Dendritic Spine Heads Research Articles

Optogenetic inhibition of cortical afferents in the nucleus accumbens simultaneously prevents cue-induced transient synaptic potentiation and cocaine-seeking behavior.

Animal models of relapse reveal that the motivation to seek drug is regulated by enduring morphological and physiological changes in the nucleus accumbens, as well as transient synaptic potentiation in the accumbens core (NAcore) that parallels drug-seeking behavior. The current study sought to examine the link between the behavioral and synaptic consequences of cue-induced cocaine seeking by optically silencing glutamatergic afferents to the NAcore from the prelimbic cortex (PL). Adeno-associated virus coding for the inhibitory opsin archaerhodopsin was microinjected into PL, and optical fibers were targeted to NAcore. Animals were trained to self-administer cocaine followed by extinction training, and then underwent cue-induced reinstatement in the presence or absence of 15 min of optically induced inhibition of PL fibers in NAcore. Inhibiting the PL-to-NAcore projection blocked reinstated behavior and was paralleled by decreased dendritic spine head diameter and AMPA/NMDA ratio relative to sham-laser control rats. Interestingly, while spine density was elevated after extinction training, no further effects were observed by cued reinstatement or optical inhibition. These findings validate the critical role for PL afferents to the NAcore in simultaneously regulating both reinstated behavior and the associated transient synaptic potentiation.

Protein kinase C is essential for kainate-induced anxiety-related behavior and glutamatergic synapse upregulation in prelimbic cortex.

Anxiety is one of common mood disorders, in which the deficit of serotonergic and GABAergic synaptic functions in the amygdala and prefrontal cortex is believed to be involved. The pathological changes at the glutamatergic synapses and neurons in these brain regions as well as their underlying mechanisms remain elusive, which we aim to investigate. An agonist of kainate-type glutamate receptors, kainic acid, was applied to induce anxiety-related behaviors. The morphology and functions of glutamatergic synapses in the prelimbic region of mouse prefrontal cortex were analyzed using cellular imaging and electrophysiology. After kainate-induced anxiety is onset, the signal transmission at the glutamatergic synapses is upregulated, and the dendritic spine heads are enlarged. In terms of the molecular mechanisms, the upregulated synaptic plasticity is associated with the expression of more protein kinase C (PKC) in the dendritic spines. Chelerythrine, a PKC inhibitor, reverses kainate-induced anxiety and anxiety-related glutamatergic synapse upregulation. The activation of glutamatergic kainate-type receptors leads to anxiety-related behaviors and glutamatergic synapse upregulation through protein kinase C in the prelimbic region of the mouse prefrontal cortex.

Myosin IIb controls actin dynamics underlying the dendritic spine maturation.

Precise control of the formation and development of dendritic spines is critical for synaptic plasticity. Consequently, abnormal spine development is linked to various neurological disorders. The actin cytoskeleton is a structural element generating specific changes in dendritic spine morphology. Although mechanisms underlying dendritic filopodia elongation and spine head growth are relatively well understood, it is still not known how spine heads are enlarged and stabilized during dendritic spine maturation. By using rat hippocampal neurons, we demonstrate that the size of the stable actin pool increases during the neuronal maturation process. Simultaneously, the treadmilling rate of the dynamic actin pool increases. We further show that myosin IIb controls dendritic spine actin cytoskeleton by regulating these two different pools of F-actin via distinct mechanisms. The findings indicate that myosin IIb stabilizes the stable F-actin pool through actin cross-linking. Simultaneously, activation of myosin IIb contractility increases the treadmilling rate of the dynamic pool of actin. Collectively, these data show that myosin IIb has a major role in the regulation of actin filament stability in dendritic spines, and elucidate the complex mechanism through which myosin IIb functions in this process. These new insights into the mechanisms underlying dendritic spine maturation further the model of dendritic spine morphogenesis.

