A competitive game of synaptic tag

LTP maintenance is thought to require an activity-dependent "synaptic tag" that allows potentiated synapses to sequester factors necessary for maintenance. In a paper in this issue of Neuron, Fonseca et al. show that synapses can compete with each other for such maintenance factors, so that additional potentiation of one input results in loss of potentiation of another. These data suggest that LTP maintenance is a dynamic and competitive process.     

Making space for rats: from synapse to place code

The formation of spatial memories is believed to depend on long-term potentiation (LTP) of synapses within the hippocampal network. In this issue of Neuron, Dragoi et al. demonstrate that LTP can cause changes in the hippocampal representation of space without disrupting network dynamics. These results help define the elusive relationship between cellular-level synaptic plasticity and systems-level neural coding.     

Long-term potentiation in cultured hippocampal neurons

Studies performed on low-density primary neuronal cultures have enabled dissection of molecular and cellular changes during N-methyl-D-aspartate (NMDA) receptor-dependent long-term potentiation (LTP). Various electrophysiological and chemical induction protocols were developed for the persistent enhancement of excitatory synaptic transmission in hippocampal neuronal cultures. The characterisation of these plasticity models confirmed that they share many key properties with the LTP of CA1 neurons, extensively studied in hippocampal slices using electrophysiological techniques. For example, LTP in dissociated hippocampal neuronal cultures is also dependent on Ca(2+) influx through post-synaptic NMDA receptors, subsequent activation and autophosphorylation of the Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and an increase in alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor insertion at the post-synaptic membrane. The availability of models of LTP in cultured hippocampal neurons significantly facilitated the monitoring of changes in endogenous postsynaptic receptor proteins and the investigation of the associated signalling mechanisms that underlie LTP. A central feature of LTP of excitatory synapses is the recruitment of AMPA receptors at the postsynaptic site. Results from the use of cell culture-based models started to establish the mechanism by which synaptic input controls a neuron's ability to modify its synapses in LTP. This review focuses on key features of various LTP induction protocols in dissociated hippocampal neuronal cultures and the applications of these plasticity models for the investigation of activity-induced changes in native AMPA receptors.     

Synaptic neurotransmission depression in ventral tegmental dopamine neurons and cannabinoid-associated addictive learning

Drug addiction is an association of compulsive drug use with long-term associative learning/memory. Multiple forms of learning/memory are primarily subserved by activity- or experience-dependent synaptic long-term potentiation (LTP) and long-term depression (LTD). Recent studies suggest LTP expression in locally activated glutamate synapses onto dopamine neurons (local Glu-DA synapses) of the midbrain ventral tegmental area (VTA) following a single or chronic exposure to many drugs of abuse, whereas a single exposure to cannabinoid did not significantly affect synaptic plasticity at these synapses. It is unknown whether chronic exposure of cannabis (marijuana or cannabinoids), the most commonly used illicit drug worldwide, induce LTP or LTD at these synapses. More importantly, whether such alterations in VTA synaptic plasticity causatively contribute to drug addictive behavior has not previously been addressed. Here we show in rats that chronic cannabinoid exposure activates VTA cannabinoid CB1 receptors to induce transient neurotransmission depression at VTA local Glu-DA synapses through activation of NMDA receptors and subsequent endocytosis of AMPA receptor GluR2 subunits. A GluR2-derived peptide blocks cannabinoid-induced VTA synaptic depression and conditioned place preference, i.e., learning to associate drug exposure with environmental cues. These data not only provide the first evidence, to our knowledge, that NMDA receptor-dependent synaptic depression at VTA dopamine circuitry requires GluR2 endocytosis, but also suggest an essential contribution of such synaptic depression to cannabinoid-associated addictive learning, in addition to pointing to novel pharmacological strategies for the treatment of cannabis addiction.     

