Postsynaptic Protein Research Articles

Memory Improving Effects in Social-Isolated Aged Rats by Seaweed Glycoproteins

Social isolation and loneliness that could induce cognitive decline are serious public health problems in elderly. The trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) is a critical process in long-term potentiation of synapses that is necessary for memory formation. We previously found that glycoproteins of an edible seaweed Capsosiphon fulvescens (C. fulvescens) prevent aging-induced cognitive impairment via regulation of brain-derived neurotrophic factor in hippocampus. This study was to investigate whether chronic administration of hydrophilic glycoproteins of C. fulvescens (Cf-hGP) prevent cognitive dysfunctions caused by social isolation in aged rats and this effect is regulated by post synaptic density protein 95 (PSD-95)-mediated AMPAR trafficking and the glycosylation of Cf-hGP. Social isolation for four weeks decreased phosphorylation of extracellular signal-regulated protein kinase 1/2 (ERK1/2), PSD-95 (Ser295) and AMPAR GluR1 (Ser845) and increased expression of metabotropic glutamate receptor 5 (mGluR5) in synaptosome of the dorsal hippocampus, which was attenuated by chronic Cf-hGP treatment (15 mg/kg/day, 4 weeks). Blockade of mGluR5 abolished decrease in ERK1/2-mediated phosphorylation of PSD-95 (Ser295) and GluR1 (Ser845) in the socially isolated rats. In particular, chronic Cf-hGP treatment enhanced binding of ERK1/2 to PSD-95 and upregulated the surface movement of AMPAR GluR1 in the dorsal hippocampus. In addition, Cf-hGP treatment prevented spatial memory impairment caused by the social isolation, which was attenuated by inhibition of ERK1/2 or deglycosylation of Cf-hGP. These findings suggest that Cf-hGP-induced clustering of ERK1/2-mediated PSD-95 in the dorsal hippocampus improves memory formation in socially isolated aged rats and protein glycosylation contributes to enhancing the Cf-hGP effect.

Reduced expression of the cell intrinsic clock protein, Bmal1, in myeloid cells accelerates cognitive decline and alters microglial function in aging mice

The circadian clock system plays a fundamental role in the temporal coordination of the external environment with internal physiological processes. These processes include the sleep/wake cycle, feeding, liver function, kidney function, core body temperature, including insulin release and heart rate, and immune function. With aging, the ability to rhythmically regulate these processes progressively declines. Furthermore, aging is associated with a chronic inflammation that is linked to multiple age-associated diseases, including cognitive decline. Several studies have demonstrated that deletion of Bmal1, a transcription factor and principal driver of cell-autonomous circadian rhythms, accelerates the aging process, exhibiting a variety of age-related diseases including sarcopenia, cataracts, organ shrinkage, bone calcification, and impaired glucose metabolism. Here, we show that myeloid-lineage specific knockdown of Bmal1 increases systemic inflammation and accelerates cognitive decline in aging mice. This was associated with a paradoxical increase in IBA1 expression and number of microglia with less morphological complexity and reduced lysosomal CD68 in the CA1 hippocampal region. Looking at individual microglia showed less CD68 in the Bmal1-deficient microglia, suggesting lysosomal dysfunction. Furthermore, myeloid Bmal1 deletion led to significant increases in pre- and post-synaptic proteins in aged mice, as well as decreased levels of C1q, which is required for synaptic pruning, suggesting impaired synapse clearance. In summary, our data indicates that Bmal1 knock-down in myeloid-lineage cells is sufficient to accelerate cognitive decline by a mechanism that may involve impaired synapse removal.