Cell type-specific expression of Eps8 in the mouse hippocampus

BackgroundEpidermal growth factor receptor substrate 8 (Eps8) is a multifunctional protein that regulates actin cytoskeleton dynamics and architecture through its barbed-end capping and bundling activities. In cultured hippocampal neurons, Eps8 is enriched at dendritic spine heads and is required for spine morphogenesis; however, the detailed expression pattern of Eps8 in the hippocampus has not yet been explored.ResultsHere, we demonstrate that endogenous Eps8 protein is restrictively expressed in neurons (NeuN-positive), but not in glial cells (glial fibrillary acidic protein-positive) in area CA1 of the mouse hippocampus. Surprisingly, Eps8 immunoreactivity is rarely found in pyramidal cell somata, but is expressed predominantly in the somata and dendrites of 67 kDa isoform of glutamic acid decarboxylase-expressing GABAergic interneurons in the stratum radiatum and at the border of stratum radiatum and lacunosum-moleculare of area CA1. On the basis of co-localizing markers, we found that Eps8 is not present in perisomatic inhibitory parvalbumin-expressing cells or calretinin-expressing interneurons. However, Eps8 is richly expressed in calbindin-expressing interneurons. Furthermore, Eps8 is also present in cholecystokinin-expressing interneurons, but not in somatostatin-expressing interneurons in area CA1 stratum pyramidale and stratum radiatum.ConclusionsThese results reveal a previously unknown cell type-specific expression pattern of endogenous Eps8 protein in the mouse hippocampus and speculate that the role of Eps8 in controlling and orchestrating neuronal morphogenesis and structural plasticity might be more prominent in interneurons than in pyramidal cells of the hippocampus.

Moderate AMPA receptor clustering on the nanoscale can efficiently potentiate synaptic current

The prevailing view at present is that postsynaptic expression of the classical NMDA receptor-dependent long-term potentiation relies on an increase in the numbers of local AMPA receptors (AMPARs). This is thought to parallel an expansion of postsynaptic cell specializations, for instance dendritic spine heads, which accommodate synaptic receptor proteins. However, glutamate released into the synaptic cleft can normally activate only a hotspot of low-affinity AMPARs that occur in the vicinity of the release site. How the enlargement of the AMPAR pool is causally related to the potentiated AMPAR current remains therefore poorly understood. To understand possible scenarios of postsynaptic potentiation, here we explore a detailed Monte Carlo model of the typical small excitatory synapse. Simulations suggest that approximately 50% increase in the synaptic AMPAR current could be provided by expanding the existing AMPAR pool at the expense of 100–200% new AMPARs added at the same packing density. Alternatively, reducing the inter-receptor distances by only 30–35% could achieve a similar level of current potentiation without any changes in the receptor numbers. The NMDA receptor current also appears sensitive to the NMDA receptor crowding. Our observations provide a quantitative framework for understanding the ‘resource-efficient’ ways to enact use-dependent changes in the architecture of central synapses.

Matrix Metalloproteinases Regulate the Formation of Dendritic Spine Head Protrusions during Chemically Induced Long-Term Potentiation

Dendritic spines are are small membranous protrusions that extend from neuronal dendrites and harbor the majority of excitatory synapses. Increasing evidence has shown that matrix metalloproteinases (MMPs), a family of extracellularly acting and Zn2+-dependent endopeptidases, are able to rapidly modulate dendritic spine morphology. Spine head protrusions (SHPs) are filopodia-like processes that extend from the dendritic spine head, representing a form of postsynaptic structural remodeling in response to altered neuronal activity. Herein, we show that chemically induced long-term potentiation (cLTP) in dissociated hippocampal cultures upregulates MMP-9 activity that controls the formation of SHPs. Blocking of MMPs activity or microtubule dynamics abolishes the emergence of SHPs. In addition, autoactive recombinant MMP-9, promotes the formation of SHPs in organotypic hippocampal slices. Furthermore, spines with SHPs gained postsynaptic α-amino-3-hydroxyl-5-methyl-4-isoxazole propionic acid (AMPA) receptors upon cLTP and the synaptic delivery of AMPA receptors was controlled by MMPs. The present results strongly imply that MMP-9 is functionally involved in the formation of SHPs and the control of postsynaptic receptor distribution upon cLTP.

