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*

OBJECTIVE: To investigate the mechanism of valproic acid (VPA) -induced impairment of the dendritic spines and synapses in the prefrontal cortex (PFC) for causing core symptoms of autism spectrum disorder (ASD) in mice. METHODS: Female C57 mice were subjected to injections of saline or VPA on gestational days 10 and 12, and the male offspring mice in the two groups were used as the normal control group and ASD model group (n=10), respectively. Another 20 male mice with fetal exposure to VPA were randomized into two groups for stereotactic injection of DMSO or Wortmannin into the PFC (n=10). Open field test, juvenile play test and 3-chamber test were used to evaluate autistic behaviors of the mice. The density of dendrite spines in the PFC was observed with Golgi staining. Western blotting and immunofluorescence staining were used to detect the expressions of p-PI3K, PI3K, p-AKT, AKT, p-mTOR, mTOR and the synaptic proteins PSD95, p-Syn, and Syn in the PFC of the mice. RESULTS: Compared with the normal control mice, the mice with fetal exposure to VPA exhibited obvious autism-like behaviors with significantly decreased density of total, mushroom and stubby dendritic spines (P < 0.05) and increased filopodia dendritic spines (P < 0.05) in the PFC. The VPA-exposed mice also showed significantly increased expressions of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR (P < 0.01) and lowered expressions of PSD95 and p-Syn/Syn in the PFC (P < 0.05 or 0.001). Wortmannin injection into the PFC obviously improved the ASD-like phenotype and dendritic spine development, down-regulated PI3K/Akt/mTOR signaling pathway and up-regulated the synaptic proteins in VPA-exposed mice. CONCLUSION In male mice with fetal exposure to VPA, excessive activation of PI3K/Akt/mTOR signaling pathway and decreased expressions of the synaptic proteins PSD95 and p-Syn cause dendritic spine damage and synaptic development disturbance in the PFC, which eventually leads to ASD-like phenotype.
Animals ; Autism Spectrum Disorder/chemically induced* ; Autistic Disorder/chemically induced* ; Dendritic Spines ; Disease Models, Animal ; Female ; Male ; Mice ; Phosphatidylinositol 3-Kinases ; Prefrontal Cortex ; Prenatal Exposure Delayed Effects ; Valproic Acid/adverse effects*

A transient ischemic attack (TIA) can cause reversible and delayed impairment of cognition, but the specific mechanisms are still unclear. Annexin a1 (ANXA1) is a phospholipid-binding protein. Here, we confirmed that cognition and hippocampal synapses were impaired in TIA-treated mice, and this could be rescued by multiple mild stimulations (MMS). TIA promoted the interaction of ANXA1 and CX3CR1, increased the membrane distribution of CX3CR1 in microglia, and thus enhanced the CX3CR1 and CX3CL1 interaction. These phenomena induced by TIA could be reversed by MMS. Meanwhile, the CX3CR1 membrane distribution and CX3CR1-CX3CL1 interaction were upregulated in primary cultured microglia overexpressing ANXA1, and the spine density was significantly reduced in co-cultured microglia overexpressing ANXA1 and neurons. Moreover, ANXA1 overexpression in microglia abolished the protection of MMS after TIA. Collectively, our study provides a potential strategy for treating the delayed synaptic injury caused by TIA.
Animals ; Annexin A1/metabolism* ; CX3C Chemokine Receptor 1/metabolism* ; Chemokine CX3CL1 ; Cognition ; Dendritic Spines/metabolism* ; Ischemic Attack, Transient ; Mice ; Microglia/metabolism*

Alzheimer's disease (AD) is the most common neurodegenerative disorder causing dementia worldwide, and is mainly characterized by aggregated β-amyloid (Aβ). Increasing evidence has shown that plant extracts have the potential to delay AD development. The plant sterol β-Sitosterol has a potential role in inhibiting the production of platelet Aβ, suggesting that it may be useful for AD prevention. In the present study, we aimed to investigate the effect and mechanism of β-Sitosterol on deficits in learning and memory in amyloid protein precursor/presenilin 1 (APP/PS1) double transgenic mice. APP/PS1 mice were treated with β-Sitosterol for four weeks, from the age of seven months. Brain Aβ metabolism was evaluated using ELISA and Western blotting. We found that β-Sitosterol treatment can improve spatial learning and recognition memory ability, and reduce plaque load in APP/PS1 mice. β-Sitosterol treatment helped reverse dendritic spine loss in APP/PS1 mice and reversed the decreased hippocampal neuron miniature excitatory postsynaptic current frequency. Our research helps to explain and support the neuroprotective effect of β-Sitosterol, which may offer a novel pharmaceutical agent for the treatment of AD. Taken together, these findings suggest that β-Sitosterol ameliorates memory and learning impairment in APP/PS1 mice and possibly decreases Aβ deposition.
Alzheimer Disease ; Amyloid ; Animals ; Blood Platelets ; Blotting, Western ; Brain ; Cognition Disorders ; Dementia ; Dendritic Spines ; Enzyme-Linked Immunosorbent Assay ; Excitatory Postsynaptic Potentials ; Learning ; Memory ; Metabolism ; Mice ; Mice, Transgenic ; Neurodegenerative Diseases ; Neurons ; Neuroprotective Agents ; Plant Extracts ; Plants ; Plaque, Amyloid ; Spatial Learning