Long-lasting potentiation of excitatory synaptic signaling to the crayfish lateral giant neuron

The neural circuit that underlies the lateral giant fiber (LG)-mediated reflex escape in crayfish has provided findings relating synaptic change to nonassociative learning such as sensitization and habituation. The LGs receive sensory inputs from the primary sensory afferents and a group of mechanosensory interneurons (MSIs). An increase of excitability by suprathreshold repetitive excitation of this circuit, which is similar to Hebbian long-term potentiation (LTP), has been reported [Miller et al. (1987) J Neurosci 7:1081]. This potentiation was previously thought to result from the enhancement of transmission at cholinergic synapses between primary afferents and MSIs but not the electrical synapses onto LG. In this study, we found that potentiation of synaptic signaling at the electrical synapse onto LG can also be induced when the synapse was activated with subthreshold repetitive pulses or with a few strong suprathreshold shocks. LG LTP was induced in the preparation which had received pulses at limited frequency range. Although whether this LTP is involved in the learning process of escape behavior in crayfish is not clear, the intensity and amount of sensory stimulation used here mimicked those that could easily be produced by a predator trying to catch a crayfish and could be of adaptive significance in life.     

Hippocampal long term potentiation: silent synapses and beyond.

Long-term, activity-driven synaptic plasticity allows neuronal networks to constantly and durably adjust synaptic gains between synaptic partners. These processes have been proposed to serve as a substrate for learning and memory. Long-term synaptic potentiation (LTP) has been observed at many central excitatory synapses and perhaps most extensively studied at Schaffer collaterals synapses onto hippocampal CA1 neurons. Multiple contradictory models were proposed to account for this form of LTP. However, recent evidence suggests that some synapses are initially devoid of functional AMPA receptors which can be incorporated during LTP. This new model appears to account for most, but not all, properties of this form of plasticity. Indeed, several mechanisms seem to act in parallel to specifically enhance AMPA-receptor mediated synaptic transmission.     

Synaptic vesicle distribution by conveyor belt

The equal distribution of synaptic vesicles among synapses along the axon is critical for robust neurotransmission. Wong et al. show that the continuous circulation of synaptic vesicles throughout the axon driven by molecular motors ultimately yields this even distribution.     

Heterosynaptic LTP in early development.

The formation of precise synaptic connections in the developing central nervous system involves activity-dependent control of synaptic strength. Correlated activity between the afferent input and postsynaptic neuron strengthens connections through long-term potentiation (LTP), while uncorrelated synaptic inputs are selectively diminished. In this issue of Neuron, Tao et al. report that LTP in young animals has less synaptic specificity than in older animals, indicating that the properties of LTP change during development.     

Glutamate uptake determines pathway specificity of long-term potentiation in the neural circuitry of fear conditioning

Long-term synaptic modifications in afferent inputs to the amygdala underlie fear conditioning in animals. Fear conditioning to a single sensory modality does not generalize to other cues, implying that synaptic modifications in fear conditioning pathways are input specific. The mechanisms of pathway specificity of long-term potentiation (LTP) are poorly understood. Here we show that inhibition of glutamate transporters leads to the loss of input specificity of LTP in the amygdala slices, as assessed by monitoring synaptic responses at two independent inputs converging on a single postsynaptic neuron. Diffusion of glutamate ("spillover") from stimulated synapses, paired with postsynaptic depolarization, is sufficient to induce LTP in the heterosynaptic pathway, whereas an enzymatic glutamate scavenger abolishes this effect. These results establish active glutamate uptake as a crucial mechanism maintaining the pathway specificity of LTP in the neural circuitry of fear conditioning.     

Multiple Forms and Distribution of Calcium/Calmodulin-Stimulated Protein Kinase II in Brain

In recent years, the enzyme Ca²⁺/calmodulin-stimulated protein kinase II¹ (CaM-PK II) as attracted a great deal of interest. CaM-PK II is the most abundant calmodulin-stimulated protein kinase in brain, where it is particularly enriched in neurons (Ouimet et al., 1984; Erondu and Kennedy, 1985; Lin et al., 1987; Scholz et al., 1988). Neuronal CaM-PK II has been suggested to be involved in several phenomena associated with synaptic plasticity (Lisman and Goldring, 1988; Kelly, 1992), including long-term potentiation (Malinow et al., 1988; Malenka et al.,1989), neurotransmission (Nichols et al., 1990; Siekevitz, 1991), and learning (for review, see Rostas, 1991). This enzyme has also been postulated to be selectively vulnerable in several pathological condition, including epilepsy/kindling (Bronstein et al.,1990; Wu et al., 1990), cerebral ischemia (Taft et al., 1988), and organophosphorus toxicity (Abou-Donia and Lapadula, 1990).     