Reinstatement of nicotine seeking is mediated by glutamatergic plasticity

Nicotine abuse and addiction is a major health liability. Nicotine, an active alkaloid in tobacco, is self-administered by animals and produces cellular adaptations in brain regions associated with drug reward, such as the nucleus accumbens. However, it is unknown whether, akin to illicit drugs of abuse such as cocaine or heroin, the adaptations endure and contribute to the propensity to relapse after discontinuing nicotine use. Using a rat model of cue-induced relapse, we made morphological and electrophysiological measures of synaptic plasticity, as well as quantified glutamate overflow, in the accumbens after 2 wk of withdrawal with extinction training. We found an enduring basal increase in dendritic spine head diameter and in the ratio of AMPA to NMDA currents in accumbens spiny neurons compared with yoked saline animals at 2 wk after the last nicotine self-administration session. This synaptic potentiation was associated with an increase in both AMPA (GluA1) and NMDA (GluN2A and GluN2B) receptor subunits, and a reduction in the glutamate transporter-1 (GLT-1). When nicotine seeking was reinstated by presentation of conditioned cues, there were parallel increases in behavioral responding, extracellular glutamate, and further increases in dendritic spine head diameter and ratio of AMPA to NMDA currents within 15 min. These findings suggest that targeting glutamate transmission might inhibit cue-induced nicotine seeking. In support of this hypothesis, we found that pharmacological inhibition of GluN2A with 3-Chloro-4-fluoro-N-[4-[[2-(phenylcarbonyl)hydrazino]carbonyl]benzyl]benzenesulfonamide (TCN-201) or GluN2B with ifenprodil abolished reinstated nicotine seeking. These results indicate that up-regulated GluN2A, GluN2B, and rapid synaptic potentiation in the accumbens contribute to cue-induced relapse to nicotine use.

Enhanced Recruitment of Endosomal Na+/H+Exchanger NHE6 into Dendritic Spines of Hippocampal Pyramidal Neurons during NMDA Receptor-Dependent Long-Term Potentiation

Postsynaptic endosomal trafficking has emerged as a principal regulatory mechanism of structural and functional plasticity of glutamatergic synapses. Recycling endosomes perform activity-dependent transport of AMPA receptors (AMPARs) and lipids to the postsynaptic membrane, activities that are known to contribute to long-term synaptic potentiation and hypothesized to subserve learning and memory processes in the brain. Recently, genetic defects in a widely expressed vesicular pH-regulating transporter, the Na(+)/H(+) exchanger NHE6 isoform, have been implicated in neurodevelopmental disorders including severe X-linked mental retardation and autism. However, little information is available regarding the cellular properties of this transporter in the CNS. Here, we show by quantitative light microscopy that the protein abundance of NHE6 is developmentally regulated in area CA1 of the mouse hippocampus. Within pyramidal neurons, NHE6 was found to localize to discrete puncta throughout the soma and neurites, with noticeable accumulation at dendritic spines and presynaptic terminals. Dual immunolabeling of dendritic spines revealed that NHE6 partially colocalizes with typical markers of early and recycling endosomes as well as with the AMPAR subunit GluA1. Significantly, NHE6-containing vesicles exhibited enhanced translocation to dendritic spine heads during NMDA receptor (NMDAR)-dependent long-term potentiation. These data suggest that NHE6 may play a unique, previously unrecognized, role at glutamatergic synapses that are important for learning and memory.