MicroRNAs (miRNAs) are critical for both development and function of the central nervous system. Significant evidence suggests that abnormal expression of miRNAs is associated with neurodevelopmental disorders. MeCP2 protein is an epigenetic regulator repressing or activating gene transcription by binding to methylated DNA. Both loss-of-function and gain-of-function mutations in the MECP2 gene lead to neurodevelopmental disorders such as Rett syndrome, autism and MECP2 duplication syndrome. In this study, we demonstrate that miR-130a inhibits neurite outgrowth and reduces dendritic spine density as well as dendritic complexity. Bioinformatics analyses, cell cultures and biochemical experiments indicate that miR-130a targets MECP2 and down-regulates MeCP2 protein expression. Furthermore, expression of the wild-type MeCP2, but not a loss-of-function mutant, rescues the miR-130a-induced phenotype. Our study uncovers the MECP2 gene as a previous unknown target for miR-130a, supporting that miR-130a may play a role in neurodevelopment by regulating MeCP2. Together with data from other groups, our work suggests that a feedback regulatory mechanism involving both miR-130a and MeCP2 may serve to ensure their appropriate expression and function in neural development.
Animals ; Dendrites ; genetics ; metabolism ; Dendritic Spines ; genetics ; metabolism ; Down-Regulation ; physiology ; Methyl-CpG-Binding Protein 2 ; biosynthesis ; genetics ; MicroRNAs ; genetics ; metabolism ; Rats

GIT1, a multifunctional signaling adaptor protein, is implicated in the development of dendritic spines and neuronal synapses. GIT1 forms a signaling complex with PIX, RAC, and PAK proteins that is known to play important roles in brain development. Here we found that Git1-knockout (Git1-/-) mice show a microcephaly-like small brain phenotype, which appears to be caused by reduced neuronal size rather than number. Git1-/- mice also show decreased dendritic spine number without morphological alterations in the hippocampus. Behaviorally, Git1-/- mice show impaired motor coordination and learning and memory. In addition, adult dGit Drosophila mutants show decreased brain size and abnormal morphology of the mushroom body. These results suggest that GIT1 is important for brain development in both rodents and flies.
Adult ; Animals ; Brain* ; Dendritic Spines ; Diptera ; Drosophila ; Hippocampus ; Humans ; Invertebrates* ; Learning ; Memory ; Mice ; Microcephaly ; Mushroom Bodies ; Neurons ; Phenotype ; Rodentia ; Synapses ; Vertebrates*

BACKGROUND: Adult neural stem cells have the potential for self-renewal and differentiation into multiple cell lineages via symmetric or asymmetric cell division. Preso1 is a recently identified protein involved in the formation of dendritic spines and the promotion of axonal growth in developing neurons. Preso1 can also bind to cell polarity proteins, suggesting a potential role for Preso1 in asymmetric cell division. METHODS: To investigate the distribution of Preso1, we performed immunohistochemistry with adult mouse brain slice. Also, polarized distribution of Preso1 was assessed by immunocytochemistry in cultured neural stem cells. RESULTS: Immunoreactivity for Preso1 (Preso1-IR) was strong in the rostral migratory stream and subventricular zone, where proliferating transit-amplifying cells and neuroblasts are prevalent. In cultured neural stem cells, Preso1-IR was unequally distributed in the cell cytosol. We also observed the distribution of Preso1 in the subgranular zone of the hippocampal dentate gyrus, another neurogenic region in the adult brain. Interestingly, Preso1-IR was transiently observed in the nuclei of doublecortin-expressing neuroblasts immediately after asymmetric cell division. CONCLUSION: Our study demonstrated that Preso1 is asymmetrically distributed in the cytosol and nuclei of neural stem/progenitor cells in the adult brain, and may play a significant role in cell differentiation via association with cell polarity machinery.
Adult* ; Animals ; Asymmetric Cell Division ; Axons ; Brain ; Cell Differentiation ; Cell Lineage ; Cell Polarity ; Cytosol ; Dendritic Spines ; Dentate Gyrus ; Humans ; Immunohistochemistry ; Mice ; Neural Stem Cells ; Neurons ; Rivers

BACKGROUNDThe Ras/Raf/ERK1/2 signaling pathway controls many cellular responses such as cell proliferation, migration, differentiation, and death. In the nervous system, emerging evidence also points to a death-promoting role for ERK1/2 in both in vitro and in vivo models of neuronal death. To further investigate how Ras/Raf/ERK1/2 up-regulation may lead to the development of spinal cord injury, we developed a cellular model of Raf/ERK up-regulation by overexpressing c-Raf in cultured spinal cord neurons (SCNs) and dorsal root ganglions (DRGs).

