miR‐181a negatively modulates synaptic plasticity in hippocampal cultures and its inhibition rescues memory deficits in a mouse model of Alzheimer’s disease

MicroRNAs play a pivotal role in rapid, dynamic, and spatiotemporal modulation of synaptic functions. Among them, recent emerging evidence highlights that microRNA‐181a (miR‐181a) is particularly abundant in hippocampal neurons and controls the expression of key plasticity‐related proteins at synapses. We have previously demonstrated that miR‐181a was upregulated in the hippocampus of a mouse model of Alzheimer's disease (AD) and correlated with reduced levels of plasticity‐related proteins. Here, we further investigated the underlying mechanisms by which miR‐181a negatively modulated synaptic plasticity and memory. In primary hippocampal cultures, we found that an activity‐dependent upregulation of the microRNA‐regulating protein, translin, correlated with reduction of miR‐181a upon chemical long‐term potentiation (cLTP), which induced upregulation of GluA2, a predicted target for miR‐181a, and other plasticity‐related proteins. Additionally, Aβ treatment inhibited cLTP‐dependent induction of translin and subsequent reduction of miR‐181a, and cotreatment with miR‐181a antagomir effectively reversed the effects elicited by Aβ but did not rescue translin levels, suggesting that the activity‐dependent upregulation of translin was upstream of miR‐181a. In mice, a learning episode markedly decreased miR‐181a in the hippocampus and raised the protein levels of GluA2. Lastly, we observed that inhibition of miR‐181a alleviated memory deficits and increased GluA2 and GluA1 levels, without restoring translin, in the 3xTg‐AD model. Taken together, our results indicate that miR‐181a is a major negative regulator of the cellular events that underlie synaptic plasticity and memory through AMPA receptors, and importantly, Aβ disrupts this process by suppressing translin and leads to synaptic dysfunction and memory impairments in AD.     

Soluble oligomers of amyloid-β peptide disrupt membrane trafficking of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor contributing to early synapse dysfunction

β-Amyloid (Aβ), a peptide generated from the amyloid precursor protein, is widely believed to underlie the pathophysiology of Alzheimer disease (AD). Emerging evidences suggest that soluble Aβ oligomers adversely affect synaptic function, leading to cognitive failure associated with AD. The Aβ-induced synaptic dysfunction has been attributed to the synaptic removal of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors (AMPARs). However, the molecular mechanisms underlying the loss of AMPAR induced by Aβ at synapses are largely unknown. In this study we have examined the effect of Aβ oligomers on phosphorylated GluA1 at serine 845, a residue that plays an essential role in the trafficking of AMPARs toward extrasynaptic sites and the subsequent delivery to synapses during synaptic plasticity events. We found that Aβ oligomers reduce basal levels of Ser-845 phosphorylation and surface expression of AMPARs affecting AMPAR subunit composition. Aβ-induced GluA1 dephosphorylation and reduced receptor surface levels are mediated by an increase in calcium influx into neurons through ionotropic glutamate receptors and activation of the calcium-dependent phosphatase calcineurin. Moreover, Aβ oligomers block the extrasynaptic delivery of AMPARs induced by chemical synaptic potentiation. In addition, reduced levels of total and phosphorylated GluA1 are associated with initial spatial memory deficits in a transgenic mouse model of AD. These findings indicate that Aβ oligomers could act as a synaptic depressor affecting the mechanisms involved in the targeting of AMPARs to the synapses during early stages of the disease.     

GluA4 Dependent Plasticity Mechanisms Contribute to Developmental Synchronization of the CA3–CA1 Circuitry in the Hippocampus

During the course of development, molecular mechanisms underlying activity-dependent synaptic plasticity change considerably. At immature CA3–CA1 synapses in the hippocampus, PKA-driven synaptic insertion of GluA4 AMPA receptors is the predominant mechanism for synaptic strengthening. However, the physiological significance of the developmentally restricted GluA4-dependent plasticity mechanisms is poorly understood. Here we have used microelectrode array (MEA) recordings in GluA4 deficient slice cultures to study the role of GluA4 in early development of the hippocampal circuit function. We find that during the first week in culture (DIV2–6) when GluA4 expression is restricted to pyramidal neurons, loss of GluA4 has no effect on the overall excitability of the immature network, but significantly impairs synchronization of the CA3 and CA1 neuronal populations. In the absence of GluA4, the temporal correlation of the population spiking activity between CA3–CA1 neurons was significantly lower as compared to wild-types at DIV6. Our data show that synapse-level defects in transmission and plasticity mechanisms are efficiently compensated for to normalize population firing rate at the immature hippocampal network. However, lack of the plasticity mechanisms typical for the immature synapses may perturb functional coupling between neuronal sub-populations, a defect frequently implicated in the context of developmentally originating neuropsychiatric disorders.

