Spines Of Hippocampal Neurons Research Articles

MiR-351-5p regulation of CPEB3 affecting aluminium-induced learning and memory impairment in SD rats

Aluminium exposure has been found to impair learning and memory abilities; however, the underlying molecular mechanisms remain unclear. In this study we conducted a double luciferase reporter assay to determine whether miR-351-5p regulates cytoplasmic polyadenylation element binding protein (CPEB) 3 mRNA. To this end, we overexpressed and inhibited miR-351-5p via stereotaxic microinjections of adeno-associated virus (AAV) into the hippocampus of Sprague Dawley rats in a sub-chronic aluminium exposure model to examine learning and memory ability using Morris water maze. Ultrastructural electron microscopy and Golgi staining were used to examine morphological changes in hippocampal neurons. In addition, we examined the levels of synaptic plasticity-related proteins (PRPs) and CPEB3 to determine the involvement of the miR-351-5P/CPEB3/PRPs pathway in aluminium neurotoxicity. Sub-chronic aluminium exposure reduced the spatial learning and memory ability of rats. Overexpression of AAV-miR-351-5P in the hippocampus aggravated the impairment of spatial learning and memory abilities of aluminium-treated rats, whereas inhibition of AAV-miR-351-5p expression alleviated it. Western blotting suggested that sub-chronic aluminium exposure increased miR-351-5p levels and reduced the expression of CPEB3 and PRPs in the hippocampus. Treatment with an AAV-miR-351-5p inhibitor partially recovered CPEB3 and PRPs. Double luciferase reporter assay results showed that CPEB3 was a direct target of miR-351-5p, while electron microscopy suggested that aluminium could damage mitochondria and synapses in the CA1 of the hippocampus. Golgi staining results indicated that aluminium could reduce the number of dendritic spines in hippocampal neurons. Inhibition of miR-351-5p restored the synaptic structure and growth of dendritic spines in the hippocampus. The involvement of the miR-351-5P/CPEB3/RPPs pathway in aluminium neurotoxicity was confirmed. Our findings suggest that inhibition of miR-351-5p can alleviate learning and memory impairments by increasing CPEB3 and PRPs.

Space-Like Irradiation Exacerbated Cognitive Deficits and Amyloid Pathology in CRND8 Mouse Model of Alzheimer's Disease.

Space radiation was linked to neurological damage and behavioral deficits which raised concerns of increased degenerative risk on the brain and development of Alzheimer's disease following space travel. In this study, we investigated the effects of irradiation by 56Fe and 28Si in CRND8 mice, an Alzheimer's disease mouse model. Six-month-old CRND8 mice were exposed to whole body irradiation by 56Fe and 28Si at 0.5 Gy and 2 Gy doses. Behavior tests were administered 1-month to 3-months post-irradiation. Amyloid deposition and other pathological changes were analyzed 3-months and/or 6-months post-irradiation. The Novel Object Recognition test showed some decline in 8-month-old mice compared to non-irradiated CRND8 mice. Male mice also showed a loss of freezing behavior in the fear conditioning contextual test following irradiation. Golgi staining revealed a loss of spines in hippocampal neurons after irradiation. Total amyloid immunohistochemistry showed a robust increase in 3-months post-irradiation 56Fe groups which became normalized to non-irradiated group by 6-months post-irradiation. However, 2 Gy 28Si caused a trend towards increased plaque load at 3-months post-irradiation which became significant at 6-months post irradiation only in male CRND8 mice. While 0.5 Gy Fe did not induce obvious changes in the total number of iba-1 positive microglia, more hippocampal microglia were found to express PCNA after 0.5 Gy Fe treatment, suggesting potential involvement of microglial dysfunction. Overall, our study provides new evidence of gender-specific and ion-dependent effects of space radiation on cognition and amyloid pathology in AD models.

Glycogen phosphorylase inhibition improves cognitive function of aged mice.

