyclic AMP-dependent protein kinase was about equal in the two fractions based on protein, but about one-third lower in cerebral cortex fraction than in cerebellar fractions. In the cerebral cortex membrane fraction the primary substrate for the cyclic AMP-dependent protein kinase is synapsin I, with much lower amounts in the cerebellar membrane fraction. The PSD fraction from the two sources also showed these differences in synapsin I content. In the cerebellar membrane fraction, the primary substrate for the enzyme is a approximately 245,000 Mr protein not found in the cerebral cortex membrane fraction. The findings that the turnover of cyclic AMP is much higher in cerebral cortex synapses than in cerebellar synapses, and that differences are found between the cerebral cortex and cerebellum with regard to the substrate proteins for the cyclic AMP-dependent protein kinase indicate a divergence in the effect of cyclic AMP between cerebral cortex and cerebellar synapses.     

Appearance and differentiation of NADPH-D-neurons in ontogeny of the cerebral cortex in rats

The appearance of presumptive NO-ergic nerve cells and their differentiation in the rat neocortex were studied. For this purpose, a comparative analysis of the development and differentiation of NADPH-D-positive neurons in the neocortex transplants taken from the embryos of different ages and transplanted in the occipital cortex of adult rats and in the normally developing cerebral cortex. The nervous tissue was stained histochemically for NADPH-D. The results we obtained suggest that no NADPH-D-containing neurons were found in the transplants from 15-day embryos, while they developed in those from 18-day embryos. Hence, precursors of NO-ergic neurons were still absent in the presumptive neocortex of 15-day embryos and appeared only on day 16-18 of embryogenesis. Expression of NADPH-D begins in them only within four to five days, but the neurons are differentiated during a relatively short period of time. Most NADPH-D-positive neurons reach their structural-functional maturity already by the end of the first week of postnatal development, while their complete maturation takes place by the end of the second week of postnatal development.     

突触中磷蛋白的生后发育变化

为了研究在突触功能中起重要作用的磷蛋白状况,利用高分辩率的放射自显影、梯度电泳和双向电泳,以及抗ＣａＮ多克隆抗体封闭ＣａＮ磷酸酶活力等技术,并运用计算机图象处理系统,对大鼠大脑皮层突触体中磷蛋白生后发育变化进行定量分析．结果表明,大鼠出生后（ＰＮＤ）３ｄ、７ｄ、２１ｄ、和成年磷蛋白表达有很大不同,在出生后早期对应突触主要形成时期,磷蛋白呈高表达;从ＰＮＤ２１ｄ开始至成年,底物蛋白磷酸化状态逐渐降低,同时研究了突触主要形成时期有显著变化的钙调神经磷酸酶,它的内源底物及其在其生后发育所发生的变化．     

Immunohistochemical Markers of Neural Progenitor Cells in the Early Embryonic Human Cerebral Cortex

The development of the human central nervous system represents a delicate moment of embryogenesis. The purpose of this study was to analyze the expression of multiple immunohistochemical markers in the stem/progenitor cells in the human cerebral cortex during the early phases of development. To this end, samples from cerebral cortex were obtained from 4 human embryos of 11 weeks of gestation. Each sample was formalin-fixed, paraffin embedded and immunostained with several markers including GFAP, WT1, Nestin, Vimentin, CD117, S100B, Sox2, PAX2, PAX5, Tβ4, Neurofilament, CD44, CD133, Synaptophysin and Cyclin D1. Our study shows the ability of the different immunohistochemical markers to evidence different zones of the developing human cerebral cortex, allowing the identification of the multiple stages of differentiation of neuronal and glial precursors. Three important markers of radial glial cells are evidenced in this early gestational age: Vimentin, Nestin and WT1. Sox2 was expressed by the stem/progenitor cells of the ventricular zone, whereas the postmitotic neurons of the cortical plate were immunostained by PAX2 and NSE. Future studies are needed to test other important stem/progenitor cells markers and to better analyze differences in the immunohistochemical expression of these markers during gestation.Key words: Cerebral cortex, human embryo, human development, immunohistochemistry, fetal stem cells     

Glutamate decarboxylase in developing rat neocortex: Does it correlate with the differentiation of GABAergic neurons and synapses?

