Structural dynamics of dendritic spines: Molecular composition,geometry and functional regulation

The development of dendritic spines with specific geometry and membrane composition is critical for proper synaptic function. Specific spine membrane architecture, sub-spine microdomains and spine head and neck geometry allow for well-coordinated and compartmentalized signaling, disruption of which could lead to various neurological diseases. Research from neuronal cell culture, brain slices and direct in vivo imaging indicates that dendritic spine development is a dynamic process which includes transition from small dendritic filopodia through a series of structural refinements to elaborate spines of various morphologies. Despite intensive research, the precise coordination of this morphological transition, the changes in molecular composition, and the relation of spines of various morphologies to function remain a central enigma in the development of functional neuronal circuits. Here, we review research so far and aim to provide insight into the key events that drive structural change during transition from immature filopodia to fully functional spines and the relevance of spine geometry to function.     

Calcium influx and postsynaptic proteins coordinate the dendritic filopodium‐spine transition

Dendritic spines are the major locations of excitatory synapses in the mammalian central nervous system. The transformation from dendritic filopodia to dendritic spines has been recognized as one type of spinogenesis. For instance, syndecan‐2 (SDC2), a synaptic heparan sulfate proteoglycan, is highly concentrated at dendritic spines and required for spinogenesis. It induces dendritic filopodia formation, followed by spine formation. However, the molecular regulation of the filopodium‐spine transition induced by SDC2 is still unclear. In this report, we show that calcium is an important signal downstream of SDC2 in regulation of filopodium‐spine transition but not filopodia formation. SDC2 interacted with the postsynaptic proteins calmodulin‐dependent serine kinase (CASK) and LIN7 and further recruited NMDAR to the tips of filopodia induced by SDC2. Calcium influx via NMDAR promoted spine maturation because addition of EGTA or AP5 to the culture medium effectively prevented morphological change from dendritic filopodia to dendritic spines. Our data also indicated that F‐actin rearrangement regulated by calcium influx is involved in the morphological change, because the knockdown of gelsolin, a calcium‐activated F‐actin severing molecule, impaired the filopodium‐spine transition induced by SDC2. In conclusion, our study demonstrates that postsynaptic proteins coordinate to trigger calcium signalling and cytoskeleton rearrangement and consequently control filopodium‐spine transition. © 2014 Wiley Periodicals, Inc. Develop Neurobiol 74: 1011–1029, 2014     

Electrical advantages of dendritic spines

Many neurons receive excitatory glutamatergic input almost exclusively onto dendritic spines. In the absence of spines, the amplitudes and kinetics of excitatory postsynaptic potentials (EPSPs) at the site of synaptic input are highly variable and depend on dendritic location. We hypothesized that dendritic spines standardize the local geometry at the site of synaptic input, thereby reducing location-dependent variability of local EPSP properties. We tested this hypothesis using computational models of simplified and morphologically realistic spiny neurons that allow direct comparison of EPSPs generated on spine heads with EPSPs generated on dendritic shafts at the same dendritic locations. In all morphologies tested, spines greatly reduced location-dependent variability of local EPSP amplitude and kinetics, while having minimal impact on EPSPs measured at the soma. Spine-dependent standardization of local EPSP properties persisted across a range of physiologically relevant spine neck resistances, and in models with variable neck resistances. By reducing the variability of local EPSPs, spines standardized synaptic activation of NMDA receptors and voltage-gated calcium channels. Furthermore, spines enhanced activation of NMDA receptors and facilitated the generation of NMDA spikes and axonal action potentials in response to synaptic input. Finally, we show that dynamic regulation of spine neck geometry can preserve local EPSP properties following plasticity-driven changes in synaptic strength, but is inefficient in modifying the amplitude of EPSPs in other cellular compartments. These observations suggest that one function of dendritic spines is to standardize local EPSP properties throughout the dendritic tree, thereby allowing neurons to use similar voltage-sensitive postsynaptic mechanisms at all dendritic locations.     