METHODSDRGs and SCNs were prepared from C57BL/6J mouse pups. DRGs or SCNs were infected with Ad-Raf-1 or Ad-Null adenovirus alone. Cell adhesion assay and cell migration assay were investigated, DiI labeling was employed to examine the effect of the up-regulation of Ras/Raf/ERK1/2 signaling on the dendritic formation of spinal neurons. We used the TO-PRO-3 staining to examine the apoptotic effect of c-Raf on DRGs or SCNs. The effect on the synapse formation of neurons was measured by using immunofluorescence.

RESULTSWe found that Raf/ERK up-regulation stimulates the migration of both SCNs and DRGs, and impairs the formation of excitatory synapses in SCNs. In addition, we found that Raf/ERK up-regulation inhibits the development of mature dendritic spines in SCNs. Investigating the possible mechanisms through which Raf/ERK up-regulation affects the excitatory synapse formation and dendritic spine development, we discovered that Raf/ERK up-regulation suppresses the development and maturation of SCNs.

CONCLUSIONThe up-regulation of the Raf/ERK signaling pathway may contribute to the pathogenesis of spinal cord injury through both its impairment of the SCN development and causing neural circuit imbalances.

Animals ; Cell Movement ; physiology ; Dendritic Spines ; metabolism ; physiology ; Female ; Ganglia, Spinal ; cytology ; MAP Kinase Signaling System ; physiology ; Mice ; Neurogenesis ; genetics ; physiology ; Neurons ; cytology ; Pregnancy ; Signal Transduction ; genetics ; physiology ; Spinal Cord ; cytology ; Synapses ; metabolism ; physiology ; Up-Regulation ; raf Kinases ; genetics ; metabolism ; ras Proteins ; genetics ; metabolism

Orthostatic hypotension (OH) is defined by a 20-mm Hg difference of systolic blood pressure (dtSBP) and/or a 10-mm Hg difference of diastolic blood pressure (dtDBP) between supine and standing, and OH is associated with a failure of the cardiovascular reflex to maintain blood pressure on standing from a supine position. To understand the underlying genetic factors for OH traits (OH, dtSBP, and dtDBP), genome-wide association studies (GWASs) using 333,651 single nucleotide polymorphisms (SNPs) were conducted separately for two population-based cohorts, Ansung (n = 3,173) and Ansan (n = 3,255). We identified 8 SNPs (5 SNPs for dtSBP and 3 SNPs for dtDBP) that were repeatedly associated in both the Ansung and Ansan cohorts and had p-values of <1 x 10(-5) in the meta-analysis. Unfortunately, the SNPs of the OH case control GWAS did not pass our p-value criteria. Four of 8 SNPs were located in the intergenic region of chromosome 2, and the nearest gene (CTNNA2) was located at 1 Mb of distance. CTNNA2 is a linker between cadherin adhesion receptors and the actin cytoskeleton and is essential for stabilizing dendritic spines in rodent hippocampal neurons. Although there is no report about the function in blood pressure regulation, hippocampal neurons interact primarily with the autonomic nervous system and might be related to OH. The remaining SNPs, rs7098785 of dtSBP trait and rs6892553, rs16887217, and rs4959677 of dtDBP trait were located in the PIK3AP1 intron, ACTBL2-3' flanking, STAR intron, and intergenic region, respectively, but there was no clear functional link to blood pressure regulation.
Actin Cytoskeleton ; Autonomic Nervous System ; Blood Pressure ; Case-Control Studies ; Chromosomes, Human, Pair 2 ; Cohort Studies ; Dendritic Spines ; DNA, Intergenic ; Genome-Wide Association Study ; Hypotension, Orthostatic ; Introns ; Neurons ; Polymorphism, Single Nucleotide ; Reflex ; Rodentia ; Supine Position

The prenatal ethanol exposure induced the alterations of dendritic spine and synapse in visual cortex and their long-term effect would be investigated in mice from P0 to P30. Pregnant mice were intubated ethanol daily from E5 through the pup's birth to establish mode of prenatal alcohol abuse. The dendritic spines of pyramidal cells in visual cortex of pups were labeled with DiI diolistic assay, and the synaptic ultrastructure was observed under transmission electron microscope. Prenatal alcohol exposure was associated with a significant decrease in the number of dendritic spines of pyramidal neurons in the visual cortex and an increase in their mean length; ultrastructural changes were also observed, with decreased numbers of synaptic vesicles, narrowing of the synaptic cleft and thickening of the postsynaptic density compared to controls. Prenatal alcohol exposure is associated with long-term changes in dendritic spines and synaptic ultrastructure. The changes were dose-dependent with long term effect even at postnatal 30.
Animals ; Dendritic Spines ; ultrastructure ; Ethanol ; toxicity ; Female ; Fetal Alcohol Spectrum Disorders ; etiology ; pathology ; Male ; Mice ; Mice, Inbred C57BL ; Microscopy, Confocal ; Microscopy, Electron, Transmission ; Pregnancy ; Prenatal Exposure Delayed Effects ; pathology ; Pyramidal Cells ; ultrastructure ; Synapses ; ultrastructure ; Visual Cortex ; ultrastructure