Single vesicle tracking for studying synaptic vesicle dynamics in small central synapses

Sustained neurotransmission is driven by a continuous supply of synaptic vesicles to the release sites and modulated by synaptic vesicle dynamics. However, synaptic vesicle dynamics in synapses remain elusive because of technical limitations. Recent advances in fluorescence imaging techniques have enabled the tracking of single synaptic vesicles in small central synapses in living neurons. Single vesicle tracking has uncovered a wealth of new information about synaptic vesicle dynamics both within and outside presynaptic terminals, showing that single vesicle tracking is an effective tool for studying synaptic vesicle dynamics. Particularly, single vesicle tracking with high spatiotemporal resolution has revealed the dependence of synaptic vesicle dynamics on the location, stages of recycling, and neuronal activity. This review summarizes the recent findings from single synaptic vesicle tracking in small central synapses and their implications in synaptic transmission and pathogenic mechanisms of neurodegenerative diseases.     

Tag-Trigger-Consolidation: A Model of Early and Late Long-Term-Potentiation and Depression

Changes in synaptic efficacies need to be long-lasting in order to serve as a substrate for memory. Experimentally, synaptic plasticity exhibits phases covering the induction of long-term potentiation and depression (LTP/LTD) during the early phase of synaptic plasticity, the setting of synaptic tags, a trigger process for protein synthesis, and a slow transition leading to synaptic consolidation during the late phase of synaptic plasticity. We present a mathematical model that describes these different phases of synaptic plasticity. The model explains a large body of experimental data on synaptic tagging and capture, cross-tagging, and the late phases of LTP and LTD. Moreover, the model accounts for the dependence of LTP and LTD induction on voltage and presynaptic stimulation frequency. The stabilization of potentiated synapses during the transition from early to late LTP occurs by protein synthesis dynamics that are shared by groups of synapses. The functional consequence of this shared process is that previously stabilized patterns of strong or weak synapses onto the same postsynaptic neuron are well protected against later changes induced by LTP/LTD protocols at individual synapses.     

Fusion pore modulation as a presynaptic mechanism contributing to expression of long-term potentiation

Working on the idea that postsynaptic and presynaptic mechanisms of long-term potentiation (LTP) expression are not inherently mutually exclusive, we have looked for the existence and functionality of presynaptic mechanisms for augmenting transmitter release in hippocampal slices. Specifically, we asked if changes in glutamate release might contribute to the conversion of 'silent synapses' that show N-methyl-D-aspartate (NMDA) responses but no detectable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) responses, to ones that exhibit both. Here, we review experiments where NMDA receptor responses provided a bioassay of cleft glutamate concentration, using opposition between peak [glu](cleft )and a rapidly reversible antagonist, L-AP5. We discuss findings of a dramatic increase in peak [glu](cleft) upon expression of pairing-induced LTP (Choi). We present simulations with a quantitative model of glutamatergic synaptic transmission that includes modulation of the presynaptic fusion pore, realistic cleft geometry and a distributed array of postsynaptic receptors and glutamate transporters. The modelling supports the idea that changes in the dynamics of glutamate release can contribute to synaptic unsilencing. We review direct evidence from Renger et al., in accord with the modelling, that trading off the strength and duration of the glutamate transient can markedly alter AMPA receptor responses with little effect on NMDA receptor responses. An array of additional findings relevant to fusion pore modulation and its proposed contribution to LTP expression are considered.     

