Autism Spectrum Disorder (ASD) is marked by early-onset neurodevelopmental anomalies, yet the temporal dynamics of genetic contributions to these processes remain insufficiently understood. This study aimed to elucidate the role of the Shank3 gene, known to be associated with monogenic causes of autism, in early developmental processes to inform the timing and mechanisms for potential interventions for ASD. Utilizing the Shank3B knockout (KO) mouse model, we examined Shank3 expression and its impact on neuronal maturation through Golgi staining for dendritic morphology and electrophysiological recordings to measure synaptic function in the anterior cingulate cortex (ACC) across different postnatal stages. Our longitudinal analysis revealed that, while Shank3B KO mice displayed normal neuronal morphology at one week postnatal, significant impairments in dendritic growth and synaptic activity emerged by two to three weeks. These findings highlight the critical developmental window during which Shank3 is essential for neuronal and synaptic maturation in the ACC.
Animals ; Nerve Tissue Proteins/metabolism* ; Mice, Knockout ; Dendrites/metabolism* ; Mice ; Synapses/metabolism* ; Gyrus Cinguli/metabolism* ; Male ; Mice, Inbred C57BL ; Autism Spectrum Disorder/genetics* ; Microfilament Proteins

Mutations in the cyclin-dependent kinase-like 5 gene (CDKL5) cause a severe neurodevelopmental disorder, yet the impact of truncating mutations remains unclear. Here, we introduce the Cdkl5^492stop mouse model, mimicking C-terminal truncating mutations in patients. 492stop/Y mice exhibit altered dendritic spine morphology and spontaneous seizure-like behaviors, alongside other behavioral deficits. After creating cell lines with various Cdkl5 truncating mutations, we found that these mutations are regulated by the nonsense-mediated RNA decay pathway. Most truncating mutations result in CDKL5 protein loss, leading to multiple disease phenotypes, and offering new insights into the pathogenesis of CDKL5 disorder.
Animals ; Disease Models, Animal ; Mice ; Protein Serine-Threonine Kinases/deficiency* ; Mutation/genetics* ; Epileptic Syndromes/genetics* ; Humans ; Dendritic Spines/pathology* ; Spasms, Infantile/genetics* ; Male ; Seizures/genetics* ; Mice, Inbred C57BL

Epilepsy is a chronic neurological disorder affecting ~65 million individuals worldwide. Abnormal synaptic plasticity is one of the most important pathological features of this condition. We investigated how ubiquitin-specific peptidase 47 (USP47) influences synaptic plasticity and its link to epilepsy. We found that USP47 enhanced excitatory postsynaptic transmission and increased the density of total dendritic spines and the proportion of mature dendritic spines. Furthermore, USP47 inhibited the degradation of the ubiquitinated α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) subunit glutamate receptor 1 (GluR1), which is associated with synaptic plasticity. In addition, elevated levels of USP47 were found in epileptic mice, and USP47 knockdown reduced the frequency and duration of seizure-like events and alleviated epileptic seizures. To summarize, we present a new mechanism whereby USP47 regulates excitatory postsynaptic plasticity through the inhibition of ubiquitinated GluR1 degradation. Modulating USP47 may offer a potential approach for controlling seizures and modifying disease progression in future therapeutic strategies.
Animals ; Receptors, AMPA/metabolism* ; Neuronal Plasticity/physiology* ; Seizures/physiopathology* ; Disease Models, Animal ; Mice, Inbred C57BL ; Mice ; Ubiquitin Thiolesterase/genetics* ; Male ; Excitatory Postsynaptic Potentials/physiology* ; Ubiquitination ; Dendritic Spines/metabolism* ; Hippocampus/metabolism*

The perception of motion is an important function of vision. Neural wiring diagrams for extracting directional information have been obtained by connectome reconstruction. Direction selectivity in Drosophila is thought to originate in T4/T5 neurons through integrating inputs with different temporal filtering properties. Through genetic screening based on synaptic distribution, we isolated a new type of TmY neuron, termed TmY-ds, that form reciprocal synaptic connections with T4/T5 neurons. Its neurites responded to grating motion along the four cardinal directions and showed a variety of direction selectivity. Intriguingly, its direction selectivity originated from temporal filtering neurons rather than T4/T5. Genetic silencing and activation experiments showed that TmY-ds neurons are functionally upstream of T4/T5. Our results suggest that direction selectivity is generated in a tripartite circuit formed among these three neurons-temporal filtering, TmY-ds, and T4/T5 neurons, in which TmY-ds plays a role in the enhancement of direction selectivity in T4/T5 neurons.
Animals ; Neurites ; Drosophila ; Neurons ; Connectome