     

MiR‐134‐dependent regulation of Pumilio‐2 is necessary for homeostatic synaptic depression

Neurons employ a set of homeostatic plasticity mechanisms to counterbalance altered levels of network activity. The molecular mechanisms underlying homeostatic plasticity in response to increased network excitability are still poorly understood. Here, we describe a sequential homeostatic synaptic depression mechanism in primary hippocampal neurons involving miRNA‐dependent translational regulation. This mechanism consists of an initial phase of synapse elimination followed by a reinforcing phase of synaptic downscaling. The activity‐regulated microRNA miR‐134 is necessary for both synapse elimination and the structural rearrangements leading to synaptic downscaling. Results from miR‐134 inhibition further uncover a differential requirement for GluA1/2 subunits for the functional expression of homeostatic synaptic depression. Downregulation of the miR‐134 target Pumilio‐2 in response to chronic activity, which selectively occurs in the synapto‐dendritic compartment, is required for miR‐134‐mediated homeostatic synaptic depression. We further identified polo‐like kinase 2 (Plk2) as a novel target of Pumilio‐2 involved in the control of GluA2 surface expression. In summary, we have described a novel pathway of homeostatic plasticity that stabilizes neuronal circuits in response to increased network activity.     

Nedd4-mediated AMPA receptor ubiquitination regulates receptor turnover and trafficking

α-Amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid receptors (AMPARs) are the primary mediators of excitatory synaptic transmission in the brain. Alterations in AMPAR localization and turnover have been considered critical mechanisms underpinning synaptic plasticity and higher brain functions, but the molecular processes that control AMPAR trafficking and stability are still not fully understood. Here, we report that mammalian AMPARs are subject to ubiquitination in neurons and in transfected heterologous cells. Ubiquitination facilitates AMPAR endocytosis, leading to a reduction in AMPAR cell-surface localization and total receptor abundance. Mutation of lysine residues to arginine residues at the glutamate receptor subunit 1 (GluA1) C-terminus dramatically reduces GluA1 ubiquitination and abolishes ubiquitin-dependent GluA1 internalization and degradation, indicating that the lysine residues, particularly K868, are sites of ubiquitination. We also find that the E3 ligase neural precursor cell expressed, developmentally down-regulated 4 (Nedd4) is enriched in synaptosomes and co-localizes and associates with AMPARs in neurons. Nedd4 expression leads to AMPAR ubiquitination, leading to reduced AMPAR surface expression and suppressed excitatory synaptic transmission. Conversely, knockdown of Nedd4 by specific siRNAs abolishes AMPAR ubiquitination. These data indicate that Nedd4 is the E3 ubiquitin ligase responsible for AMPAR ubiquitination, a modification that regulates multiple aspects of AMPAR molecular biology including trafficking, localization and stability.     

Adenosine A1 Receptor-Mediated Endocytosis of AMPA Receptors Contributes to Impairments in Long-Term Potentiation (LTP) in the Middle-Aged Rat Hippocampus