AMPA receptor regulation during synaptic plasticity in hippocampus and neocortex

Discovery of long-term potentiation (LTP) in the dentate gyrus of the rabbit hippocampus by Bliss and L?mo opened up a whole new field to study activity-dependent long-term synaptic modifications in the brain. Since then hippocampal synapses have been a key model system to study the mechanisms of different forms of synaptic plasticity. At least for the postsynaptic forms of LTP and long-term depression (LTD), regulation of AMPA receptors (AMPARs) has emerged as a key mechanism. While many of the synaptic plasticity mechanisms uncovered in at the hippocampal synapses apply to synapses across diverse brain regions, there are differences in the mechanisms that often reveal the specific functional requirements of the brain area under study. Here we will review AMPAR regulation underlying synaptic plasticity in hippocampus and neocortex. The main focus of this review will be placed on postsynaptic forms of synaptic plasticity that impinge on the regulation of AMPARs using hippocampal CA1 and primary sensory cortices as examples. And through the comparison, we will highlight the key similarities and functional differences between the two synapses.     

Molecular constraints on synaptic tagging and maintenance of long-term potentiation: a predictive model

Protein synthesis-dependent, late long-term potentiation (LTP) and depression (LTD) at glutamatergic hippocampal synapses are well characterized examples of long-term synaptic plasticity. Persistent increased activity of protein kinase M ζ (PKMζ) is thought essential for maintaining LTP. Additional spatial and temporal features that govern LTP and LTD induction are embodied in the synaptic tagging and capture (STC) and cross capture hypotheses. Only synapses that have been "tagged" by a stimulus sufficient for LTP and learning can "capture" PKMζ. A model was developed to simulate the dynamics of key molecules required for LTP and LTD. The model concisely represents relationships between tagging, capture, LTD, and LTP maintenance. The model successfully simulated LTP maintained by persistent synaptic PKMζ, STC, LTD, and cross capture, and makes testable predictions concerning the dynamics of PKMζ. The maintenance of LTP, and consequently of at least some forms of long-term memory, is predicted to require continual positive feedback in which PKMζ enhances its own synthesis only at potentiated synapses. This feedback underlies bistability in the activity of PKMζ. Second, cross capture requires the induction of LTD to induce dendritic PKMζ synthesis, although this may require tagging of a nearby synapse for LTP. The model also simulates the effects of PKMζ inhibition, and makes additional predictions for the dynamics of CaM kinases. Experiments testing the above predictions would significantly advance the understanding of memory maintenance.     