Inhibition of glycogen breakdown blocks memory formation in young animals, but it stimulates the maintenance of the long-term potentiation, a cellular mechanism of memory formation, in hippocampal slices of old animals. Here, we report that a 2-week treatment with glycogen phosphorylase inhibitor BAY U6751 alleviated memory deficits and stimulated neuroplasticity in old mice. Using the 2-Novel Object Recognition and Novel Object Location tests, we discovered that the prolonged intraperitoneal administration of BAY U6751 improved memory formation in old mice. This was accompanied by changes in morphology of dendritic spines in hippocampal neurons, and by "rejuvenation" of hippocampal proteome. In contrast, in young animals, inhibition of glycogen degradation impaired memory formation; however, as in old mice, it did not alter significantly the morphology and density of cortical dendritic spines. Our findings provide evidence that prolonged inhibition of glycogen phosphorolysis improves memory formation of old animals. This could lead to the development of new strategies for treatment of age-related memory deficits.

Arc-Mediated Synaptic Plasticity Regulates Cognitive Function in a Migraine Mouse Model

Previous clinical and basic studies have shown that migraine is associated with cognitive impairment, anxiety, and depression. It severely affects the quality of life. In this study, C57BL/6 mice were randomly divided into four groups: IS group, IS+M group, and IS+S group with repeated application of dural inflammatory soup (IS) stimulation to establish a migraine model, followed by PBS, memantine, and sumatriptan interventions, respectively; the blank control group underwent the same treatment procedure but with PBS instead of IS and intervention drugs. The cognitive function of the mice was used as the main outcome indicator. After application of the IS, mice showed reduced pain threshold for mechanical stimulation, decreased learning memory capacity, attention deficit, a reduced number of dendritic spines in hippocampal neurons, and altered synaptic ultrastructure. The cognitive function indexes of mice in the IS+M group recovered with changes in Arc protein expression to a level not statistically different from that of the Control group, while the IS and IS+S groups remained at lower levels. The present results suggest that Arc-mediated synaptic plasticity may be an essential mechanism of cognitive dysfunction in migraine.

In Vitro Neurotrophic Properties and Structural Characterization of a New Polysaccharide LTC-1 from Pyrola corbieri Levl (Luticao).

Pyrola corbieri Levl has been used to strengthen bones and nourish the kidney (the kidney governs the bone and is beneficial to the brain) by the local Miao people in China. However, the functional components and neurotrophic activity have not been reported. A new acidic homogeneous heteropolysaccharide named LTC-1 was obtained and characterized by periodate oxidation, Smith degradation, partial acid hydrolysis, GC-MS spectrometry, methylation analysis, and Fourier transform infrared spectroscopy, and its molecular weight was 3239 Da. The content of mannuronic acid (Man A) in LTC-1 was 46%, and the neutral sugar was composed of L-rhamnose (L-Rha), L-arabinose (L-Ara), D-xylose (D-Xyl), D-mannose (D-Man), D-glucose (D-Glc) and D-galactose (D-Gal) with a molar ratio of 1.00:3.63:0.86:1.30:6.97:1.30. The main chain of LTC-1 was composed of Glc, Gal, Man, Man A and the branched chain Ara, Glc, Gal. The terminal residues were composed of Glc and Gal. The main chain and branched chains were linked by (1→5)-linked-Ara, (1→3)-linked-Glc, (1→4)-linked-Glc, (1→6)-linked-Glc, (1→3)-linked-Gal, (1→6)-linked-Gal, (1→3, 6)-linked-Man and ManA. Meanwhile, neurotrophic activity was evaluated through PC12 and primary hippocampal neuronal cell models. LTC-1 exhibited neurotrophic activity in a concentration-dependent manner, which significantly induced the differentiation of PC12 cells, promoted the neurite outgrowth of PC12 cells, enhanced the formation of the web architecture of dendrites, and increased the density of dendritic spines in hippocampal neurons and the expression of PSD-95. These results displayed significant neurotrophic factor-like activity of LTC-1, which suggests that LTC-1 is a potential treatment option for neurodegenerative diseases.

Overexpressing lnc240 Rescues Learning and Memory Dysfunction in Hepatic Encephalopathy Through miR-1264-5p/MEF2C Axis.