A Novel Missense Variant in DLG2 /PSD‐93 Disrupts Protein Folding and Binding In Neurodevelopmental Disorders

The genetic basis of neurodevelopmental disorders (NDDs) such as autism spectrum disorder and intellectual disability is complex, involving multiple genes with mutations that can be rare, common, de novo or inherited. However, amidst this complexity, genomic studies continue to reveal a convergence of mutations in synaptic genes in NDDs. At excitatory synapses, in the vertebrate brain, the postsynaptic terminal contains highly conserved multiprotein complexes critical for synaptic signalling and behavior. These multiprotein complexes are organised by postsynaptic scaffold proteins, such as PSD-93 (encoded by the DLG2 gene). PSD-93 directly binds and anchors NMDA receptors to the membrane and assembles a downstream network of proteins to form the postsynaptic signalling machinery. We have identified a novel missense variant in DLG2 (R837N mutation in PSD-93) in a family with two brothers diagnosed with intellectual disability and autistic features. Here, we investigated the biological impact of this DLG2/PSD-93 mutation on protein structure and function. We show that the mouse PSD-93 R714N mutation (corresponding to human PSD-93 R837N) did not significantly alter PSD-93 protein expression nor density of PSD-93-positive synapses in mouse hippocampal neurons in vitro. However, protein structural analyses revealed that this mutation impairs PSD-93 protein folding and, as a consequence, decreases protein stability, and disrupts binding of PSD-93 to other key postsynaptic interactors. Our results show that the DLG2/PSD-93 R837N missense variant causes significant changes in the shape, stability and binding of the scaffold protein. This likely affects the structural capacity of PSD-93 to dynamically bind and reorganise the postsynaptic terminal and thus, potentially disrupts synaptic signalling and behaviour. Elucidating the functional impacts of pathogenic disease variants at the structural and molecular levels provide important biological insights into the dysfunctional mechanisms that underlie NDDs.

CaMKII activation persistently segregates postsynaptic proteins via liquid phase separation.

Transient information input to the brain leads to persistent changes in synaptic circuits, contributing to the formation of memory engrams. Pre- and postsynaptic structures undergo coordinated functional and structural changes during this process, but how such changes are achieved by their component molecules remains largely unknown. We found that activated CaMKII, a central player of synaptic plasticity, undergoes liquid-liquid phase separation with the NMDA-type glutamate receptor subunit GluN2B. Due to CaMKII autophosphorylation, the condensate stably persists even after Ca2+ is removed. The selective binding of activated CaMKII with GluN2B cosegregates AMPA receptors and the synaptic adhesion molecule neuroligin into a phase-in-phase assembly. In this way, Ca2+-induced liquid-liquid phase separation of CaMKII has the potential to act as an activity-dependent mechanism to crosslink postsynaptic proteins, which may serve as a platform for synaptic reorganization associated with synaptic plasticity.

Experiences affect social behaviors via altering neuronal morphology and oxytocin system

Life experiences, such as maternal deprivation (MD) and environment enrichment (EE), affect social behaviors in the adult. But, the underlying mechanism remains unclear. In the present study, we determined whether neonatal MD induces social deficits, whether postweaning EE restores the deficits, and their effects on neuron morphology and oxytocin (OT)-oxytocin receptor (OTR) system. We found that MD induced repetitive behavior and deficits in novel object recognition and sociability, and EE alleviated these deficits. MD decreased oxytocinergic neurons in the magnocellular hypothalamic paraventricular nucleus (mPVH), which was parallel to the increased OTR levels and dendritic branches of projection neurons in the basolateral amygdala (BLA). EE increased the OTR levels in the prelimbic cortex (PL) and the oxytocinergic neurons in the parvocellular PVH (vPVH), which were parallel to the increased dendritic branches of small pyramidal neurons in the PL and synaptic connections marked with synaptophysin and postsynaptic density protein 95 in the BLA and PL. Together, the results suggest that postweaning EE alleviates the social impairments induced by neonatal MD and OT-OTR system are experience-dependent and associated with social behaviors and neuron morphology.

Electrical synaptic transmission requires a postsynaptic scaffolding protein.