Cortical Dendritic Spine Heads Are Not Electrically Isolated by the Spine Neck from Membrane Potential Signals in Parent Dendrites

The evidence for an important hypothesis that cortical spine morphology might participate in modifying synaptic efficacy that underlies plasticity and possibly learning and memory mechanisms is inconclusive. Both theory and experiments suggest that the transfer of excitatory postsynaptic potential signals from spines to parent dendrites depends on the spine neck morphology and resistance. Furthermore, modeling of signal transfer in the opposite direction predicts that synapses on spine heads are not electrically isolated from voltages in the parent dendrite. In sharp contrast to this theoretical prediction, one of a very few measurements of electrical signals from spines reported that slow hyperpolarizing membrane potential changes are attenuated considerably by the spine neck as they spread from dendrites to synapses on spine heads. This result challenges our understanding of the electrical behavior of spines at a fundamental level. To re-examine the specific question of the transfer of dendritic signals to synapses of spines, we took advantage of a high-sensitivity Vm-imaging technique and carried out optical measurements of electrical signals from 4 groups of spines with different neck length and simultaneously from parent dendrites. The results show that spine neck does not filter membrane potential signals as they spread from the dendrites into the spine heads.

A mathematical model of the tripartite synapse: astrocyte-induced synaptic plasticity

In this paper, we present a biologically detailed mathematical model of tripartite synapses, where astrocytes modulate short-term synaptic plasticity. The model consists of a pre-synaptic bouton, a post-synaptic dendritic spine-head, a synaptic cleft and a peri-synaptic astrocyte controlling Ca(2 + ) dynamics inside the synaptic bouton. This in turn controls glutamate release dynamics in the cleft. As a consequence of this, glutamate concentration in the cleft has been modeled, in which glutamate reuptake by astrocytes has also been incorporated. Finally, dendritic spine-head dynamics has been modeled. As an application, this model clearly shows synaptic potentiation in the hippocampal region, i.e., astrocyte Ca(2 + ) mediates synaptic plasticity, which is in conformity with the majority of the recent findings (Perea and Araque (Science 317, 1083-1086, 2007); Henneberger et al. (Nature 463, 232-236, 2010); Navarrete et al. (PLoS Biol. 10, e1001259, 2012)).

N-Methyl-d-aspartate (NMDA) Receptor Composition Modulates Dendritic Spine Morphology in Striatal Medium Spiny Neurons

Dendritic spines of medium spiny neurons represent an essential site of information processing between NMDA and dopamine receptors in striatum. Even if activation of NMDA receptors in the striatum has important implications for synaptic plasticity and disease states, the contribution of specific NMDA receptor subunits still remains to be elucidated. Here, we show that treatment of corticostriatal slices with NR2A antagonist NVP-AAM077 or with NR2A blocking peptide induces a significant increase of spine head width. Sustained treatment with D1 receptor agonist (SKF38393) leads to a significant decrease of NR2A-containing NMDA receptors and to a concomitant increase of spine head width. Interestingly, co-treatment of corticostriatal slices with NR2A antagonist (NVP-AAM077) and D1 receptor agonist augmented the increase of dendritic spine head width as obtained with SKF38393. Conversely, NR2B antagonist (ifenprodil) blocked any morphological effect induced by D1 activation. These results indicate that alteration of NMDA receptor composition at the corticostriatal synapse contributes not only to the clinical features of disease states such as experimental parkinsonism but leads also to a functional and morphological outcome in dendritic spines of medium spiny neurons.

Glial glutamate transport modulates dendritic spine head protrusions in the hippocampus

Accumulating evidence supports the idea that synapses are tripartite, whereby perisynaptic astrocytes modulate both pre- and postsynaptic function. Although some of these features have been uncovered by using electrophysiological methods, less is known about the structural interplay between synapses and glial processes. Here, we investigated how astrocytes govern the plasticity of individual hippocampal dendritic spines. Recently, we uncovered that a subgroup of innervated dendritic spines is able to undergo remodeling by extending spine head protrusions (SHPs) toward neighboring functional presynaptic boutons, resulting in new synapses. Although glutamate serves as a trigger, how this behavior is regulated is unknown. As astrocytes control extracellular glutamate levels through their high-affinity uptake transporters, together with their privileged access to synapses, we investigated a role for astrocytes in SHP formation. Using time-lapse confocal microscopy, we found that the volume overlap between spines and astrocytic processes decreased during the formation of SHPs. Focal application of glutamate also reduced spine-astrocyte overlap and induced SHPs. Importantly, SHP formation was prevented by blocking glial glutamate transporters, suggesting that glial control of extracellular glutamate is important for SHP-mediated plasticity of spines. Hence, the dynamic changes of both spines and astrocytes can rapidly modify synaptic connectivity.