Postnatal development of glutamate decarboxylase was studied in the rat cerebral cortex. Two methods were used: estimation of the enzymatic activity of glutamate decarboxylase in homogenates of developing cortical tissue and visualization of structures containing glutamate decarboxylase-like immunoreactivity. Glutamate decarboxylase-like immunoreactivity appeared first in perikarya and dendrites and only later in axons and axon varicosities. The most rapid increase in the glutamate decarboxylase activity took place during the second postnatal week and this coincided with a rapid increase in the density of axon varicosities containing glutamate decarboxylase-like immunoreactivity but preceded the most rapid phase in the formation of GABAergic synapses by several days. However, there was a change in the characteristics of glutamate decarboxylase which correlated with GABA synaptogenesis: two fractions of glutamate decarboxylase with different sensitivities to the activating effects of Triton X-100 could be distinguished as from about the time when most of the GABAergic synapses are formed.     

Appearance and Differentiation of NADPH-D-Neurons in Ontogeny of Cerebral Cortex in Rats

The appearance of presumptive NO-ergic nerve cells and their differentiation in the rat neocortex were studied. For this purpose, a comparative analysis of the development and differentiation of NADPH-D-positive neurons in the neocortex transplants taken from the embryos of different ages and transplanted in the occipital cortex of adult rats and in the normally developing cerebral cortex was undertaken. The nervous tissue was stained histochemically for NADPH-D. The results we obtained suggest that no NADPH-D-containing neurons were found in the transplants from 15-day embryos, while they developed in those from 18-day embryos. Hence, precursors of NO-ergic neurons were still absent in the presumptive neocortex of 15-day embryos and appeared only on day 16–18 of embryogenesis. Expression of NADPH-D begins in them only within four to five days, but the neurons are differentiated during a relatively short period of time. Most NADPH-D-positive neurons reach their structural–functional maturity already by the end of the first week of postnatal development, while their complete maturation takes place by the end of the second week of postnatal development.     

PACAP decides neuronal laminar fate via PKA signaling in the developing cerebral cortex

Laminar formation in the developing cerebral cortex requires the precisely regulated generation of phenotype-specified neurons. To test the possible involvement of pituitary adenylate cyclase-activating polypeptide (PACAP) in this formation, we investigated the effects of PACAP administered into the telencephalic ventricular space of 13.5-day-old mouse embryos. PACAP partially inhibited the proliferation of cortical progenitors and altered the position and gene-expression profiles of newly generated neurons otherwise expected for layer IV to those of neurons for the deeper layers, V and VI, of the cerebral cortex. The former and latter effects were seen only when the parent progenitor cells were exposed to PACAP in the later and in earlier G1 phase, respectively; and these effects were suppressed by co-treatment with a protein kinase A (PKA) inhibitor. These observations suggest that PACAP participates in the processes forming the neuronal laminas in the developing cortex via the intracellular PKA pathway.     

Paramembraneous microfilamentous structures of the synapses of rat cerebral cortex during sensitization by brain antigens

On the material contrasted by phosphotungstic acid the state of paramembranes microfilament structures of interneuronal contacts of molecular layer of sensomotor field cortex of rat brain of Krushynsky-Molodkina line during sensitization by homologous brain antigens was studied. Sharp reduction of general density of synapses and symmetric contacts because of damaging of paramembranes microfilament structures of cerebral cortex synapses and reduction of new contacts was proved that plays an essential role in changing of brain integrating activity during different pathological processes connected with autoimmune mechanism of neuronal tissue damaging.     

Mosaic Expression of the Reeler and Normal Phenotypes in the Cerebral Cortex in Reeler-Normal Chimeras at a Late Embryonic Stage

The cerebral cortex of reeler -normal chimera embryos was studied by hematoxylin-eosin staining and fractographic scanning electron microscopy in comparison with the cortices of normal and reeler mutant mice. The cerebral cortex of normal mice had a plexiform layer, which was composed of a fine meshwork of matrix cell processes. Spindle shaped neuroblasts formed a radial lining columnar structure, which was formed by attachment of migrating neuroblasts to the radial bundles. The cerebral cortex of reeler mutants did not show a plexiform layer and the cells were round with no radially columnar structures, and no radial bundles. In reeler -normal chimera embryos, the thickness of the plexiform layer varied in different parts of the cerebral cortex. In parts where the plexiform layer was present, neuroblasts were spindle-shaped and had a radially oriented columnar structure (normal type). But where the plexiform layer was absent, the neuroblasts were round with a radial architecture ( reeler type). Intermediates between the reeler and control types were also observed. Since mosaic expression of the two phenotypes, was observed in chimeras, the reeler abnormality is apparently not caused by humoral factors. The possible mechanism of cell migration is discussed.     

The role of spike timing for thalamocortical processing

Although the response properties of sensory neurons in the thalamus and cerebral cortex have been studied for decades, relatively few studies have examined how sensory information is processed at thalamocortical synapses. Recent studies now show that the strength of thalamocortical connections is very dynamic and spike timing plays an important role in determining whether action potentials will be transferred from thalamus to cortex.     