Electrical synaptic transmission relies on neuronal gap junctions containing channels constructed by Connexins. While at chemical synapses neurotransmitter-gated ion channels are critically supported by scaffolding proteins, it is unknown if channels at electrical synapses require similar scaffold support. Here, we investigated the functional relationship between neuronal Connexins and Zonula Occludens 1 (ZO1), an intracellular scaffolding protein localized to electrical synapses. Using model electrical synapses in zebrafish Mauthner cells, we demonstrated that ZO1 is required for robust synaptic Connexin localization, but Connexins are dispensable for ZO1 localization. Disrupting this hierarchical ZO1/Connexin relationship abolishes electrical transmission and disrupts Mauthner cell-initiated escape responses. We found that ZO1 is asymmetrically localized exclusively postsynaptically at neuronal contacts where it functions to assemble intercellular channels. Thus, forming functional neuronal gap junctions requires a postsynaptic scaffolding protein. The critical function of a scaffolding molecule reveals an unanticipated complexity of molecular and functional organization at electrical synapses.

The Speed of Allosteric Signaling Within a Single-Domain Protein.

While much is known about different allosteric regulation mechanisms, the nature of the allosteric signal and the time scale on which it propagates remains elusive. The PDZ3 domain from postsynaptic density-95 protein is a small protein domain with a terminal third α-helix, i.e., the α3-helix, which is known to be allosterically active. By cross-linking the allosteric helix with an azobenzene moiety, we obtained a photocontrollable PDZ3 variant. Photoswitching triggers its allosteric transition, resulting in a change in binding affinity of a peptide to the remote binding pocket. Using time-resolved infrared and UV/vis spectroscopy, we follow the allosteric signal transduction and reconstruct the timeline in which the allosteric signal propagates through the protein within 200 ns.

Expression Analysis of Zinc Transporters in Nervous Tissue Cells Reveals Neuronal and Synaptic Localization of ZIP4.

In the last years, research has shown that zinc ions play an essential role in the physiology of brain function. Zinc acts as a potent neuromodulatory agent and signaling ions, regulating healthy brain development and the function of both neurons and glial cells. Therefore, the concentration of zinc within the brain and its cells is tightly controlled. Zinc transporters are key regulators of (extra-) cellular zinc levels, and deregulation of zinc homeostasis and zinc transporters has been associated with neurodegenerative and neuropsychiatric disorders. However, to date, the presence of specific family members and their subcellular localization within brain cells have not been investigated in detail. Here, we analyzed the expression of all zinc transporters (ZnTs) and Irt-like proteins (ZIPs) in the rat brain. We further used primary rat neurons and rat astrocyte cell lines to differentiate between the expression found in neurons or astrocytes or both. We identified ZIP4 expressed in astrocytes but significantly more so in neurons, a finding that has not been reported previously. In neurons, ZIP4 is localized to synapses and found in a complex with major postsynaptic scaffold proteins of excitatory synapses. Synaptic ZIP4 reacts to short-term fluctuations in local zinc levels. We conclude that ZIP4 may have a so-far undescribed functional role at excitatory postsynapses.

Ketamine Induces Lasting Antidepressant Effects by Modulating the NMDAR/CaMKII-Mediated Synaptic Plasticity of the Hippocampal Dentate Gyrus in Depressive Stroke Model.