Aging causes multiple changes in the mammalian brain, including changes in synaptic signaling. Previous reports have shown increased extracellular adenosine in the aging brain, and we recently reported that activation of adenosine A1 receptors (A1Rs) induces AMPA receptor (AMPAR) internalization in rat hippocampus. This study investigated whether aging-related changes in the rat hippocampus include altered surface expression of adenosine A1 and A2A receptors, and whether these changes correspond to changes in AMPAR surface expression and altered synaptic plasticity. We found reduced A1R surface expression in middle-aged rat hippocampus, and also reduced GluA1 and GluA2 AMPAR subunit surface expression. Using a chemically-induced LTP (cLTP) experimental protocol, we recorded fEPSPs in young (1 month old) and middle-aged (7–12 month old) rat hippocampal slices. There were significant impairments in cLTP in middle-aged slices, suggesting impaired synaptic plasticity. Since we previously showed that the A1R agonist N⁶-cyclopentyladenosine (CPA) reduced both A1Rs and GluA2/GluA1 AMPARs, we hypothesized that the observed impaired synaptic plasticity in middle-aged brains is regulated by A1R-mediated AMPAR internalization by clathrin-mediated endocytosis. Following cLTP, we found a significant increase in GluA1 and GluA2 surface expression in young rats, which was blunted in middle-aged brains or in young brains pretreated with CPA. Blocking A1Rs with 8-cyclopentyl-1,3-dipropylxanthine or AMPAR endocytosis with either Tat-GluA2-3Y peptide or dynasore (dynamin inhibitor) similarly enhanced AMPAR surface expression following cLTP. These data suggest that age-dependent alteration in adenosine receptor expression contributes to increased AMPAR endocytosis and impaired synaptic plasticity in aged brains.     

Small GTPase Rab17 Regulates the Surface Expression of Kainate Receptors but Not α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid (AMPA) Receptors in Hippocampal Neurons via Dendritic Trafficking of Syntaxin-4 Protein

Glutamate receptors are fundamental for control synaptic transmission, synaptic plasticity, and neuronal excitability. However, many of the molecular mechanisms underlying their trafficking remain elusive. We previously demonstrated that the small GTPase Rab17 regulates dendritic trafficking in hippocampal neurons. Here, we investigated the role(s) of Rab17 in AMPA receptor (AMPAR) and kainate receptor (KAR) trafficking. Although Rab17 knockdown did not affect surface expression of the AMPAR subunit GluA1 under basal or chemically induced long term potentiation conditions, it significantly reduced surface expression of the KAR subunit GluK2. Rab17 co-localizes with Syntaxin-4 in the soma, dendritic shaft, the tips of developing hippocampal neurons, and in spines. Rab17 knockdown caused Syntaxin-4 redistribution away from dendrites and into axons in developing hippocampal neurons. Syntaxin-4 knockdown reduced GluK2 but had no effect on GluA1 surface expression. Moreover, overexpression of constitutively active Rab17 promoted dendritic surface expression of GluK2 by enhancing Syntaxin-4 translocation to dendrites. These data suggest that Rab17 mediates the dendritic trafficking of Syntaxin-4 to selectively regulate dendritic surface insertion of GluK2-containing KARs in rat hippocampal neurons.     

Sequential Delivery of Synaptic GluA1- and GluA4-containing AMPA Receptors (AMPARs) by SAP97 Anchored Protein Complexes in Classical Conditioning

Multiple signaling pathways are involved in AMPAR trafficking to synapses during synaptic plasticity and learning. The mechanisms for how these pathways are coordinated in parallel but maintain their functional specificity involves subcellular compartmentalization of kinase function by scaffolding proteins, but how this is accomplished is not well understood. Here, we focused on characterizing the molecular machinery that functions in the sequential synaptic delivery of GluA1- and GluA4-containing AMPARs using an in vitro model of eyeblink classical conditioning. We show that conditioning induces the interaction of selective protein complexes with the key structural protein SAP97, which tightly regulates the synaptic delivery of GluA1 and GluA4 AMPAR subunits. The results demonstrate that in the early stages of conditioning the initial activation of PKA stimulates the formation of a SAP97-AKAP/PKA-GluA1 protein complex leading to synaptic delivery of GluA1-containing AMPARs through a SAP97-PSD95 interaction. This is followed shortly thereafter by generation of a SAP97-KSR1/PKC-GluA4 complex for GluA4 AMPAR subunit delivery again through a SAP97-PSD95 interaction. These data suggest that SAP97 forms the molecular backbone of a protein scaffold critical for delivery of AMPARs to the PSD during conditioning. Together, the findings reveal a cooperative interaction of multiple scaffolding proteins for appropriately timed delivery of subunit-specific AMPARs to synapses and support a sequential two-stage model of AMPAR synaptic delivery during classical conditioning.     