Studying Signal Transduction in Single Dendritic Spines

Many forms of synaptic plasticity are triggered by biochemical signaling that occurs in small postsynaptic compartments called dendritic spines, each of which typically houses the postsynaptic terminal associated with a single glutamatergic synapse. Recent advances in optical techniques allow investigators to monitor biochemical signaling in single dendritic spines and thus reveal the signaling mechanisms that link synaptic activity and the induction of synaptic plasticity. This is mostly in the study of Ca²⁺-dependent forms of synaptic plasticity for which many of the steps between Ca²⁺ influx and changes to the synapse are now known. This article introduces the new techniques used to investigate signaling in single dendritic spines and the neurobiological insights that they have produced.Each neuron typically receives 1000–10,000 synaptic inputs and sends information to an axon, which branches to produce a similar number of synaptic outputs. Most excitatory postsynaptic terminals are associated with dendritic spines, small protrusions emanating from the dendritic surface (Nimchinsky et al. 2002; Alvarez and Sabatini 2007). Each spine has a volume of ∼0.1 femtoliter, and connects to the parent dendrite through a narrow neck, which acts as a diffusion barrier and compartmentalizes biochemical reactions. Ca²⁺ influx into spines initiates a cascade of biochemical signals leading to various forms of synaptic plasticity including long-term potentiation (LTP).Because LTP in hippocampal CA1 pyramidal neurons is a cellular mechanism that may underlie long-term memory formation, the signal transduction underlying LTP has been extensively studied by pharmacological and genetic methods (Bliss and Collingridge 1993; Derkach et al. 2007). It is now well established that LTP is induced by Ca²⁺ influx into dendritic spines through NMDA-type glutamate receptors (NMDARs), which induces the insertion of AMPA-type glutamate receptors (AMPARs) into the synapse, thereby increasing the sensitivity of the postsynaptic terminal to glutamate (Derkach et al. 2007; Kessels and Malinow 2009). An increase of release probability during LTP has also been reported (Enoki et al. 2009), and thus both pre- and postsynaptic mechanisms may contribute to LTP (Lisman and Raghavachari 2006).Manipulations of signal transduction using specific pharmacological inhibitors or genetic perturbations have identified many signaling pathways that connect Ca²⁺ to LTP induction. For example, LTP requires the activation of many signaling proteins, including Ca²⁺/calmodulin-dependent kinase II (CaMKII), extracellular signal-related kinase (ERK), Phoshoinositide 3 kinase (PI3K), protein kinase A and C, and GTPases such as Ras, Rab, and Rho (Kennedy et al. 2005). The list is continually growing, and the hundreds of implicated proteins form a complex signaling network whose contribution to LTP is still unclear (Bromberg et al. 2008).Signaling dynamics in neurons have traditionally been measured using biochemical analyses (Bromberg et al. 2008). However, the spatiotemporal resolution of conventional biochemistry is limited, restricting analysis to the time scale of many minutes and requiring the homogenization of tissue containing millions of synapses and other cellular elements. Furthermore, resolving synaptically induced changes in signaling by biochemical analysis typically requires stimulating many synapses at the same time, which may produce unintended effects, for instance, excitotoxicity or homeostatic plasticity.The size of dendritic spines is similar to the resolution of an optical microscope, permitting the optical analysis of biochemical signaling in each dendritic spine (Svoboda and Yasuda 2006). In particular, the advent of two-photon-based FRET techniques and the development of appropriate fluorescent reporters of specific biochemical reactions (see below) have provided readouts for signal transduction with high spatiotemporal resolution in live brain tissue (Svoboda and Yasuda 2006; Yasuda 2006). This has provided detailed information about the dynamics of signal transduction in spines and dendrites, and insights into the molecular mechanisms of synaptic plasticity.     

States of high conductance in a large-scale model of the visual cortex

This paper reports on the consequences of large, activity dependent, synaptic conductances for neurons in a large-scale neuronal network model of the input layer 4C of the Macaque primary visual cortex (Area V1). This high conductance state accounts for experimental observations about orientation selectivity, dynamics, and response magnitude (D. McLaughlin et al. (2000) Proc. Natl. Acad. Sci. USA 97: 8087–8092), and the linear dependence of Simple cells on visual stimuli (J. Wielaard et al. (2001) J. Neuroscience 21: 5203–5211). The source of large conductances in the model can be traced to inhibitory corticocortical synapses, and the model's predictions of large conductance changes are consistent with recent intracellular measurements (L. Borg-Graham et al. (1998) Nature 393: 369–373; J. Hirsch et al. (1998) J. Neuroscience 15: 9517–9528; J.S. Anderson et al. (2000) J. Neurophysiol. 84: 909–926). During visual stimulation, these conductances are large enough that their associated time-scales become the shortest in the model cortex, even below that of synaptic interactions. One consequence of this activity driven separation of time-scales is that a neuron responds very quickly to temporal changes in its synaptic drive, with its intracellular membrane potential tracking closely an effective reversal potential composed of the instantaneous synaptic inputs. From the effective potential and large synaptic conductance, the spiking activity of a cell can be expressed in an interesting and simplified manner, with the result suggesting how accurate and smoothly graded responses are achieved in the model network. Further, since neurons in this high-conductance state respond quickly, they are also good candidates as coincidence detectors and burst transmitters.     

Breaking down to build up: Neuroligin’s C-terminal domain strengthens the synapse