Background Ketamine has been shown to possess lasting antidepressant properties. However, studies of the mechanisms involved in its effects on poststroke depression are nonexistent. Methods To investigate these mechanisms, Sprague-Dawley rats were treated with a single local dose of ketamine after middle cerebral artery occlusion and chronic unpredicted mild stress. The effects on the hippocampal dentate gyrus were analyzed through assessment of the N-methyl-D-aspartate receptor/calcium/calmodulin-dependent protein kinase II (NMDAR/CaMKII) pathway, synaptic plasticity, and behavioral tests. Results Ketamine administration rapidly exerted significant and lasting improvements of depressive symptoms. The biochemical analysis showed rapid, selective upregulation and downregulation of the NMDAR2-β and NMDAR2-α subtypes as well as their downstream signaling proteins β-CaMKII and α-phosphorylation in the dentate gyrus, respectively. Furthermore, the colocalization analysis indicated a significant and selectively increased conjunction of β-CaMKII and postsynaptic density protein 95 (PSD95) coupled with a notable decrease in NMDAR2-β association with PSD95 after ketamine treatment. These changes translated into significant and extended synaptic plasticity in the dentate gyrus. Conclusions These findings not only suggest that ketamine represents a viable candidate for the treatment of poststroke depression but also that ketamine's lasting antidepressant effects might be achieved through modulation of NMDAR/CaMKII-induced synaptic plasticity in key brain regions.

Early postnatal allergic airway inflammation induces dystrophic microglia leading to excitatory postsynaptic surplus and autism-like behavior

Microglia play key roles in synaptic pruning, which primarily occurs from the postnatal period to adolescence. Synaptic pruning is essential for normal brain development and its impairment is implicated in neuropsychiatric developmental diseases such as autism spectrum disorders (ASD). Recent epidemiological surveys reported a strong link between ASD and atopic/allergic diseases. However, few studies have experimentally investigated the relationship between allergy and ASD-like manifestations, particularly in the early postnatal period, when allergic disorders occur frequently. Therefore, we aimed to characterize how allergic inflammation in the early postnatal period influences microglia and behavior using mouse models of short- and long-term airway allergy. Male mice were immunized by an intraperitoneal injection of aluminum hydroxide and ovalbumin (OVA) or phosphate-buffered saline (control) on postnatal days (P) 3, 7, and 11, followed by intranasal challenge with OVA or phosphate-buffered saline solution twice a week until P30 or P70. In the hippocampus, Iba-1-positive areas, the size of Iba-1-positive microglial cell bodies, and the ramification index of microglia by Sholl analysis were significantly smaller in the OVA group than in the control group on P30 and P70, although Iba-1-positive microglia numbers did not differ significantly between the two groups. In Iba-1-positive cells, postsynaptic density protein 95 (PSD95)-occupied areas and CD68-occupied areas were significantly decreased on P30 and P70, respectively, in the OVA group compared with the control group. Immunoblotting using hippocampal tissues demonstrated that amounts of PSD95, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor 2, and N-methyl-D-aspartate (NMDA) receptor 2B were significantly increased in the OVA group compared with the control group on P70, and a similar increasing trend for PSD95 was observed on P30. Neurogenesis was not significantly different between the two groups on P30 or P70 by doublecortin immunohistochemistry. The social preference index was significantly lower in the three chamber test and the number of buried marbles was significantly higher in the OVA group than in the control group on P70 but not on P30, whereas locomotion and anxiety were not different between the two groups. Compared with the control group, serum basal corticosterone levels were significantly elevated and hippocampal glucocorticoid receptor (GR) amounts and nuclear GR translocation in microglia, but not in neurons or astrocytes, were significantly decreased in the OVA group on P70 but not on P30. Gene set enrichment analysis of isolated microglia revealed that genes related to immune responses including Toll-like receptor signaling and chemokine signaling pathways, senescence, and glucocorticoid signaling were significantly upregulated in the OVA group compared with the control group on P30 and P70. These findings suggest that early postnatal allergic airway inflammation induces dystrophic microglia that exhibit defective synaptic pruning upon short- and long-term allergen exposure. Furthermore, long-term allergen exposure induced excitatory postsynaptic surplus and ASD-like behavior. Hypothalamo-pituitary-adrenal axis activation and the compensatory downregulation of microglial GR during long-term allergic airway inflammation may also facilitate these changes.