Hepatic encephalopathy (HE) is a nervous system disease caused by severe liver diseases and different degrees of learning and memory dysfunction. Long non-coding RNA (lncRNA) is highly expressed in the brain and plays important roles in central nervous system diseases like Alzheimer's disease. In the present work, we found that the expression of lnc240 in the hippocampus of HE mice was significantly downregulated, but its pathogenesis in HE has not been clarified. This study aimed to explore the effects of lnc240 on the cognitive function of HE. The expression of lnc240, miR-1264-5p, and MEF2C was analyzed with RNA-seq and further determined by qRT-PCR in HE mouse. Double luciferase reporter gene testing confirmed the relationship between lnc240, MEF2C, and miR-1264-5p. The functional role of lnc240 and MEF2C in vitro and in vivo was evaluated by qRT-PCR, western blot analysis, immunofluorescence staining, Golgi staining, electrophysiology, and Morris water maze. The expression of lnc240 was decreased in HE mice. The overexpression of lnc240 could significantly downregulate miR-1264-5p and upregulate MEF2C, also increasing the amplitude and frequency of mEPSC in primary cultured hippocampal neurons. The overexpression of miR-1264-5p reversed the effect of lnc240 on MEF2C. Moreover, in vivo experiments have shown that the overexpression of lnc240 could improve HE mice's spatial learning and memory functions. Golgi staining suggested that overexpression of lnc240 could increase the density and maturity of dendritic spines in hippocampal neurons of HE mice. Lnc240 can regulate the expression of MEF2C through miR-1264-5p and regulate the synaptic plasticity of hippocampal neurons, thereby saving the learning and memory dysfunction in HE mice, suggesting that lnc240 might be a potential therapeutic target for the treatment of HE.

The role of α-tubulin tyrosination in controlling the structure and function of hippocampal neurons.

Microtubules (MTs) are central components of the neuronal cytoskeleton and play a critical role in CNS integrity, function, and plasticity. Neuronal MTs are diverse due to extensive post-translational modifications (PTMs), particularly detyrosination/tyrosination, in which the C-terminal tyrosine of α-tubulin is cyclically removed by a carboxypeptidase and reattached by a tubulin-tyrosine ligase (TTL). The detyrosination/tyrosination cycle of MTs has been shown to be an important regulator of MT dynamics in neurons. TTL-null mice exhibit impaired neuronal organization and die immediately after birth, indicating TTL function is vital to the CNS. However, the detailed cellular role of TTL during development and in the adult brain remains elusive. Here, we demonstrate that conditional deletion of TTL in the neocortex and hippocampus during network development results in a pathophysiological phenotype defined by incomplete development of the corpus callosum and anterior commissures due to axonal growth arrest. TTL loss was also associated with a deficit in spatial learning, impaired synaptic plasticity, and reduced number of spines in hippocampal neurons, suggesting that TTL also plays a critical role in hippocampal network development. TTL deletion after postnatal development, specifically in the hippocampus and in cultured hippocampal neurons, led to a loss of spines and impaired spine structural plasticity. This indicates a novel and important function of TTL for synaptic plasticity in the adult brain. In conclusion, this study reveals the importance of α-tubulin tyrosination, which defines the dynamics of MTs, in controlling proper network formation and suggests TTL-mediated tyrosination as a new key determinant of synaptic plasticity in the adult brain.

Ca2+ signaling in postsynaptic neurons: Neuroplastin-65 regulates the interplay between plasma membrane Ca2+ ATPases and ionotropic glutamate receptors

Upon postsynaptic glutamate receptor activation, the cytosolic Ca2+ concentration rises and initiates signaling and plasticity in spines. The plasma membrane Ca2+ ATPase (PMCA) is a major player to limit the duration of cytosolic Ca2+ signals. It forms complexes with the glycoprotein neuroplastin (Np) isoforms Np55 and Np65 and functionally interplays with N-methyl-D-aspartate (NMDA)-type ionotropic glutamate receptors (iGluNRs). Moreover, binding of the Np65-specific extracellular domain to Ca2+-permeable GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-type ionotropic glutamate receptors (iGluA1Rs) was found to be required for long-term potentiation (LTP). However, the link between PMCA and iGluRs function to regulate cytosolic Ca2+ signals remained unclear. Here, we report that Np65 coordinates PMCA and iGluRs’ functions to modulate the duration and amplitude of cytosolic Ca2+ transients in dendrites and spines of hippocampal neurons. Using live-cell Ca2+ imaging, acute pharmacological treatments, and GCaMP5G-expressing hippocampal neurons, we discovered that endogenous or Np65-promoted PMCA activity contributes to the restoration of basal Ca2+ levels and that this effect is dependent on iGluR activation. Super-resolution STED and confocal microscopy revealed that electrical stimulation increases the abundance of synaptic neuroplastin-PMCA complexes depending on iGluR activation and that low-rate overexpression of Np65 doubled PMCA levels and decreased cell surface levels of GluN2A and GluA1 in dendrites and Shank2-positive glutamatergic synapses. In neuroplastin-deficient hippocampi, we observed reduced PMCA and unchanged GluN2B levels, while GluN2A and GluA1 levels were imbalanced. Our electrophysiological data from hippocampal slices argues for an essential interplay of PMCA with GluN2A- but not with GluN2B-containing receptors upon induction of synaptic plasticity. Accordingly, we conclude that Np65 may interconnect PMCA with core players of glutamatergic neurotransmission to fine-tune the Ca2+ signal regulation in basal synaptic function and plasticity.