TRPV1 activation alleviates cognitive and synaptic plasticity impairments through inhibiting AMPAR endocytosis in APP23/PS45 mouse model of Alzheimer’s disease

Alzheimer's disease (AD) is one of the most common causes of neurodegenerative diseases in the elderly. The accumulation of amyloid‐β (Aβ) peptides is one of the pathological hallmarks of AD and leads to the impairments of synaptic plasticity and cognitive function. The transient receptor potential vanilloid 1 (TRPV1), a nonselective cation channel, is involved in synaptic plasticity and memory. However, the role of TRPV1 in AD pathogenesis remains largely elusive. Here, we reported that the expression of TRPV1 was decreased in the brain of APP23/PS45 double transgenic AD model mice. Genetic upregulation of TRPV1 by adeno‐associated virus (AAV) inhibited the APP processing and Aβ deposition in AD model mice. Meanwhile, upregulation of TRPV1 ameliorated the deficits of hippocampal CA1 long‐term potentiation (LTP) and spatial learning and memory through inhibiting GluA2‐containing α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid receptor (AMPAR) endocytosis. Furthermore, pharmacological activation of TRPV1 by capsaicin (1 mg/kg, i.p.), an agonist of TRPV1, dramatically reversed the impairments of hippocampal CA1 LTP and spatial learning and memory in AD model mice. Taken together, these results indicate that TRPV1 activation effectively ameliorates cognitive and synaptic functions through inhibiting AMPAR endocytosis in AD model mice and could be a novel molecule for AD treatment.     

Phosphorylation of AMPA receptors is required for sensory deprivation-induced homeostatic synaptic plasticity

Sensory experience, and the lack thereof, can alter the function of excitatory synapses in the primary sensory cortices. Recent evidence suggests that changes in sensory experience can regulate the synaptic level of Ca(2+)-permeable AMPA receptors (CP-AMPARs). However, the molecular mechanisms underlying such a process have not been determined. We found that binocular visual deprivation, which is a well-established in vivo model to produce multiplicative synaptic scaling in visual cortex of juvenile rodents, is accompanied by an increase in the phosphorylation of AMPAR GluR1 (or GluA1) subunit at the serine 845 (S845) site and the appearance of CP-AMPARs at synapses. To address the role of GluR1-S845 in visual deprivation-induced homeostatic synaptic plasticity, we used mice lacking key phosphorylation sites on the GluR1 subunit. We found that mice specifically lacking the GluR1-S845 site (GluR1-S845A mutants), which is a substrate of cAMP-dependent kinase (PKA), show abnormal basal excitatory synaptic transmission and lack visual deprivation-induced homeostatic synaptic plasticity. We also found evidence that increasing GluR1-S845 phosphorylation alone is not sufficient to produce normal multiplicative synaptic scaling. Our study provides concrete evidence that a GluR1 dependent mechanism, especially S845 phosphorylation, is a necessary pre-requisite step for in vivo homeostatic synaptic plasticity.     

PIKE-mediated PI3-kinase activity is required for AMPA receptor surface expression

AMPAR (α-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid receptor) is an ion channel involved in the formation of synaptic plasticity. However, the molecular mechanism that couples plasticity stimuli to the trafficking of postsynaptic AMPAR remains poorly understood. Here, we show that PIKE (phosphoinositide 3-kinase enhancer) GTPases regulate neuronal AMPAR activity by promoting GluA2/GRIP1 association. PIKE-L directly interacts with both GluA2 and GRIP1 and forms a tertiary complex upon glycine-induced NMDA receptor activation. PIKE-L is also essential for glycine-induced GluA2-associated PI3K activation. Genetic ablation of PIKE (PIKE(-/-)) in neurons suppresses GluA2-associated PI3K activation, therefore inhibiting the subsequent surface expression of GluA2 and the formation of long-term potentiation. Our findings suggest that PIKE-L is a critical factor in controlling synaptic AMPAR insertion.     