Neurogranin Regulates Adult-Born Olfactory Granule Cell Spine Density and Odor-Reward Associative Memory in Mice

Neurogranin (Ng) is a brain-specific postsynaptic protein, whose role in modulating Ca2+/calmodulin signaling in glutamatergic neurons has been linked to enhancement in synaptic plasticity and cognitive functions. Accordingly, Ng knock-out (Ng-ko) mice display hippocampal-dependent learning and memory impairments associated with a deficit in long-term potentiation induction. In the adult olfactory bulb (OB), Ng is expressed by a large population of GABAergic granule cells (GCs) that are continuously generated during adult life, undergo high synaptic remodeling in response to the sensory context, and play a key role in odor processing. However, the possible implication of Ng in OB plasticity and function is yet to be investigated. Here, we show that Ng expression in the OB is associated with the mature state of adult-born GCs, where its active-phosphorylated form is concentrated at post-synaptic sites. Constitutive loss of Ng in Ng-ko mice resulted in defective spine density in adult-born GCs, while their survival remained unaltered. Moreover, Ng-ko mice show an impaired odor-reward associative memory coupled with reduced expression of the activity-dependent transcription factor Zif268 in olfactory GCs. Overall, our data support a role for Ng in the molecular mechanisms underlying GC plasticity and the formation of olfactory associative memory.

Role of mTOR-Regulated Autophagy in Synaptic Plasticity Related Proteins Downregulation and the Reference Memory Deficits Induced by Anesthesia/Surgery in Aged Mice.

Postoperative cognitive dysfunction increases mortality and morbidity in perioperative patients and has become a major concern for patients and caregivers. Previous studies demonstrated that synaptic plasticity is closely related to cognitive function, anesthesia and surgery inhibit synaptic function. In central nervous system, autophagy is vital to synaptic plasticity, homeostasis of synapticproteins, synapse elimination, spine pruning, proper axon guidance, and when dysregulated, is associated with behavioral and memory functions disorders. The mammalian target of rapamycin (mTOR) negatively regulates the process of autophagy. This study aimed to explore whether rapamycin can ameliorate anesthesia/surgery-induced cognitive deficits by inhibiting mTOR, activating autophagy and rising synaptic plasticity-related proteins in the hippocampus. Aged C57BL/6J mice were used to establish POCD models with exploratory laparotomy under isoflurane anesthesia. The Morris Water Maze (MWM) was used to measure reference memory after anesthesia and surgery. The levels of mTOR phosphorylation (p-mTOR), Beclin-1 and LC3-II were examined on postoperative days 1, 3 and 7 by western blotting. The levels of synaptophysin (SYN) and postsynaptic density protein 95 (PSD-95) in the hippocampus were also examined by western blotting. Here we showed that anesthesia/surgery impaired reference memory and induced the activation of mTOR, decreased the expression of autophagy-related proteins such as Beclin-1 and LC3-II. A corresponding decline in the expression of neuronal/synaptic, plasticity-related proteins such as SYN and PSD-95 was also observed. Pretreating mice with rapamycin inhibited the activation of mTOR and restored autophagy function, also increased the expression of SYN and PSD-95. Furthermore, anesthesia/surgery-induced learning and memory deficits were also reversed by rapamycin pretreatment. In conclusion, anesthesia/surgery induced mTOR hyperactivation and autophagy impairments, and then reduced the levels of SYN and PSD-95 in the hippocampus. An mTOR inhibitor, rapamycin, ameliorated anesthesia/surgery-related cognitive impairments by inhibiting the mTOR activity, inducing activation of autophagy, enhancing SYN and PSD-95 expression.