Nanoscale imaging of dendritic spines

Stimulated emission depletion (STED) microscopy has an extremely high (nanoscale) resolution, but this technique has not been applied in vivo in rodents. Here, the authors used STED microscopy to study neuron dynamics in vivo in the somatosensory cortex of adult mice. The images showed structures as small as <70 nm. Moreover, serial imaging revealed morphological changes and movement in the head and neck regions of dendritic spines over minutes. STED and other forms of nanoscopy could be used to investigate spine dynamics in brain development and function as well as in disease processes.

Pathology of Synapses and Dendritic Spines

Excitatory synapses represent the sites in which axons make a functional contact with their target neurons and they are typically located at the head of dendritic spines. Due to their critical role as mediators of interneuronal interactions, insults to synapses may result with significant clinical manifestations such as dementia or movement disorders. This special issue of Neural Plasticity discusses various aspects of synapse and spine pathology associated with Alzheimer's disease, perinatal asphyxia, and neuropathic pain. Synapse and spine loss may represent early and profound neuropathological changes potentially underlying cognitive deficits in Alzheimer's disease (AD). In fact, synapse loss is the strongest pathological correlate of dementia in AD. The inherent plasticity of synapses makes them an attractive therapeutic target. W. Yu and B. Lu, “Synapses and dendritic spines as pathogenic targets in Alzheimer's disease,” review the important field of synapse and dendritic loss in AD. In their paper, the authors discuss the well-established role of oligomeric amyloid beta in causing synaptic dysfunction and loss through signaling mechanisms associated with long-term depression. They also explore the exciting new relationship between amyloid beta and tau at the postsynaptic density. Several recent studies have placed tau in dendritic spines—a surprising place for a protein that dogma places firmly on microtubules in axons—where it is turning out to mediate amyloid-beta-induced synaptic dysfunction and loss. In another review T. Spires-Jones and S. Knafo, “Spines, plasticity, and cognition in Alzheimer's model mice,” provide a comprehensive analysis on recent works describing the morphological, synaptic, and behavioral characteristics of the different transgenic models of AD. Results from various models and in variety of ages show a gradual deterioration in synaptic and cognitive functions. The accumulating evidence from transgenic models discussed in this review appears to support a model of AD pathogenesis in which oligomeric amyloid beta initiates synaptic dysfunction or degeneration and induces pathological changes in tau, leading to neuronal loss and ultimately to cognitive deficits. G. E. Saraceno et al., “Hippocampal dendritic spines modifications induced by perinatal asphyxia,” investigated the effect of perinatal asphyxia (PA) on the hippocampal postsynaptic density (PSD). They report an unexpected increased thickness and dispersed appearance of the PSD in the asphyctic brains. Correlative fluorescent and electron microscopy showed a decline of F-actin-stained spines in hippocampal excitatory synapses after the insult that may suggest that PA is harmful to the actin cytoskeleton. These data suggest that perinatal asphyxia may lead to long-term changes in hippocampal synapses. NMDA receptors are located at synapses and modulate various forms of synaptic plasticity. Grin1b gene encodes the postsynaptic NMDA receptor in zebrafish. RNA and various RNA products play critical roles in regulating gene expression and production of neuroactive proteins in the nervous system. P. Pozo and B. Hoopengardner, “Identification and characterization of two novel RNA editing sites in grin1b transcripts of embryonic Danio rerio,” identified two novel RNA editing events for the grin1b gene in zebrafish. This new information may have implications for transcriptional regulation of mammalian glutamate receptors, which play essential roles in neuronal transmission and plasticity, but also mediate neuronal toxicity in neurodegenerative disorders. S. K. Kim et al., “Synaptic structure and function in the mouse somatosensory cortex during chronic pain: in vivo two-photon imaging,” summarize recent developments in studying in vivo spine dynamics in thesomatosensory cortex of adult mice in a model of chronic neuropathic pain. Thearticle highlights theimportance of neural plasticity in pain research. Employingmultiphoton microscopy, they describe remarkably rapid synaptic remodeling inlayer 5 neurons of somatosensory cortexwithin days of the peripheral nerveinjury (partial sciatic nerve ligation). Peripheral nerve injury via peripheralhyperactivity causes a rapid rewiring of distinct somatosensory cortexsynapticconnections, leading to local somatosensory cortex hyperexcitability. Theauthors postulate that these local cortical changes in spine turnover followinginduction of neuropathicpain play an important role in chronic pain conditions. Shira Knafo Gunnar K. Gouras Xiao-Xin Yan Tara Spires-Jones