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*

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*

The back-propagating action potential (bpAP) is crucial for neuronal signal integration and synaptic plasticity in dendritic trees. Its properties (velocity and amplitude) can be affected by dendritic morphology. Due to limited spatial resolution, it has been difficult to explore the specific propagation process of bpAPs along dendrites and examine the influence of dendritic morphology, such as the dendrite diameter and branching pattern, using patch-clamp recording. By taking advantage of Optopatch, an all-optical electrophysiological method, we made detailed recordings of the real-time propagation of bpAPs in dendritic trees. We found that the velocity of bpAPs was not uniform in a single dendrite, and the bpAP velocity differed among distinct dendrites of the same neuron. The velocity of a bpAP was positively correlated with the diameter of the dendrite on which it propagated. In addition, when bpAPs passed through a dendritic branch point, their velocity decreased significantly. Similar to velocity, the amplitude of bpAPs was also positively correlated with dendritic diameter, and the attenuation patterns of bpAPs differed among different dendrites. Simulation results from neuron models with different dendritic morphology corresponded well with the experimental results. These findings indicate that the dendritic diameter and branching pattern significantly influence the properties of bpAPs. The diversity among the bpAPs recorded in different neurons was mainly due to differences in dendritic morphology. These results may inspire the construction of neuronal models to predict the propagation of bpAPs in dendrites with enormous variation in morphology, to further illuminate the role of bpAPs in neuronal communication.
Action Potentials/physiology* ; Dendrites/physiology* ; Neurons/physiology* ; Neuronal Plasticity ; Pyramidal Cells/physiology*

Diffusion tensor imaging technology can provide information on the white matter of the brain, which can be used to explore changes in brain tissue structure, but it lacks the specific description of the microstructure information of brain tissue. The neurite orientation dispersion and density imaging make up for its shortcomings. But in order to accurately estimate the brain microstructure, a large number of diffusion gradients are needed, and the calculation is complex and time-consuming through maximum likelihood fitting. Therefore, this paper proposes a kind of microstructure parameters estimation method based on the proximal gradient network, which further avoids the classic fitting paradigm. The method can accurately estimate the parameters while reducing the number of diffusion gradients, and achieve the purpose of imaging quality better than the neurite orientation dispersion and density imaging model and accelerated microstructure imaging via convex optimization model.
Brain/diagnostic imaging* ; Diffusion Magnetic Resonance Imaging ; Diffusion Tensor Imaging ; Neurites ; White Matter

OBJECTIVE: To assess the value of Ploton silver staining and phalloidin-iFlour 488 staining in observation of the morphology of osteocyte dendrites of mice at different developmental stages. METHODS: The humerus and femurs were harvested from mice at 0 (P0), 5 (P5), 15 (P15), 21 (P21), 28 (P28), and 35 days (P35) after birth to prepare cryo-sections and paraffin sections. HE staining of P35 mouse femur sections served as a reference for observing osteocytes in the trabecular bone and cortical bone. The humeral sections at different developmental stages were stained with Ploton silver staining to observe the morphology of osteocytes and canaliculi, and the canalicular lengths in the cortical and trabecular bones of the humerus of the mice in each developmental stage were recorded. The cryo-sections of the humerus from P10 and P15 mice were stained with phalloidin iFlour-488 to observe the morphology of osteocytes and measurement of the length of osteocyte dendrites in the cortical bone. RESULTS: In the trabecular bone of the humerus of P0-P15 mice, Ploton silver staining only visualized the outline of the osteocytes, and the morphology of the canaliculi was poorly defined. In P21 or older mice, Ploton silver staining revealed the morphology of the trabecular bone osteocytes and the canaliculi, which were neatly arranged and whose lengths increased significantly with age (P21 CONCLUSIONS Mouse osteocyte dendrites elongate progressively and their arrangement gradually becomes regular with age. Ploton silver staining can clearly visualize the morphology of the osteocytes and the canaliculi in adult mice but not in mice in early stages of development. Phalloidin iFlour-488 staining for labeling the cytoskeleton can be applied for mouse osteocytes at all developmental stages and allows morphological observation of mouse osteocytes in early developmental stages.
Animals ; Bone and Bones ; Dendrites ; Mice ; Osteocytes ; Phalloidine ; Silver Staining