Bi-Directional Protein-Protein Interactions Control Liquid-Liquid Phase Separation of PSD-95 and Its Interaction Partners

The organization of the postsynaptic density (PSD), a protein-dense semi-membraneless organelle, is mediated by numerous specific protein-protein interactions (PPIs) which constitute a functional post-synapse. Postsynaptic density protein 95 (PSD-95) interacts with a manifold of proteins, including the C-terminal of transmembrane AMPA receptor (AMAPR) regulatory proteins (TARPs). Here, we uncover the minimal essential peptide responsible for the stargazin (TARP-γ2) mediated liquid-liquid phase separation (LLPS) formation of PSD-95 and other key protein constituents of the PSD. Furthermore, we find that pharmacological inhibitors of PSD-95 can facilitate formation of LLPS. We found that in some cases LLPS formation is dependent on multivalent interactions while in other cases short peptides carrying a high charge are sufficient to promote LLPS in complex systems. This study offers a new perspective on PSD-95 interactions and their role in LLPS formation, while also considering the role of affinity over multivalency in LLPS systems.

Sensorimotor Perturbation Induces Late and Transient Molecular Synaptic Proteins Activation and Expression Changes.

Plasticity of the cerebral cortex following a modification of the sensorimotor experience takes place in several steps that can last from few hours to several months. Among the mechanisms involved in the dynamic modulation of the cerebral cortex in adults, it is commonly proposed that short-term plasticity reflects changes in synaptic connections. Here, we were interested in the time-course of synaptic plasticity taking place in the somatosensory primary cortex all along a 14-day period of sensorimotor perturbation (SMP), as well as during a recovery phase up to 24h. Activation and expression level of pre- (synapsin 1, synaptophysin, synaptotagmin 1) and postsynaptic (AMPA and NMDA receptors) proteins, postsynaptic density scaffold proteins (PSD-95 and Shank2), and cytoskeletal proteins (neurofilaments-L and M, β3-tubulin, synaptopodin, N-cadherin) were determined in cortical tissue enriched in synaptic proteins. During the SMP period, most changes were observed as soon as D7 in the presynaptic compartment and were followed, at D14, by changes in the postsynaptic compartment. These changes persisted at least until 24h of recovery. Proteins involved in synapse structure (scaffolding, adhesion, cytoskeletal) were mildly affected and almost exclusively at D14. We concluded that experience-dependent reorganization of somatotopic cortical maps is accompanied by changes in synaptic transmission with a very close time-course.

Neurodegeneration Induced by Anti-IgLON5 Antibodies Studied in Induced Pluripotent Stem Cell-Derived Human Neurons.

Anti-IgLON5 disease is a progressive neurological disorder associated with autoantibodies against a neuronal cell adhesion molecule, IgLON5. In human postmortem brain tissue, the neurodegeneration and accumulation of hyperphosphorylated tau (p-tau) are found. Whether IgLON5 antibodies induce neurodegeneration or neurodegeneration provokes an immune response causing inflammation and antibody formation remains to be elucidated. We investigated the effects of anti-IgLON5 antibodies on human neurons. Human neural stem cells were differentiated for 14–48 days and exposed from Days 9 to 14 (short-term) or Days 13 to 48 (long-term) to either (i) IgG from a patient with confirmed anti-IgLON5 antibodies or (ii) IgG from healthy controls. The electrical activity of neurons was quantified using multielectrode array assays. Cultures were immunostained for β-tubulin III and p-tau and counterstained with 4′,6-Diamidine-2′-phenylindole dihydrochloride (DAPI). To study the impact on synapses, cultures were also immunostained for the synaptic proteins postsynaptic density protein 95 (PSD95) and synaptophysin. A lactate dehydrogenase release assay and nuclei morphology analysis were used to assess cell viability. Cultures exposed to anti-IgLON5 antibodies showed reduced neuronal spike rate and synaptic protein content, and the proportion of neurons with degenerative appearance including p-tau (T205)-positive neurons was higher when compared to cultures exposed to control IgG. In addition, cell death was increased in cultures exposed to anti-IgLON5 IgG for 21 days. In conclusion, pathological anti-IgLON5 antibodies induce neurodegenerative changes and cell death in human neurons. This supports the hypothesis that autoantibodies may induce the neurodegenerative changes found in patients with anti-IgLON5-mediated disease. Furthermore, this study highlights the potential use of stem cell-based in vitro models for investigations of antibody-mediated diseases. As anti-IgLON5 disease is heterogeneous, more studies with different IgLON5 antibody samples tested on human neurons are needed.