The neuropeptide PACAP38 induces dendritic spine remodeling through ADAM10/N-Cadherin signaling pathway

The neuropeptide pituitary adenylate cyclase-activating polypeptide 38 (PACAP38) has been implicated in the induction of synaptic plasticity at the excitatory glutamatergic synapse. In particular, recent studies have shown that it is involved in the regulation of N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor activation. Here we demonstrate the effect of PACAP38 on the modulation of dendritic spine morphology through a disintegrin and metalloproteinase 10 (ADAM10)-N-cadherin-AMPA receptor signaling pathway. Treatment of primary hippocampal neurons with PACAP38 induced an accumulation of ADAM10 at the postsynaptic membrane. This event led to a significant decrease of dendritic spine head width and to a concomitant reduction of GluR1 colocalization with postsynaptic markers. The PACAP38-induced effect on dendritic spine head width was prevented by either treatment with the ADAM10-specific inhibitor or transfection of a cleavage-defective N-cadherin construct mutated in the ADAM10 cleavage site. Overall, our findings reveal that PACAP38 is involved in the modulation of dendritic spine morphology in hippocampal neurons, and assign to the ADAM10-N-cadherin signaling pathway a crucial role in this modification of the excitatory glutamatergic synapse.

Essential Role for Vav Guanine Nucleotide Exchange Factors in Brain-Derived Neurotrophic Factor-Induced Dendritic Spine Growth and Synapse Plasticity

Brain-derived neurotrophic factor (BDNF) and its cognate receptor, TrkB, regulate a wide range of cellular processes, including dendritic spine formation and functional synapse plasticity. However, the signaling mechanisms that link BDNF-activated TrkB to F-actin remodeling enzymes and dendritic spine morphological plasticity remain poorly understood. We report here that BDNF/TrkB signaling in neurons activates the Vav family of Rac/RhoA guanine nucleotide exchange factors through a novel TrkB-dependent mechanism. We find that Vav is required for BDNF-stimulated Rac-GTP production in cortical and hippocampal neurons. Vav is partially enriched at excitatory synapses in the postnatal hippocampus but does not appear to be required for normal dendritic spine density. Rather, we observe significant reductions in both BDNF-induced, rapid, dendritic spine head growth and in CA3-CA1 theta burst-stimulated long-term potentiation in Vav-deficient mouse hippocampal slices, suggesting that Vav-dependent regulation of dendritic spine morphological plasticity facilitates normal functional synapse plasticity.

Anomalous diffusion imposed by dendritic spines (Commentary on Santamaria et al.)