Molecular mechanisms coordinating functional and morphological plasticity at the synapse: Role of GluA2/N-cadherin interaction-mediated actin signaling in mGluR-dependent LTD

Long-lasting synaptic plasticity involves changes in both synaptic morphology and electrical signaling (here referred to as structural and functional plasticity). Recent studies have revealed a myriad of molecules and signaling processes that are critical for each of these two forms of plasticity, but whether and how they are mechanistically linked to achieve coordinated changes remain controversial.It is well accepted that functional plasticity at the excitatory synapse is dependent upon the activities of glutamate receptors. While the activation of NMDARs (N-methyl-D-aspartic acid receptors) and/or mGluRs (metabotropic glutamate receptors) is required for the induction of many forms of plasticity, AMPARs (alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptors), the principal mediators of fast excitatory synaptic transmission, are the ultimate targets of modifications that express functional plasticity. Investigations exploring structural plasticity have been mainly focused on the small membranous protrusions on the dendrites called spines. The morphological regulation of these spines is mediated by the reorganization of the actin cytoskeleton, the predominant structural component of the synapse. In this regard, the Rho family of GTPases, particularly Rac1, RhoA and Cdc42, is found to be the central regulator of spine actin and structural plasticity of the synapse.It is thought that the collaborative interaction between functional and structural factors underlies the sustained or permanent nature of long-lasting synaptic plasticity such as long-term potentiation (LTP) and long-term depression (LTD), the most extensively studied forms of synaptic plasticity widely regarded as cellular mechanisms for learning and memory. However, data specifically pertaining to whether and how these two distinct components are linked at the molecular level remain sparse. In this regard, we have identified a number of synaptic proteins that are involved in both structural and functional changes during mGluR-dependent LTD (mGluR-LTD). Among these are the GluA2 (formerly called GluR2) subunit of AMPARs, Rac1 and Rac1-activated kinases. We have discovered that these proteins interact and reciprocally regulate each other, which led us to hypothesize that the GluA2–Rac1 interaction may serve as a coordinator between functional and morphological plasticity. In this review, we will briefly discuss the available evidence to support such a hypothesis.     

Endocytic Adaptor Epidermal Growth Factor Receptor Substrate 15 (Eps15) Is Involved in the Trafficking of Ubiquitinated α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid Receptors

AMPA-type glutamate receptors (AMPARs) play a critical role in mediating fast excitatory synaptic transmission in the brain. Alterations in receptor expression, distribution, and trafficking have been shown to underlie synaptic plasticity and higher brain functions, including learning and memory, as well as brain dysfunctions such as drug addiction and psychological disorders. Therefore, it is essential to elucidate the molecular mechanisms that regulate AMPAR dynamics. We have shown previously that mammalian AMPARs are subject to posttranslational modification by ubiquitin, with AMPAR ubiquitination enhancing receptor internalization and reducing AMPAR cell surface expression. Here we report a crucial role for epidermal growth factor receptor substrate 15 (Eps15), an endocytic adaptor, in ubiquitination-dependent AMPAR internalization. We find that suppression or overexpression of Eps15 results in changes in AMPAR surface expression. Eps15 interacts with AMPARs, which requires Nedd4-mediated GluA1 ubiquitination and the ubiquitin-interacting motif of Eps15. Importantly, we find that Eps15 plays an important role in AMPAR internalization. Knockdown of Eps15 suppresses the internalization of GluA1 but not the mutant GluA1 that lacks ubiquitination sites, indicating a role of Eps15 for the internalization of ubiquitinated AMPARs. These results reveal a novel molecular mechanism employed specifically for the trafficking of the ubiquitin-modified AMPARs.     

The Biochemistry,Ultrastructure, and Subunit Assembly Mechanism of AMPA Receptors