The mechanisms by which neuroligin adhesion molecules modulate synaptic plasticity remain unclear. In this issue, Liu et al. (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201509023) demonstrate that neuroligin 1 promotes actin assembly associated with synaptic strengthening independent of adhesion, suggesting additional ways for neuroligins to contribute to neuronal development and disease pathology.Spines are actin-enriched dendritic protrusions that serve as the major site of excitatory neurotransmission, underlying learning and memory formation (Lynch et al., 2007). Spines associate with presynaptic axon terminals through diverse adhesion molecules to form synapses (Siddiqui and Craig, 2011). Dynamic rearrangements of these synaptic adhesions and of the underlying actin cytoskeleton lead to either strengthening or weakening of particular synaptic connections. Synaptic strengthening, or long-term potentiation (LTP), is initiated by excitation of glutamate N-methyl-d-aspartate (NMDA) receptors, which promotes cleavage of synaptic adhesion molecules and disassembly of actin filaments (Lynch et al., 2007). Actin disassembly is mediated in part by recruitment of the actin-severing protein cofilin into the spine (Bosch et al., 2014). After the breakdown of the existing synaptic architecture, the actin cytoskeleton is stabilized again via Rac1-driven actin polymerization (Rex et al., 2009) and phosphorylation-mediated cofilin inactivation (Bosch et al., 2014). In parallel, recruitment and anchoring of synaptic adhesion molecules, including neuroligin 1 (NLG1; Schapitz et al., 2010) and glutamate receptors, increases the size of the postsynaptic signaling scaffold (PSD) across from the presynaptic terminal. In the final stage of LTP, the changes in synaptic morphology are consolidated by stabilization of actin filaments through actin capping and cross-linking together with the insertion of newly synthesized synaptic proteins (Lynch et al., 2007). Although the different steps of LTP shaping spine morphology and stability are generally understood, the signaling events that coordinate the initial disassembly of the existing synaptic architecture with reassembly of a stronger synaptic connection remain unclear.Neuroligins (NLGs) are a family of four transmembrane postsynaptic adhesion molecules (NLGs 1–4) that form heterotypic adhesions with presynaptic neurexins via an extracellular acetylcholinesterase-like domain (Südhof, 2008). Of the four NLG family members, NLG1 localizes predominantly to excitatory glutamatergic synapses (Song et al., 1999). Both in vitro and in vivo evidence demonstrate that the NLG–neurexin binding interaction is sufficient to promote synapse formation (Südhof, 2008; Chen et al., 2010). However, NLG knockout mice exhibit normal spine density but impaired synaptic transmission, suggesting that NLGs may regulate synaptic function independent of adhesion (Südhof, 2008). In addition to trans-synaptic adhesion mediated by the extracellular domain of NLGs, their short intracellular C-terminal domain (CTD) contains a PDZ binding domain (PBD) that facilitates binding and recruitment of postsynaptic density scaffold proteins, such as PSD95 (Irie et al., 1997; Dresbach et al., 2004). NLG1 is cleaved in an activity-dependent manner, leading to the release of an extracellular fragment that destabilizes synaptic adhesion and of the intracellular CTD (Suzuki et al., 2012).In this issue, Liu et al. focused on how activity-dependent cleavage of NLG1 and the subsequent release of its CTD affect actin organization and spine stability at excitatory synapses. They first observed that NLG1 knockout mouse brains, as well as cultured neurons infected with an shRNA targeting NLG1, exhibit decreased cofilin-S3 phosphorylation when compared with wild-type levels. Cofilin-S3 phosphorylation functions as a marker of mature dendritic spines, as cofilin inactivation results in F-actin assembly and is associated with the later stages of LTP (Calabrese et al., 2014). In addition, the absence of NLG1 prevented dynamic regulation of cofilin phosphorylation in response to KCl-induced neuronal excitation of brain slices, suggesting that cofilin phosphorylation depends on NLG1 both basally and in an activity-dependent manner. Remarkably, incubation with recombinant NLG1-CTD increased spine-associated cofilin phosphorylation in cultured neurons and rescued cofilin phosphorylation in NLG1 knockout mouse brain slices. Using full-length or truncated NLG1 constructs with a wild-type or mutated PDB sequence, Liu et al. (2016) demonstrated that NLG1-induced cofilin phosphorylation depends on both NLG1 cleavage and an intact PBD sequence within the released CTD. As the NLG1-CTD alone induced spine-associated cofilin phosphorylation, the researchers investigated its impact on actin assembly associated with synapse formation and function. In cultured neurons, recombinant NLG1-CTD increased F-actin levels together with spine and synapse formation. Similarly, intravenous injection of NLG1-CTD increased spine density in the CA1 region of the mouse hippocampus. This increased spine and synapse formation resulted in a corresponding increase in the frequency of excitatory postsynaptic currents, which was inhibited by a peptide that blocked cofilin phosphorylation. Together, these results establish that the NLG1-CTD requires cofilin phosphorylation to strengthen synaptic connections, prompting Liu et al. (2016) to investigate the mechanism underlying NLG1-induced cofilin phosphorylation.SPAR is a known regulator of the actin cytoskeleton that is hypothesized to bind to NLG1 (Craig and Kang, 2007). Using brain lysates and HEK293 cells expressing both NLG1 and SPAR, Liu et al. (2016) demonstrated that SPAR interacts with NLG1-CTD via its PBD domain. In brain slices, KCl-mediated excitation, which induces proteolytic cleavage of endogenous NLG1, increased the association of NLG1 and SPAR, suggesting that the interaction occurs in response to activity-dependent release of an intracellular CTD. To test whether this interaction regulates cofilin phosphorylation, Liu et al. (2016) expressed SPAR in HEK293 cells, where