An exciting decade of two-photon fluorescence imaging has revealed that the shape and turnover of dendritic spines are dynamic and modulated by synaptic plasticity (Matsuzaki et al., 2004). Such structural changes can be induced via focal glutamate uncaging at individual spines, but they can also be driven by experience (Holtmaat et al., 2006). Indeed, the size of both the head and neck of dendritic spines can be modulated by neuronal activity (Matsuzaki et al., 2004; Bloodgood & Sabatini, 2005). In turn, the geometry of spines will affect the compartmentalization of signals generated at the spine head and the extent to which ions and second messengers can spread via the neck to the parent dendrite and nearby spines. Interestingly, Santamaria et al. (2006) demonstrated in an earlier study that diffusion in spiny dendrites cannot be described by the standard diffusion equation. Instead, spines temporarily ‘trap’ molecules causing anomalous diffusion in the parent dendrite (Fig. 1). Uncaging dextran-conjugated fluorescent probes and imaging the fluorescence changes over time along a stretch of dendrite enabled calculation of the spatial variance of the diffusing molecules and estimation of the anomalous exponent dw. In smooth dendrites diffusion had an anomalous exponent of dw = 2, whereas in spiny dendrites dw turned out to be high (dw > 2) resulting in slowed diffusion. The importance of this finding is that the diffusion of second messengers within dendrites could be controlled by changes in spine shape or density. It is known that spine density in CA1 pyramidal neurons varies considerably across their dendritic arbors, with a progressive increase in spine density and spine size in the distal segments of apical dendrites (Konur et al., 2003). Thus, different dendritic compartments may exhibit different rates of diffusion of postsynaptic signaling molecules. In their study published in this issue of EJN, Santamaria et al. (2011) show that, in both Purkinje cells and CA1 pyramidal neurons, the anomalous exponent dw scales linearly with spine density. Anomalous diffusion in dendrites appears to be a hallmark of all neurons with spines. In addition, these authors provide empirical evidence that neither branching geometry nor dendrite diameter account for anomalous diffusion, confirming their previous theoretical models (Santamaria et al., 2006). The mechanistic studies by Santamaria et al. (2011) might be relevant to explain, for example, the functional impact of estradiol on memory formation (Andreescu et al., 2007) or of the pathological impact of a lack of fragile X mental retardation protein on motor behavior (Koekkoek et al., 2005). In both cases, altered geometry of spines and ⁄ or their inputs are likely to affect the diffusion of second messengers in the dendritic network. Therefore, it will be of general interest to identify the molecules for which anomalous diffusion plays a relevant role under physiological and pathophysiological conditions. For instance, active extrusion of calcium is likely to supersede the effects of spine geometry on diffusion of this ion in dendrites, whereas diffusion of inositol triphosphate (IP3) is slowed down (Santamaria et al., 2006) and that of GTPase signaling proteins varies among their family (Yasuda & Murakoshi, 2011). Anomalous diffusion imposed by spine geometry may thus represent a mechanism modulating the function of a selected set of signaling molecules.

Sexually dimorphic effects of estrogen on spines in cultures of accessory olfactory bulb

A sex difference has been reported in the responsiveness of the vomeronasal (VN) system to pheromones. In the present study, to clarify a direct and acute influence of 17β-estradiol (E2) on the accessory olfactory bulb (AOB) neurons, we investigated the effect of E2 on dendritic spines in cultured AOB cells derived from male and female neonatal rats. After 17–18 days in vitro (DIV), cultured AOB cells were transfected with GFP expression vectors. At 21–23 DIV, cells were treated with E2, and time-lapse images of transfected AOB neurons identified as granule cells were taken under a confocal laser scanning microscope for 3 h. The dendritic spine head area of granule cells was quantitatively evaluated, and spine heads were classified into larger (≥1 μm 2) and smaller (<1 μm 2) ones before E2-treatment (0 h). In cultured cells derived from both sexes, the larger spines were not significantly changed at 1, 2 and 3 h after E2-treatment. In contrast, E2-treatment significantly enlarged the head area of the smaller spines of granule cells derived from the female, whereas E2 did not cause any significant effects on those from the male. Our results provide evidence for the sexually-dimorphic effect of E2 on spine development in AOB granule cells.