The AMPA-type ionotropic glutamate receptors (AMPA-Rs) are tetrameric ligand-gated ion channels that play crucial roles in synaptic transmission and plasticity. Our knowledge about the ultrastructure and subunit assembly mechanisms of intact AMPA-Rs was very limited. However, the new studies using single particle EM and X-ray crystallography are revealing important insights. For example, the tetrameric crystal structure of the GluA2cryst construct provided the atomic view of the intact receptor. In addition, the single particle EM structures of the subunit assembly intermediates revealed the conformational requirement for the dimer-to-tetramer transition during the maturation of AMPA-Rs. These new data in the field provide new models and interpretations. In the brain, the native AMPA-R complexes contain auxiliary subunits that influence subunit assembly, gating, and trafficking of the AMPA-Rs. Understanding the mechanisms of the auxiliary subunits will become increasingly important to precisely describe the function of AMPA-Rs in the brain. The AMPA-R proteomics studies continuously reveal a previously unexpected degree of molecular heterogeneity of the complex. Because the AMPA-Rs are important drug targets for treating various neurological and psychiatric diseases, it is likely that these new native complexes will require detailed mechanistic analysis in the future. The current ultrastructural data on the receptors and the receptor-expressing stable cell lines that were developed during the course of these studies are useful resources for high throughput drug screening and further drug designing. Moreover, we are getting closer to understanding the precise mechanisms of AMPA-R-mediated synaptic plasticity.     

Identification of a novel signaling pathway and its relevance for GluA1 recycling

We previously showed that the serum- and glucocorticoid-inducible kinase 3 (SGK3) increases the AMPA-type glutamate receptor GluA1 protein in the plasma membrane. The activation of AMPA receptors by NMDA-type glutamate receptors eventually leads to postsynaptic neuronal plasticity. Here, we show that SGK3 mRNA is upregulated in the hippocampus of new-born wild type Wistar rats after NMDA receptor activation. We further demonstrate in the Xenopus oocyte expression system that delivery of GluA1 protein to the plasma membrane depends on the small GTPase RAB11. This RAB-dependent GluA1 trafficking requires phosphorylation and activation of phosphoinositol-3-phosphate-5-kinase (PIKfyve) and the generation of PI(3,5)P(2). In line with this mechanism we could show PIKfyve mRNA expression in the hippocampus of wild type C57/BL6 mice and phosphorylation of PIKfyve by SGK3. Incubation of hippocampal slices with the PIKfyve inhibitor YM201636 revealed reduced CA1 basal synaptic activity. Furthermore, treatment of primary hippocampal neurons with YM201636 altered the GluA1 expression pattern towards reduced synaptic expression of GluA1. Our findings demonstrate for the first time an involvement of PIKfyve and PI(3,5)P(2) in NMDA receptor-triggered synaptic GluA1 trafficking. This new regulatory pathway of GluA1 may contribute to synaptic plasticity and memory.     

STIM2 regulates PKA-dependent phosphorylation and trafficking of AMPARs

STIMs (STIM1 and STIM2 in mammals) are transmembrane proteins that reside in the endoplasmic reticulum (ER) and regulate store-operated Ca²⁺ entry (SOCE). The function of STIMs in the brain is only beginning to be explored, and the relevance of SOCE in nerve cells is being debated. Here we identify STIM2 as a central organizer of excitatory synapses. STIM2, but not its paralogue STIM1, influences the formation of dendritic spines and shapes basal synaptic transmission in excitatory neurons. We further demonstrate that STIM2 is essential for cAMP/PKA-dependent phosphorylation of the AMPA receptor (AMPAR) subunit GluA1. cAMP triggers rapid migration of STIM2 to ER–plasma membrane (PM) contact sites, enhances recruitment of GluA1 to these ER-PM junctions, and promotes localization of STIM2 in dendritic spines. Both biochemical and imaging data suggest that STIM2 regulates GluA1 phosphorylation by coupling PKA to the AMPAR in a SOCE-independent manner. Consistent with a central role of STIM2 in regulating AMPAR phosphorylation, STIM2 promotes cAMP-dependent surface delivery of GluA1 through combined effects on exocytosis and endocytosis. Collectively our results point to a unique mechanism of synaptic plasticity driven by dynamic assembly of a STIM2 signaling complex at ER-PM contact sites.     

