OBJECTIVE: To clarify the functional effects of differential expression of ring finger and tryptophan-aspartic acid 2 (RFWD2) on dendritic development and formation of dendritic spines in cerebral cortex neurons of mice. METHODS: Immunofluorescent staining was used to identify the location and global expression profile of RFWD2 in mouse brain and determine the co-localization of RFWD2 with the synaptic proteins in the cortical neurons. We also examined the effects of RFWD2 over-expression (RFWD2-Myc) and RFWD2 knockdown (RFWD2-shRNA) on dendritic development, dendritic spine formation and synaptic function in cultured cortical neurons. RESULTS: RFWD2 is highly expressed in the cerebral cortex and hippocampus of mice, and its expression level was positively correlated with the development of cerebral cortex neurons and dendrites. RFWD2 expression was detected on the presynaptic membrane and postsynaptic membrane of the neurons, and its expression levels were positively correlated with the length, number of branches and complexity of the dendrites. In cultured cortical neurons, RFWD2 overexpression significantly lowered the expressions of the synaptic proteins synaptophysin (P < 0.01) and postsynapic density protein 95 (P < 0.01), while RFWD2 knockdown significantly increased their expressions (both P < 0.05). Compared with the control and RFWD2-overexpressing cells, the neurons with RFWD2 knockdown showed significantly reduced number of dendritic spines (both P < 0.05). CONCLUSION RFWD2 can regulate the expression of the synaptic proteins, the development of the dendrites, the formation of the dendritic spines and synaptic function in mouse cerebral cortex neurons through ubiquitination of Pea3 family members and c-Jun, which may serve as potential treatment targets for neurological diseases.
Animals ; Aspartic Acid/metabolism* ; Cerebral Cortex ; Dendritic Spines/metabolism* ; Mice ; Neurons/metabolism* ; Synapses ; Tryptophan/metabolism*

OBJECTIVETo observe a turning performance in the rats excited by using a proper electrical stimuli of the barrel cortex region (BC), and the expression of choline acetyltransferase (ChAT) in the BC regions after electoral stimulation.

METHODSSD rats were divided into three groups. The stimulation electrodes were surgically implanted into the bilateral BC regions in the control group and the experimental group rats. The experiment group post surgery for seven days was given the electrical impulses via connection with the electrodes for three times each day through consecutive three days. Three groups of the rats were killed and the brains were quickly removed for frozen sections and then performed with conventional HE and immunohistochemistry staining. And protein samples were prepared from brain and the hippocampus tissues of the three groups to detect the level of the ChAT protein by Western blot.

RESULTSThe experimental rats turn left or right when continuously stimulation in the bilateral BC regions with electric pulse. HE staining showed no significant damage around electrodes in the cerebral cortex. Compared with the control and blank groups, the ChAT positive rate in the brain section in the experimental rats was significantly high by immunohistochemistry assay; the level of the ChAT protein in the rats given the electrical stimulation increased.

CONCLUSIONTurnings performance of the rat could be initiated hy electrical stimuli in the BC region. Expression of ChAT is significantly higher in the BC regions of rat under electrical stimulation, suggesting that acetylcholine might be associated with signal transmission between senses and movement behavior in the nervous central system.

Acetylcholine ; metabolism ; Animals ; Cerebral Cortex ; metabolism ; Choline O-Acetyltransferase ; metabolism ; Electric Stimulation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the differences of protein expression levels in the brain cortex of human fetus and adult with proteomics technique, and provide preliminary data on the change of proteins during brain development.

METHODSProteins extracted from human temporal lobes in fetal (3 month and 5 month respectively) and adult (30 years old) brain were separated by two-dimensional gel electrophoresis (2DE). The proteins were then stained with colloidal Coomassie blue to produce a high-resolution map of the proteiome. The differential protein spots were analyzed by PDQuest 7.0 software and 8 spots, which were gradually reduced or gradually increased in brain development process and the protein spots of difference over two-fold in the brain, were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF/TOF).

RESULTS(1) On average, 642, 511 and 527 protein spots could be obtained in the temporal lobes of adult, 3 month and 5 month fetus. The matching rate of images was 87%. The basic proteins in adult brain were obviously much more than that in the fetus; (2) There were 172, 171 and 152 singular protein spots in temporal lobes of adult, 3 month and 5 month fetus respectively.(3) Compared with adult, there were 131 and 115 different protein spots in the 3 month and 5 month fetus respectively. There were 60 and 40 protein spots with more than 2 fold difference, among which 24 and 17 were down-regulated, and 36 and 23 were up-regulated respectively. (4) There was different expression in proteins such as serum albumin, triosephosphate isomerase, etc. in the 3 groups. Fatty acid binding protein 7 and unnamed proteins were only highly expressed in the 3 month brain; ribulose-1, 5-bisphosphate carboxylase/oxygenase large subunit and transducin beta-1 subunit were up-regulated in adult brain. Serum albumin decreases gradually with brain development. However, ATP synthase, mitochondrial F0 complex, and triosephosphate isomerase increase gradually with brain development.