Ginkgo Biloba Extract Ameliorates Scopolamine-induced Memory Deficits via Rescuing Synaptic Damage

ObjectiveAlzheimer’s disease (AD) is the most common age-related neurodegenerative disorder. Emerging evidence suggests that synaptic dysfunction is associated with the onset and progression of AD. Interestingly, Ginkgo biloba extract (EGb) is one of the most frequently investigated herbal medicines for enhancing cognition and alleviating neurodegenerative dementia. This study aimed to investigate the effect and the mechanism of EGb on AD-like synaptic disorders.MethodsScopolamine (SCO)-induced rats were used to mimic AD-like memory deficits. Morris water maze test and fear conditioning test were conducted to evaluate the memory status of rats in response to different treatments. Then, the synapse alterations were assessed by Golgi staining, and Western blotting was conducted to assess the protein expression of PSD95, GluN2B, synapsin-1, and synaptophysin. Reverse transcription quantitative polymerase chain reaction was applied to detect the mRNA expression of PSD95 and the levels of miR-1-3p/miR-206-3p.ResultsEGb supplement alleviated the learning and memory deficits induced by SCO in behavioral experiments. Moreover, EGb treatment attenuated synaptic damage elicited by SCO, manifested as increased dendritic spine density and the proportion of mushroom-type spines in hippocampal neurons. Further investigation indicated that EGb rescued the expression of synaptic-related proteins, especially PSD95, and decreased the levels of miR-1-3p/miR-206-3p in the rat hippocampus.ConclusionThe application of EGb effectively treats SCO-induced memory impairments probably by suppressing miR-1-3p/miR-206-3p and elevating the expression of PSD95.

Overexpression of the dystrophins Dp40 and Dp40L170P modifies neurite outgrowth and the protein expression profile of PC12 cells

Dp40 is ubiquitously expressed including the central nervous system. In addition to being present in the nucleus, membrane, and cytoplasm, Dp40 is detected in neurites and postsynaptic spines in hippocampal neurons. Although Dp40 is expressed from the same promoter as Dp71, its role in the cognitive impairment present in Duchenne muscular dystrophy patients is still unknown. Here, we studied the effects of overexpression of Dp40 and Dp40L170P during the neuronal differentiation of PC12 Tet-On cells. We found that Dp40 overexpression increased the percentage of PC12 cells with neurites and neurite length, while Dp40L170P overexpression decreased them compared to Dp40 overexpression. Two-dimensional gel electrophoresis analysis showed that the protein expression profile was modified in nerve growth factor-differentiated PC12-Dp40L170P cells compared to that of the control cells (PC12 Tet-On). The proteins α-internexin and S100a6, involved in cytoskeletal structure, were upregulated. The expression of vesicle-associated membrane proteins increased in differentiated PC12-Dp40 cells, in contrast to PC12-Dp40L170P cells, while neurofilament light-chain was decreased in both differentiated cells. These results suggest that Dp40 has an important role in the neuronal differentiation of PC12 cells through the regulation of proteins involved in neurofilaments and exocytosis of synaptic vesicles, functions that might be affected in PC12-Dp40L170P.

Sleep Deprivation Aggravates Cognitive Impairment by the Alteration of Hippocampal Neuronal Activity and the Density of Dendritic Spine in Isoflurane-Exposed Mice.