Rapid eye movement sleep deprivation impairs neuronal plasticity and reduces hippocampal neuronal arborization in male albino rats: Noradrenaline is involved in the process.

Rapid eye movement sleep (REMS) favors brain development and memory, while it is decreased in neurodegenerative diseases. REMS deprivation (REMSD) affects several physiological processes including memory consolidation; however, its detailed mechanism(s) of action was unknown. REMS reduces, while REMSD elevates noradrenaline (NA) level in the brain; the latter induces several deficiencies and disorders, including changes in neuronal cytomorphology and apoptosis. Therefore, we proposed that REMS- and REMSD-associated modulation of NA level might affect neuronal plasticity and affect brain functions. Male albino rats were REMS deprived by flower-pot method for 6days, and its effects were compared with home cage and large platform controls as well as post-REMSD recovered and REMS-deprived prazosin (α1-adrenoceptor antagonist)-treated rats. We observed that REMSD reduced CA1 and CA3 neuronal dendritic length, branching, arborization, and spine density, while length of active zone and expressions of pre- as well as post-synaptic proteins were increased as compared to controls; interestingly, prazosin prevented most of the effects in vivo. Studies on primary culture of neurons from chick embryo brain confirmed that NA at lower concentration(s) induced neuronal branching and arborization, while higher doses were destructive. The findings support our contention that REMSD adversely affects neuronal plasticity, branching, and synaptic scaffold, which explain the underlying cytoarchitectural basis of REMSD-associated patho-physio-behavioral changes. Consolidation of findings of this study along with that of our previous reports suggest that the neuronal disintegration could be due to either withdrawal of direct protective and proliferative role of low dose of NA or indirect effect of high dose of NA or both.

Recordings from neuron-HEK cell cocultures reveal the determinants of miniature excitatory postsynaptic currents.

Spontaneous exocytosis of single synaptic vesicles generates miniature synaptic currents, which provide a window into the dynamic control of synaptic transmission. To resolve the impact of different factors on the dynamics and variability of synaptic transmission, we recorded miniature excitatory postsynaptic currents (mEPSCs) from cocultures of mouse hippocampal neurons with HEK cells expressing the postsynaptic proteins GluA2, neuroligin 1, PSD-95, and stargazin. Synapses between neurons and these heterologous cells have a molecularly defined postsynaptic apparatus, while the compact morphology of HEK cells eliminates the distorting effect of dendritic filtering. HEK cells in coculture produced mEPSCs with a higher frequency, larger amplitude, and more rapid rise and decay than neurons from the same culture. However, mEPSC area indicated that nerve terminals in synapses with both neurons and HEK cells release similar populations of vesicles. Modulation by the glutamate receptor ligand aniracetam revealed receptor contributions to mEPSC shape. Dendritic cable effects account for the slower mEPSC rise in neurons, whereas the slower decay also depends on other factors. Lastly, expression of synaptobrevin transmembrane domain mutants in neurons slowed the rise of HEK cell mEPSCs, thus revealing the impact of synaptic fusion pores. In summary, we show that cocultures of neurons with heterologous cells provide a geometrically simplified and molecularly defined system to investigate the time course of synaptic transmission and to resolve the contribution of vesicles, fusion pores, dendrites, and receptors to this process.