CONCLUSIONThe proteins of human brain cortex were obviously changed from embryonic stage to adult. The differentially displayed proteins may provide further insight into the understanding of development of human brain.

Adult ; Cerebral Cortex ; metabolism ; Electrophoresis, Gel, Two-Dimensional ; Fetus ; metabolism ; Humans ; Mass Spectrometry ; Proteins ; metabolism ; Proteomics

The aim of this study was to investigate the harmful effects of acute hypoxia on mouse cerebral cortex and hippocampus and the underlying mechanism. Mouse model of acute hypoxia was constructed by using a sealed glass jar. Laser speckle contrast imaging was used to detect the changes of cerebral blood flow after different time duration of hypoxia. Total superoxide dismutase (T-SOD) and malondialdehyde (MDA) assay kits were used to detect oxidative stress in cerebral cortex and hippocampus. Immunofluorescent staining was used to detect neuroinflammatory response of microglia in the cerebral cortex and hippocampus. One-step TUNEL method was used to detect neuronal apoptosis. The results showed that, compared with non-hypoxia (0 min hypoxia) group, 30 min hypoxia group exhibited decreased cerebral blood flow, higher percentage of CD68⁺/Iba1⁺ microglia, and increased neural apoptosis in the cerebral cortex and hippocampus. Compared with 30 min group, 60 min hypoxia group showed significantly decreased cerebral blood flow, increased MDA content in the cortex, as well as greater percentage of CD68⁺/Iba1⁺ microglia and neuronal apoptosis in the cerebral cortex and hippocampus. These results suggest that acute hypoxia damages brain tissue in a time-dependent manner and the oxidative stress and neuroinflammation are important mechanisms.
Animals ; Cerebral Cortex/metabolism* ; Hippocampus/metabolism* ; Hypoxia ; Malondialdehyde ; Mice ; Oxidative Stress ; Superoxide Dismutase/pharmacology*

Heterozygous loss-of-function variants of FOXP4 are associated with neurodevelopmental disorders (NDDs) that exhibit delayed speech development, intellectual disability, and congenital abnormalities. The etiology of NDDs is unclear. Here we found that FOXP4 and N-cadherin are expressed in the nuclei and apical end-feet of radial glial cells (RGCs), respectively, in the mouse neocortex during early gestation. Knockdown or dominant-negative inhibition of Foxp4 abolishes the apical condensation of N-cadherin in RGCs and the integrity of neuroepithelium in the ventricular zone (VZ). Inhibition of Foxp4 leads to impeded radial migration of cortical neurons and ectopic neurogenesis from the proliferating VZ. The ectopic differentiation and deficient migration disappear when N-cadherin is over-expressed in RGCs. The data indicate that Foxp4 is essential for N-cadherin-based adherens junctions, the loss of which leads to periventricular heterotopias. We hypothesize that FOXP4 variant-associated NDDs may be caused by disruption of the adherens junctions and malformation of the cerebral cortex.
Mice ; Animals ; Ependymoglial Cells/physiology* ; Cadherins ; Neurons/metabolism* ; Cerebral Cortex/metabolism* ; Cell Differentiation ; Cell Movement

TrkA is essential components of the high-affinity NGF receptor necessary to mediate biological effects of the neurotrophins NGF. Here we report on the expression of trkA in the cerebral cortex and diencephalon of mongolian gerbils during postnatal development. The expression of trkA was identified by immunohistochemical method. In parietal cortex and piriform cortex, higher levels of trkA-IR (immunoreactivity) were detected at 3 days postnatal (P3) and at P9. Although trkA was not expressed till P3 in the parietal cortex, it was detectable at birth in the piriform cortex. Several regions, such as Layers I, IV & VI, did not show much expression. Layer I showed especially weak labeling. In the hippocampus, thalamus, and hypothalamus, higher levels of trkA-IR were detected at P6 and P12 than earlier days. But trkA was not expressed at birth in the hippocampus, at P3 in the reticular thalamic nucleus (Rt), or neonatally in the dorsomedial hypothalamic nucleus (DM). This data shows that expression of trkA is developmentally regulated and suggests that high affinity neurotrophin-receptors mediate a transient response to neurotrophines in the cerebral cortex and diencephalon during mongolian gerbil brain ontogeny.
Animals ; Animals, Newborn ; Cerebral Cortex/*metabolism ; Diencephalon/*metabolism ; Gerbillinae/*metabolism ; Immunohistochemistry/veterinary ; Nerve Growth Factor/metabolism ; Receptor, trkA/*metabolism

OBJECTIVETo screen differentially expressed brain proteins with proteomic method in cerebral cortex of neonatal rats with congenital hypothyroidism.