Isoflurane contributes to cognitive deficits when used as a general anesthetic, and so does sleep deprivation (SD). Patients usually suffer from insomnia before an operation due to anxiety, fear, and other factors. It remains unclear whether preoperative SD exacerbates cognitive impairment induced by isoflurane. In this study, we observed the effects of pretreated 24-h SD in adult isoflurane-exposed mice on the cognitive behaviors, the Ca2+ signals of dorsal hippocampal CA1 (dCA1) neurons in vivo with fiber photometry, and the density of dendritic spines in hippocampal neurons. Our results showed that in cognitive behavior tasks, short-term memory damages were more severe with SD followed by isoflurane exposure than that with SD or isoflurane exposure separately, and interestingly, severe long-term memory deficits were induced only by SD followed by isoflurane exposure. Only the treatment of SD followed by isoflurane exposure could reversibly decrease the amplitude of Ca2+ signals when mice were freely moving and increase the duration of Ca2+ signals during the long-term memory behavior test. The density of dendritic spines with both SD and isoflurane exposure was lower than that with SD alone. This study suggests that SD should be avoided preoperatively in patients undergoing elective surgery under isoflurane anesthesia.

The Long Isoform of Intersectin-1 Has a Role in Learning and Memory.

Down syndrome is caused by partial or total trisomy of chromosome 21 and is characterized by intellectual disability and other disorders. Although it is difficult to determine which of the genes over-expressed on the supernumerary chromosome contribute to a specific abnormality, one approach is to study each gene in isolation. This can be accomplished either by using an over-expression model to study increased gene dosage or a gene-deficiency model to study the biological function of the gene. Here, we extend our examination of the function of the chromosome 21 gene, ITSN1. We used mice in which the long isoform of intersectin-1 was knocked out (ITSN1-LKO) to understand how a lack of the long isoform of ITSN1 affects brain function. We examined cognitive and locomotor behavior as well as long term potentiation (LTP) and the mitogen-activated protein kinase (MAPK) and 3′-kinase-C2β-AKT (AKT) cell signaling pathways. We also examined the density of dendritic spines on hippocampal pyramidal neurons. We observed that ITSN1-LKO mice had deficits in learning and long term spatial memory. They also exhibited impaired LTP, and no changes in the levels of the phosphorylated extracellular signal-regulated kinase (ERK) 1/2. The amount of phosphorylated AKT was reduced in the ITSN1-LKO hippocampus and there was a decrease in the number of apical dendritic spines in hippocampal neurons. Our data suggest that the long isoform of ITSN1 plays a part in normal learning and memory.

Benzothiazole amphiphiles promote RasGRF1-associated dendritic spine formation in human stem cell-derived neurons.

Synaptic dysfunction has been implicated as an early cause of cognitive decline in neurodegenerative diseases (NDDs) such as Alzheimer’s disease (AD). Methods to slow down or reverse the loss of functional synapses, therefore, represent a promising avenue to explore for treating NDDs. We have previously reported the development of a class of benzothiazole amphiphiles (BAMs) that exhibited the capability to improve memory and learning both in wild‐type mice and in an AD rodent model, putatively through promoting RasGRF1‐associated formation of dendritic spines in hippocampal neurons. While these results represent a good first step in exploring a new approach to treating NDDs, the capability of these compounds to increase spine density has not been previously examined in a human neuronal model. Here, we found that neurons derived from differentiated human induced pluripotent stem cells exhibited both an increase in RasGRF1 expression and a phenotypic increase in the density of postsynaptic density protein 95‐positive puncta (which we use to provide an estimate of dendritic spine density) in BAM‐treated vs. control neurons. These results demonstrate that the previously observed spinogenic effects of BAMs in rodent neurons can be recapitulated in a human neuronal model, which further supports the potential utility of BAM agents for treating human diseases associated with spine deficits such as AD or other NDDs.

Loss of drebrin from dendritic spines in hippocampal neurons from Alzheimer's disease model mouse

Loss of drebrin from dendritic spines in hippocampal neurons from Alzheimer's disease model mouse

Rac1 and Rac3 GTPases differently influence the morphological maturation of dendritic spines in hippocampal neurons.