Lock and chop: A novel method for the generation of a PICK1 PDZ domain and piperidine‐based inhibitor co‐crystal structure

The membrane protein interacting with kinase C1 (PICK1) plays a trafficking role in the internalization of neuron receptors such as the amino‐3‐hydroxyl‐5‐methyl‐4‐isoxazole‐propionate (AMPA) receptor. Reduction of surface AMPA type receptors on neurons reduces synaptic communication leading to cognitive impairment in progressive neurodegenerative diseases such as Alzheimer disease. The internalization of AMPA receptors is mediated by the PDZ domain of PICK1 which binds to the GluA2 subunit of AMPA receptors and targets the receptor for internalization through endocytosis, reducing synaptic communication. We planned to block the PICK1‐GluA2 protein–protein interaction with a small molecule inhibitor to stabilize surface AMPA receptors as a therapeutic possibility for neurodegenerative diseases. Using a fluorescence polarization assay, we identified compound BIO124 as a modest inhibitor of the PICK1‐GluA2 interaction. We further tried to improve the binding affinity of BIO124 using structure‐aided drug design but were unsuccessful in producing a co‐crystal structure using previously reported crystallography methods for PICK1. Here, we present a novel method through which we generated a co‐crystal structure of the PDZ domain of PICK1 bound to BIO124.     

Incorporation of inwardly rectifying AMPA receptors at silent synapses during hippocampal long-term potentiation

Despite decades of study, the mechanisms by which synapses express the increase in strength during long-term potentiation (LTP) remain an area of intense interest. Here, we have studied how AMPA receptor subunit composition changes during the early phases of hippocampal LTP in CA1 pyramidal neurons. We studied LTP at silent synapses that initially lack AMPA receptors, but contain NMDA receptors. We show that strongly inwardly rectifying AMPA receptors are initially incorporated at silent synapses during LTP and are then subsequently replaced by non-rectifying AMPA receptors. These findings suggest that silent synapses initially incorporate GluA2-lacking, calcium-permeable AMPA receptors during LTP that are then replaced by GluA2-containing calcium-impermeable receptors. We also show that LTP consolidation at CA1 synapses requires a rise in intracellular calcium concentration during the early phase of expression, indicating that calcium influx through the GluA2-lacking AMPA receptors drives their replacement by GluA2-containing receptors during LTP consolidation. Taken together with previous studies in hippocampus and in other brain regions, these findings suggest that a common mechanism for the expression of activity-dependent glutamatergic synaptic plasticity involves the regulation of GluA2-subunit composition and highlights a critical role for silent synapses in this process.     

Synaptically Released Matrix Metalloproteinase Activity in Control of Structural Plasticity and the Cell Surface Distribution of GluA1-AMPA Receptors

Synapses are particularly prone to dynamic alterations and thus play a major role in neuronal plasticity. Dynamic excitatory synapses are located at the membranous neuronal protrusions called dendritic spines. The ability to change synaptic connections involves both alterations at the morphological level and changes in postsynaptic receptor composition. We report that endogenous matrix metalloproteinase (MMP) activity promotes the structural and functional plasticity of local synapses by its effect on glutamate receptor mobility and content. We used live imaging of cultured hippocampal neurons and quantitative morphological analysis to show that chemical long-term potentiation (cLTP) induces the permanent enlargement of a subset of small dendritic spines in an MMP-dependent manner. We also used a superresolution microscopy approach and found that spine expansion induced by cLTP was accompanied by MMP-dependent immobilization and synaptic accumulation as well as the clustering of GluA1-containing AMPA receptors. Altogether, our results reveal novel molecular and cellular mechanisms of synaptic plasticity.     

mGluR long‐term depression regulates GluA2 association with COPII vesicles and exit from the endoplasmic reticulum

mGluR long‐term depression (mGluR‐LTD) is a form of synaptic plasticity induced at excitatory synapses by metabotropic glutamate receptors (mGluRs). mGluR‐LTD reduces synaptic strength and is relevant to learning and memory, autism, and sensitization to cocaine; however, the mechanism is not known. Here we show that activation of Group I mGluRs in medium spiny neurons induces trafficking of GluA2 from the endoplasmic reticulum (ER) to the synapse by enhancing GluA2 binding to essential COPII vesicle proteins, Sec23 and Sec13. GluA2 exit from the ER further depends on IP3 and Ryanodine receptor‐controlled Ca²⁺ release as well as active translation. Synaptic insertion of GluA2 is coupled to removal of high‐conducting Ca²⁺‐permeable AMPA receptors from synapses, resulting in synaptic depression. This work demonstrates a novel mechanism in which mGluR signals release AMPA receptors rapidly from the ER and couple ER release to GluA2 synaptic insertion and GluA1 removal.