METHODFrom the 13th day of gestation, pregnant Wistar rats from the experimental group were given intragastrically with 2.5 ml of 1% propylthiouracil daily. Cerebral cortex specimens were collected from the control and hypothyroidism neonatal rats. Two-directional electrophoresis (2-DE) was applied to analyze protein expression diversities between the euthyroid and hypothyroidism neonatal rat cerebral cortex. Protein spots with significantly different expression were screened and identified by mass spectrometry. Radioimmunoassay (RIA) was used to analyze serum FT(3), FT(4) levels of each groups.

RESULTThe body weight of hypothyroid neonatal rats were lower than those in the corresponding control group (t = -8.07, P < 0.01). The FT(3) levels of hypothyroid neonatal rats were lower than those in the corresponding control group (t = 5.39, P < 0.01). The FT(4) levels of hypothyroid neonatal rats were lower than those in the corresponding control group (t = 7.62, P < 0.01). Stable 2-DE maps of normal and CH neonatal rat were constantly obtained. The maps were analyzed by software. Seven protein spots with high reproducibility, high resolution and significantly different expression were chosen and identified by mass spectrometry, including collapsing response mediator protein 2, actin related protein 2/3 complex subunit 5, ubiquitin-conjugating enzyme E2-25K, ATP synthase subunit d, Cu-Zn superoxide dismutase, synuclein alpha, and nucleoside diphosphate kinase.

CONCLUSIONThe value of this research is demonstrated here by the identification of several proteins known to be associated with nerve synapse structures formation, cell survival, metabolism, cell signal transduction, neural differentiation and nerve growth in the central nervous system. Furthermore this study identified several proteins except for collapsing response mediator protein 2 and Cu-Zn superoxide dismutase that have not previously been described in the literature and which may play an important role as either sensitive biomarkers of brain dysfunction caused by congenital hypothyroidism. In congenital hypothyroidism, brain development retardation may be related with some important processes, including abnormal synaptic formation, excess ROS production and apoptosis. The above-mentioned proteins may play critical roles in the processes, which provide valuable clues to clarify the pathogenesis of brain developmental disorders induced by congenital hypothyroidism.

Animals ; Animals, Newborn ; metabolism ; Brain ; metabolism ; Cerebral Cortex ; metabolism ; Congenital Hypothyroidism ; metabolism ; Female ; Pregnancy ; Proteome ; analysis ; Proteomics ; Rats

Animals ; Cerebral Cortex ; metabolism ; Disease Models, Animal ; Male ; Mice ; Mice, Inbred Strains ; Morphine Dependence ; metabolism ; Neurotrophin 3 ; metabolism ; Prosencephalon ; metabolism

ER-α36, a novel variant of ER-α, is expressed in breast, uterus, digestive tract, respiratory tract etc. The aim of the present study was to investigate the distribution and expression of ER-α36 in the central nervous system (CNS). Here, we comparatively analyzed the expression pattern of ER-α36 in the hippocampus and cortex of neonatal (1-day-old) and adult (12-week-old) Sprague-Dawley (SD) rats by using immunohistochemistry/immunocytochemistry analysis and Western blot. The results showed that ER-α36 was expressed both in hippocampus and cortex of adult rats, but mainly distributed in pyramidal neurons. ER-α36 was mainly located on the cytomembrane of hippocampal and cortical neurons from neonatal rats. Compared with the cortical neurons, the hippocampal neurons showed lower ER-α36 protein expression in the neonatal rats, but exhibited higher level of ER-α36 in the adult rats. Furthermore, the adult rats showed higher levels of ER-α36 expression in both hippocampus and cortex compared with the neonatal rats. These results suggest that ER-α36 might be involved in the regulation of membrane-initiated estrogen signaling throughout the postnatal development of diverse brain regions, and thus will be a potential target for the treatment of degenerative diseases in nervous system.
Animals ; Cerebral Cortex ; metabolism ; Estrogen Receptor alpha ; metabolism ; Hippocampus ; metabolism ; Pyramidal Cells ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

Animals ; Brain Chemistry ; Brain Ischemia ; metabolism ; physiopathology ; Cerebral Cortex ; metabolism ; Dementia ; metabolism ; physiopathology ; Humans ; Microdialysis ; Reperfusion Injury ; metabolism ; physiopathology