The Rac1 and Rac3 GTPases are co-expressed in the developing nervous system, where they are involved in different aspects of neuronal development, including the formation of synapses. The deletion of both Rac genes determines a stronger reduction of dendritic spines in vitro compared to the knockout of either gene, indicating that Rac1 and Rac3 play a synergistic role in the formation of these structures. Here, we have addressed the role of each GTPase in the formation of dendritic spines by overexpressing either Rac1 or Rac3 in wildtype neurons, or by re-expressing either GTPase in double knockout hippocampal cultures. We show that the Rac3 protein is expressed with Rac1 in developing hippocampal neurons. Overexpression of either GTPase in WT neurons increases the density of dendritic spines, suggesting the involvement of both GTPases in their formation. We also found that the re-expression of either Rac1 or Rac3 in double knockout neurons is sufficient to restore spinogenesis. Rac1 is significantly more efficient than Rac3 in restoring the formation of spines. On the other hand the quantitative analysis in neurons overexpressing or re-expressing either GTPase shows that Rac3 induces a more pronounced increase in the size of the spines compared to Rac1. These enlarged spines form morphological synapses identified by the juxtaposition of postsynaptic and presynaptic markers. Thus, while Rac1 appears more efficient in inducing the formation of mature spines, Rac3 is more efficient in promoting their enlargement. Our study highlights specific roles of Rac1 and Rac3, which may be functionally relevant also to synaptic plasticity.

Oleanolic acid decreases SGK1 in the hippocampus in corticosterone-induced mice

Our previous study has demonstrated that oleanolic acid produced an antidepressant-like effect in mice exposed to chronic stress. Considering that serine/threonine-protein kinase 1 (SGK1) is involved in stress response, the present study aimed to evaluate the involvement of SGK1 in the antidepressant-like effects of oleanolic acid in depression-like mice induced by long term corticosterone (CORT) injection. Behaviors, SGK1, brain-derived neurotrophic factor (BDNF) and its downstream targets were assessed after administration with oleanolic for three weeks. The results indicated that oleanolic acid increased the sucrose preference and decreased the immobility time. In addition, oleanolic acid decreased SGK1 and activated BDNF-AKT/mTOR signaling in the hippocampus of CORT-induced animals. However, we found that GSK650394, an inhibitor of SGK1 did not exert any effects on the behaviors, GR levels and BDNF signaling. The number of spines in hippocampal neurons was not changed by GSK650394 as well. Taken together, this study demonstrated that oleanolic acid produced the antidepressant-like effects, which might be related to the down-regulation of SGK1. However, inhibition of SGK1 directly lacks the effects in the treatment of depression.

Glucagon-Like Peptide-2 Receptor is Involved in Spatial Cognitive Dysfunction in Rats After Chronic Cerebral Hypoperfusion.

Chronic cerebral hypoperfusion (CCH) affects the aging population and especially patients with neurodegenerative diseases, such as Alzheimer's disease or Parkinson's disease. CCH is closely related to the cognitive dysfunction in these diseases. Glucagon-like peptide-2 receptor (GLP2R) mRNA and protein are highly expressed in the gut and in hippocampal neurons. This receptor is involved in the regulation of food intake and the control of energy balance and glucose homeostasis. The present study employed behavioral techniques, electrophysiology, western blotting, immunohistochemistry, quantitative real time polymerase chain reaction (qRT-PCR), and Golgi staining to investigate whether the expression of GLP2R changes after CCH and whether GLP2R is involved in cognitive impairment caused by CCH. Our findings show that CCH significantly decreased hippocampal GLP2R mRNA and protein levels. GLP2R upregulation could prevent CCH-induced cognitive impairment. It also improved the CCH-induced impairment of long-term potentiation and long-term depression. Additionally, GLP2R modulated after CCH the AKT-mTOR-p70S6K pathway in the hippocampus. Moreover, an upregulation of the GLP2R increased the neurogenesis in the dentate gyrus, neuronal activity, and density of dendritic spines and mushroom spines in hippocampal neurons. Our findings reveal the involvement of GLP2R via a modulation of the AKT-mTOR-p70S6K pathway in the mechanisms underlying CCH-induced impairments of spatial learning and memory. We suggest that the GLP2R and the AKT-mTOR-p70S6K pathway in the hippocampus are promising targets to treat cognition deficits in CCH.