Structural and functional brain-wide alterations in A350V Iqsec2 mutant mice displaying autistic-like behavior

IQSEC2 is an X-linked gene that is associated with autism spectrum disorder (ASD), intellectual disability, and epilepsy. IQSEC2 is a postsynaptic density protein, localized on excitatory synapses as part of the NMDA receptor complex and is suggested to play a role in AMPA receptor trafficking and mediation of long-term depression. Here, we present brain-wide structural volumetric and functional connectivity characterization in a novel mouse model with a missense mutation in the IQ domain of IQSEC2 (A350V). Using high-resolution structural and functional MRI, we show that animals with the A350V mutation display increased whole-brain volume which was further found to be specific to the cerebral cortex and hippocampus. Moreover, using a data-driven approach we identify putative alterations in structure–function relations of the frontal, auditory, and visual networks in A350V mice. Examination of these alterations revealed an increase in functional connectivity between the anterior cingulate cortex and the dorsomedial striatum. We also show that corticostriatal functional connectivity is correlated with individual variability in social behavior only in A350V mice, as assessed using the three-chamber social preference test. Our results at the systems-level bridge the impact of previously reported changes in AMPA receptor trafficking to network-level disruption and impaired social behavior. Further, the A350V mouse model recapitulates similarly reported brain-wide changes in other ASD mouse models, with substantially different cellular-level pathologies that nonetheless result in similar brain-wide alterations, suggesting that novel therapeutic approaches in ASD that result in systems-level rescue will be relevant to IQSEC2 mutations.

Inhibition of soluble epoxide hydrolase (sEH) protects hippocampal neurons and reduces cognitive decline in type 2 diabetic mice.

Diabetes mellitus is a metabolic disorder that can lead to cognitive dysfunction. The hippocampus plays an important role in the cognitive function. Research has identified correlations between hippocampal impairment and diabetes, yet their intermediate remains unclear. Soluble epoxide hydrolase (sEH) is an enzyme that degrades epoxyeicosatrienoic acids (EETs), which have multiple protective effects by suppressing inflammation, apoptosis and oxidative stress. In this study, under diabetic conditions both hippocampal injury and cognitive decline are accompanied by upregulation of sEH. Moreover, the sEH inhibitor trans-4-[4-(3-adamantan-1-y1-ureido)-cyclohexyloxy]-benzoic acid (t-AUCB) prevents cognitive dysfunction and decreased ROS accumulation and apoptosis in the diabetic hippocampus. t-AUCB treatment restored neuronal synaptic plasticity by restoring the expression of the postsynaptic proteins Postsynaptic density protein-95 (PSD95) and N-methyl-d-aspartate receptor subunit 2B (NR2B), the levels of which were positively correlated with Proline-rich tyrosine kinase 2 (Pyk2) levels under diabetic conditions. Thus, we suggest that hippocampal protection via sEH inhibition might be a potential therapeutic approach to attenuate the progression of cognitive decline in diabetes.

Fructose Removal from the Diet Reverses Inflammation, Mitochondrial Dysfunction, and Oxidative Stress in Hippocampus.

Young age is often characterized by high consumption of processed foods and fruit juices rich in fructose, which, besides inducing a tendency to become overweight, can promote alterations in brain function. The aim of this study was therefore to (a) clarify brain effects resulting from fructose consumption in juvenile age, a critical phase for brain development, and (b) verify whether these alterations can be rescued after removing fructose from the diet. Young rats were fed a fructose-rich or control diet for 3 weeks. Fructose-fed rats were then fed a control diet for a further 3 weeks. We evaluated mitochondrial bioenergetics by high-resolution respirometry in the hippocampus, a brain area that is critically involved in learning and memory. Glucose transporter-5, fructose and uric acid levels, oxidative status, and inflammatory and synaptic markers were investigated by Western blotting and spectrophotometric or enzyme-linked immunosorbent assays. A short-term fructose-rich diet induced mitochondrial dysfunction and oxidative stress, associated with an increased concentration of inflammatory markers and decreased Neurofilament-M and post-synaptic density protein 95. These alterations, except for increases in haptoglobin and nitrotyrosine, were recovered by returning to a control diet. Overall, our results point to the dangerous effects of excessive consumption of fructose in young age but also highlight the effect of partial recovery by switching back to a control diet.