it decreased cofilin-S3 phosphorylation. However, incubation with a recombinant NLG1-CTD containing an intact PBD restored cofilin phosphorylation, demonstrating that this interaction alleviates SPAR-mediated repression of cofilin phosphorylation. In neurons, NLG1-CTD reduced the levels of synaptic SPAR, as assessed by both immunofluorescence and Western blotting of purified synaptosomes. SPAR is known to negatively regulate Rap1 signaling, and Rap1 signaling is important for Rac1 activation and spine morphogenesis (Pak et al., 2001; Maillet et al., 2003). In cultured neurons, a Rap1 inhibitor prevented NLG1-CTD–induced cofilin phosphorylation, whereas treatment with recombinant NLG1-CTD without Rap1 inhibition activated Rac1 signaling, leading to phosphorylation of its downstream targets, LIMK1 and cofilin. The results demonstrate that the CTD of NLG1 binds and displaces SPAR from the spine, alleviating its inhibition on Rap1 signaling. In turn, increased Rap1 signaling promotes Rac1 activation, leading to LIMK-1 and cofilin phosphorylation (Fig. 1). Lastly, these NLG-driven changes in actin assembly were found to simultaneously inhibit long-term depression, an activity-dependent reduction in the efficacy of synapses, and facilitate LTP, as determined by whole-cell patch clamping of brain slices incubated with NLG1-CTD.Open in a separate windowFigure 1.NLG’s CTD strengthens the synapse from within through dynamic actin remodeling. Excitatory activation of NMDA receptors (NMDAR) results in sequential cleavage of NLG1 (Suzuki et al., 2012). Liu et al. (2016) describe how the CTD of NLG1 interacts with SPAR, a negative regulator of Rap GTPase activity. This activity-dependent interaction displaces SPAR and alleviates the local inhibition of Rap activity within the dendritic spine. Rap drives a corresponding increase in Rac activation and phosphorylation of its downstream target, the actin regulator cofilin, thereby increasing F-actin filament assembly within spines. These changes in actin organization ultimately result in increased spine density and promote LTP.This work provides important insights into the mechanism by which NLG1 impacts synapse development and function by highlighting a critical role for SPAR in the regulation of actin assembly mediating synaptic strengthening. Interestingly, the temporal delay between the release of the NLG1-CTD and the subsequent sequestration of SPAR from the PSD could serve to distinguish an early disassembly phase following excitatory stimulation from later LTP consolidation, which is known to rely on both Rac1 activation (Rex et al., 2009) and cofilin phosphorylation (Bosch et al., 2014). Furthermore, it will be of interest to determine whether NLG1’s CTD affects the localization of other proteins known to bind its PBD, such as PSD95 (Irie et al., 1997), and whether these dynamic rearrangements at the postsynaptic scaffold also serve to simultaneously promote actin assembly while alleviating SPAR-mediated negative regulation of actin remodeling. For example, NLG1 has been shown to interact with Kalirin-7 (Owczarek et al., 2015), an activator of Rac1 that binds to PSD95 at the synapse; however, binding to PSD95 reduces Kalirin-7–mediated activation of Rac1 (Penzes et al., 2001). It is therefore attractive to speculate that the activity-dependent release of protein fragments, such as the CTD of NLG1, might alter postsynaptic density interactions that further promote localized Rac1-driven F-actin assembly. Consistent with this hypothesis, adhesion disassembly triggered by the extracellular domain of NLG1’s binding partner (β-neurexin) increases Rac1 activation (Owczarek et al., 2015). Ultimately, more work is necessary to determine how the strengthening effects of the intracellular CTD compete with the destabilizing effects of the extracellular domain (Suzuki et al., 2012). Recent research demonstrates that CAMKII phosphorylates and increases NLG1 surface expression in response to NMDA receptor activation (Bemben et al., 2014). If this phosphorylation event protects NLG1 from cleavage, it could serve to stabilize an adhesive pool of NLG1 while allowing for the release of the CTD from an unprotected population. Alternatively, this phosphorylation event could serve to recruit new NLG1 proteins to the synapse later in the LTP process when adhesions are reestablished. Further research is needed to understand how the adhesive and intracellular signaling capabilities of NLG1 are balanced at discrete stages of synaptic plasticity, and in particular how phosphorylation of NLG1 regulates both its surface expression as well as its cleavage.Consistent with the multiple roles of NLGs in modulating synaptic architecture, it is not surprising that NLG mutations have been implicated in diverse cognitive and neurodevelopmental disorders, such as Alzheimer’s disease and autism (Südhof, 2008; Tristán-Clavijo et al., 2015). In light of this study, it will be interesting to determine how disease-associated NLG mutations contribute to both synaptic adhesion as well as stabilization of the actin cytoskeleton that supports synaptic strengthening. This is particularly important because both Alzheimer’s disease and autism-associated NLG mutant proteins exhibit decreased surface expression (Chubykin et al., 2005; Tristán-Clavijo et al., 2015), although the autism-associated mutant NLG proteins present at the cell surface still promote synapse formation (Chubykin et al., 2005). However, the decreased postsynaptic NLG pool could impair subsequent activity-dependent synaptic strengthening. Likewise, understanding whether binding of the postsynaptic scaffolding protein Shank3 to the CTD of NLG1 (Arons et al., 2012) affects NLG1 cleavage could provide insights into the mechanism by which Shank3 affects activity-dependent synaptic remodeling in autism pathogenesis. The work by Liu et al. (2016), demonstrating that adhesion disassembly coordinates subsequent actin assembly underlying synaptic strengthening, takes an important step toward shedding light on the altered synaptic plasticity underlying both complex neurodevelopmental and neurodegenerative pathologies.     