Ketamine destabilizes growth of dendritic spines in developing hippocampal neurons invitro via a Rho‑dependent mechanism.

The safety of anesthetics on the developing brain has caused concern. Ketamine, an N-methyl-D-aspartate receptor antagonist, is widely used as a general pediatric anesthetic. Recent studies suggested that ketamine alters the plasticity of dendritic spines in the developing brain and may be an important contributing factor to learning and cognitive impairment. However, the underlying molecular mechanism remains poorly understood. Therefore, the aim of the present study was to investigate the effect of ketamine on the plasticity of dendritic spines in cultured hippocampal neurons and the potential underlying mechanisms. After 5 days in vitro, rat hippocampal neurons were exposed to different concentrations (100, 300 and 500 µM) of ketamine for 6 h. Ketamine decreased the number and length of dendritic spines in a dose-dependent manner. Ketamine at a concentration of 300 µM caused an upregulation of transforming protein RhoA (RhoA) and Rho-associated kinase (ROCK) protein. These effects were inhibited by the ROCK inhibitor Y27632. These results suggested that ketamine induces loss and shortening of dendritic spines in hippocampal neurons via activation of the RhoA/ROCK signaling pathway.

Antidepression action of BDNF requires and is mimicked by Gαi1/3 expression in the hippocampus

Stress-related alterations in brain-derived neurotrophic factor (BDNF) expression, a neurotrophin that plays a key role in synaptic plasticity, are believed to contribute to the pathophysiology of depression. Here, we show that in a chronic mild stress (CMS) model of depression the Gαi1 and Gαi3 subunits of heterotrimeric G proteins are down-regulated in the hippocampus, a key limbic structure associated with major depressive disorder. We provide evidence that Gαi1 and Gαi3 (Gαi1/3) are required for the activation of TrkB downstream signaling pathways. In mouse embryonic fibroblasts (MEFs) and CNS neurons, Gαi1/3 knockdown inhibited BDNF-induced tropomyosin-related kinase B (TrkB) endocytosis, adaptor protein activation, and Akt-mTORC1 and Erk-MAPK signaling. Functional studies show that Gαi1 and Gαi3 knockdown decreases the number of dendrites and dendritic spines in hippocampal neurons. In vivo, hippocampal Gαi1/3 knockdown after bilateral microinjection of lentiviral constructs containing Gαi1 and Gαi3 shRNA elicited depressive behaviors. Critically, exogenous expression of Gαi3 in the hippocampus reversed depressive behaviors in CMS mice. Similar results were observed in Gαi1/Gαi3 double-knockout mice, which exhibited severe depressive behaviors. These results demonstrate that heterotrimeric Gαi1 and Gαi3 proteins are essential for TrkB signaling and that disruption of Gαi1 or Gαi3 function could contribute to depressive behaviors.

Stim2-Eb3 Association and Morphology of Dendritic Spines in Hippocampal Neurons

Mushroom spines form strong synaptic contacts and are essential for memory storage. We have previously demonstrated that neuronal store-operated calcium entry (nSOC) in hippocampal neurons is regulated by STIM2 protein. This pathway plays a key role in stability of mushroom spines and is compromised in different mice models of Alzheimer’s disease (AD). Actin was thought to be the sole cytoskeleton compartment presented in dendritic spines, however, recent studies demonstrated that dynamic microtubules with EB3 capped plus-ends transiently enter spines. We showed that STIM2 forms an endoplasmic reticulum (ER) Ca2+ -dependent complex with EB3 via Ser-x-Ile-Pro aminoacid motif and that disruption of STIM2-EB3 interaction resulted in loss of mushroom spines in hippocampal neurons. Overexpression of EB3 causes increase of mushroom spines fraction and is able to restore their deficiency in hippocampal neurons obtained from PS1-M146V-KI AD mouse model. STIM2 overexpression failed to restore mushroom dendritic spines after EB3 knockdown, while in contrast EB3 overexpression rescued loss of mushroom spines resulting from STIM2 depletion. We propose that EB3 is involved in regulation of dendritic spines morphology, in part due to its association with STIM2, and that modulation of EB3 expression is a potential way to overcome synaptic loss during AD.