Quantitative studies of postnatal changes in synapses in rat superficial motor cerebral cortex

Summary Quantitation of synapses at different postnatal ages has been undertaken in the cerebral cortex of the rat. In this study axial ratios of presynaptic bags, proportion of cortex occupied by presynaptic bags and numbers of synapses per unit volume of cortex have been estimated. Observations on synaptic vesicle packing densities have also been made.Synaptic bags become increasingly spherical up to 7 days of age and become more elongated thereafter. The proportion of cortex occupied by presynaptic bags increases rapidly up to 7 days of age and then at a decelerated rate up to maturity. The number of synapses per unit volume increases slowly over the first four days after which there is a rapid increase to 14 days, followed by a decelerated rate.The average presynaptic bag shows marked changes in volume with increasing age which indicate the probability of two stages of synaptic development. This two stage development is further reflected in the estimates on vesicle packing densities. The implications of the results are discussed in relationship to changes in functional activity of the cortex during postnatal development.The authors wish to express their thanks to Mr. R. Birchenough and Mr. J. Manston for much technical assistance.     

Distribution of N-cadherin in human cerebral cortex during prenatal development

An important subgroup of adhesion molecules is the superfamily of cadherins, which takes part in cell recognition and differentiation during development. To our knowledge only one study describing N-cadherin expression in developing human brain has been performed so far. Our aim is to identify N-cadherin expression to establish a relationship between its expression and function in human cerebral cortex during prenatal development. In the present study, localization and intensity of N-cadherin was investigated in developing cerebral cortex. Fetuses from spontaneous abortions (n=13) were obtained from first, second, and third trimesters. Western blot analysis revealed three bands and the third trimester samples showed the strongest bands for N-cadherin. Cell processes, axon bundles, and some of the developing neurons revealed immunoreactivity for N-cadherin throughout pregnancy. The immunoreactivity increased in the developing neocortex and expanded from the ventricular layer toward the marginal zone as development progressed. Moreover, the immunoreactivity was strong in vascular endothelium during all three trimesters. We conclude that N-cadherin is dynamically related to the organization of cerebral cortex layers during prenatal development. The dynamic expression pattern implicates N-cadherin as a potential regulator of cell migration, axon extension and fasciculation, the establishment of synaptic contacts, and neurovascular angiogenesis in the developing human cerebral cortex.     

Effect of delta-sleep peptide on the ultrastructural features of the rat sensomotor cortex

The performed studies have shown that injection of DSIP causes an activation of nuclear apparatus and plastic metabolism in III-Y layer of the rat cerebral cortex. The activation of axo-axon and axosomatic synapses indicates the modulation of inhibitory processes in the cerebral cortex.     

An electron microscopic study of neuronal development in organotypic cultures of the anlage of the cerebral cortex from a human embryo

Data are provided on cytodifferentiation of the cerebral cortex cultured cells taken from 10-12 week old embryos of man. It is shown that low differentiated neuroblasts well survive in culture for 21 days. Mature granular cells and middle pyramidal neurons are revealed in cultures. The number of morphological criteria may testify to the maturity of neurons: the presence of the Nissl substance, differentiation of dendrites and axons; the presence of various types of synapses. The absence of myelinized fibres testifies to the insufficient maturity of the cultures, that is probably associated with employing the low differentiated nervous tissue for cultivation and with insufficient cultivation period.     

Approximate invariance of metabolic energy per synapse during development in mammalian brains

During mammalian development the cerebral metabolic rate correlates qualitatively with synaptogenesis, and both often exhibit bimodal temporal profiles. Despite these non-monotonic dependencies, it is found based on empirical data for different mammals that regional metabolic rate per synapse is approximately conserved from birth to adulthood for a given species (with a slight deviation from this constancy for human visual and temporal cortices during adolescence). A typical synapse uses about (7±2)×10(3) glucose molecules per second in primate cerebral cortex, and about five times of that amount in cat and rat visual cortices. A theoretical model for brain metabolic expenditure is used to estimate synaptic signaling and neural spiking activity during development. It is found that synaptic efficacy is generally inversely correlated with average firing rate, and, additionally, synapses consume a bulk of metabolic energy, roughly 50-90% during most of the developmental process (except human temporal cortex < 50%). Overall, these results suggest a tight regulation of brain electrical and chemical activities during the formation and consolidation of neural connections. This presumably reflects strong energetic constraints on brain development.