Surface dynamics of GluN2B‐NMDA receptors controls plasticity of maturing glutamate synapses

NMDA‐type glutamate receptors (NMDAR) are central actors in the plasticity of excitatory synapses. During adaptive processes, the number and composition of synaptic NMDAR can be rapidly modified, as in neonatal hippocampal synapses where a switch from predominant GluN2B‐ to GluN2A‐containing receptors is observed after the induction of long‐term potentiation (LTP). However, the cellular pathways by which surface NMDAR subtypes are dynamically regulated during activity‐dependent synaptic adaptations remain poorly understood. Using a combination of high‐resolution single nanoparticle imaging and electrophysiology, we show here that GluN2B‐NMDAR are dynamically redistributed away from glutamate synapses through increased lateral diffusion during LTP in immature neurons. Strikingly, preventing this activity‐dependent GluN2B‐NMDAR surface redistribution through cross‐linking, either with commercial or with autoimmune anti‐NMDA antibodies from patient with neuropsychiatric symptoms, affects the dynamics and spine accumulation of CaMKII and impairs LTP. Interestingly, the same impairments are observed when expressing a mutant of GluN2B‐NMDAR unable to bind CaMKII. We thus uncover a non‐canonical mechanism by which GluN2B‐NMDAR surface dynamics plays a critical role in the plasticity of maturing synapses through a direct interplay with CaMKII.