Complementary synaptic distribution of enzymes responsible for synthesis and inactivation of the endocannabinoid 2-arachidonoylglycerol in the human hippocampus

Clinical and experimental evidence demonstrates that endocannabinoids play either beneficial or adverse roles in many neurological and psychiatric disorders. Their medical significance may be best explained by the emerging concept that endocannabinoids are essential modulators of synaptic transmission throughout the central nervous system. However, the precise molecular architecture of the endocannabinoid signaling machinery in the human brain remains elusive. To address this issue, we investigated the synaptic distribution of metabolic enzymes for the most abundant endocannabinoid molecule, 2-arachidonoylglycerol (2-AG), in the postmortem human hippocampus. Immunostaining for diacylglycerol lipase-α (DGL-α), the main synthesizing enzyme of 2-AG, resulted in a laminar pattern corresponding to the termination zones of glutamatergic pathways. The highest density of DGL-α-immunostaining was observed in strata radiatum and oriens of the cornu ammonis and in the inner third of stratum moleculare of the dentate gyrus. At higher magnification, DGL-α-immunopositive puncta were distributed throughout the neuropil outlining the immunonegative main dendrites of pyramidal and granule cells. Electron microscopic analysis revealed that this pattern was due to the accumulation of DGL-α in dendritic spine heads. Similar DGL-α-immunostaining pattern was also found in hippocampi of wild-type, but not of DGL-α knockout mice. Using two independent antibodies developed against monoacylglycerol lipase (MGL), the predominant enzyme inactivating 2-AG, immunostaining also revealed a laminar and punctate staining pattern. However, as observed previously in rodent hippocampus, MGL was enriched in axon terminals instead of postsynaptic structures at the ultrastructural level. Taken together, these findings demonstrate the post- and presynaptic segregation of primary enzymes responsible for synthesis and elimination of 2-AG, respectively, in the human hippocampus. Thus, molecular architecture of the endocannabinoid signaling machinery supports retrograde regulation of synaptic activity, and its similar blueprint in rodents and humans further indicates that 2-AG's physiological role as a negative feed-back signal is an evolutionarily conserved feature of excitatory synapses.

NMDA-Mediated Ca2+Influx Drives Aberrant Ryanodine Receptor Activation in Dendrites of Young Alzheimer's Disease Mice

Deficits in synaptic function, particularly through NMDA receptors (NMDARs), are linked to late-stage cognitive impairments in Alzheimer's disease (AD). At earlier disease stages, however, there is evidence for altered endoplasmic reticulum (ER) calcium signaling in human cases and in neurons from AD mouse models. Despite the fundamental importance of calcium to synaptic function, neither the extent of ER calcium dysregulation in dendrites nor its interaction with synaptic function in AD pathophysiology is known. Identifying the mechanisms underlying early synaptic calcium dysregulation in AD pathogenesis is likely a key component to understanding, and thereby preventing, the synapse loss and downstream cognitive impairments. Using two-photon calcium imaging, flash photolysis of caged glutamate, and patch-clamp electrophysiology in cortical brain slices, we examined interactions between synaptically and ER-evoked calcium release at glutamatergic synapses in young AD transgenic mice. We found increased ryanodine receptor-evoked calcium signals within dendritic spine heads, dendritic processes, and the soma of pyramidal neurons from 3xTg-AD and TAS/TPM AD mice relative to NonTg controls. In addition, synaptically evoked postsynaptic calcium responses were larger in the AD strains, as were calcium signals generated from NMDAR activation. However, calcium responses triggered by back-propagating action potentials were not different. Concurrent activation of ryanodine receptors (RyRs) with either synaptic or NMDAR stimulation generated a supra-additive calcium response in the AD strains, suggesting an aberrant calcium-induced calcium release (CICR) effect within spines and dendrites. We propose that presenilin-linked disruptions in RyR signaling and subsequent CICR via NMDAR-mediated calcium influx alters synaptic function and serves as an early pathogenic factor in AD.