Cortical area and species differences in dendritic spine morphology

Dendritic spines receive most excitatory inputs in the neocortex and are morphologically very diverse. Recent evidence has demonstrated linear relationships between the size and length of dendritic spines and important features of its synaptic junction and time constants for calcium compartmentalisation. Therefore, the morphologies of dendritic spines can be directly interpreted functionally. We sought to explore whether there were potential differences in spine morphologies between areas and species that could reflect potential functional differences. For this purpose, we reconstructed and measured thousands of dendritic spines from basal dendrites of layer III pyramidal neurons from mouse temporal and occipital cortex and from human temporal cortex. We find systematic differences in spine densities, spine head size and spine neck length among areas and species. Human spines are systematically larger and longer and exist at higher densities than those in mouse cortex. Also, mouse temporal spines are larger than mouse occipital spines. We do not encounter any correlations between the size of the spine head and its neck length. Our data suggests that the average synaptic input is modulated according to cortical area and differs among species. We discuss the implications of these findings for common algorithms of cortical processing.     

Morphological constraints on calcium dependent glutamate receptor trafficking into individual dendritic spine

Glutamate receptor trafficking into dendritic spines is a pivotal step in synaptic plasticity, yet the relevance of plasticity-producing rise of [Ca2+]i and of spine morphology to subsequent delivery of glutamate receptors into dendritic spine heads are still not well understood. Following chemical induction of LTP, an increase in eGFP-GluR1 fluorescence in short but not long dendritic spines of cultured hippocampal neurons was found. Repeated flash photolysis of caged calcium, which produced a transient rise of [Ca2+]i inside spine heads caused a selective, actin and protein synthesis dependent increase of eGFP-GluR1 in these spines. Strikingly, GluR1 increase was correlated with the ability of a calcium transient generated in the spine head to diffuse into the parent dendrite, and inversely correlated with the length of the spine: short spines were more likely to raise GluR1 than long ones. These observations link, for the first time, calcium transients in dendritic spines with spine morphology and its ability to undergo synaptic plasticity.     

Calcium dynamics in dendritic spines and spine motility

A dendritic spine is an intracellular compartment in synapses of central neurons. The role of the fast twitching of spines, brought about by a transient rise of internal calcium concentration above that of the parent dendrite, has been hitherto unclear. We propose an explanation of the cause and effect of the twitching and its role in the functioning of the spine as a fast calcium compartment. Our molecular model postulates that rapid spine motility is due to the concerted contraction of calcium-binding proteins. The contraction induces a stream of cytoplasmic fluid in the direction of the dendritic shaft, thus speeding up the time course of spine calcium dynamics, relative to pure diffusion. Simulations indicate that chemical reaction rate theory at the molecular level can explain spine motility. They reveal two time periods in calcium dynamics, as measured recently by other researchers. It appears that rapid motility in dendritic spines increases the efficiency of calcium conduction to the dendrite and speeds up the emptying of the spine. This could play a major role in the induction of synaptic plasticity. A prediction of the model is that alteration of spine motility will modify the time course of calcium in the dendritic spine and could be tested experimentally.     

Calcium dynamics in dendritic spines: a link to structural plasticity

Calcium signals evoked either by action potential or by synaptic activity play a crucial role for the synaptic plasticity within an individual spine. Because of the small size of spine and the indicators commonly used to measure spine calcium activity, calcium function can be severely disrupted. Therefore, it is very difficult to explain the exact relationship between spine geometry and spine calcium dynamics. Recently, it has been suggested that the medium range of calcium which induces long term potentiation leads to the structural stability stage of spines, while very low or very high amount of calcium leads to the long term depression stage which results in shortening and eventually pruning of spines. Here we propose a physiologically realistic computational model to examine the role of calcium and the mechanisms that govern its regulation in the spine morphology. Calcium enters into spine head through NMDA and AMPA channels and is regulated by internal stores. Contribution of this calcium in the induction of long term potentiation and long term depression is also discussed. Further it has also been predicted that the presence of internal stores depletes the total calcium accumulation in cytosol which is in agreement with the recent experimental and theoretical studies.     

Caldendrin and myosin V regulate synaptic spine apparatus localization via ER stabilization in dendritic spines

Excitatory synapses of principal hippocampal neurons are frequently located on dendritic spines. The dynamic strengthening or weakening of individual inputs results in structural and molecular diversity of dendritic spines. Active spines with large calcium ion (Ca²⁺) transients are frequently invaded by a single protrusion from the endoplasmic reticulum (ER), which is dynamically transported into spines via the actin‐based motor myosin V. An increase in synaptic strength correlates with stable anchoring of the ER, followed by the formation of an organelle referred to as the spine apparatus. Here, we show that myosin V binds the Ca²⁺ sensor caldendrin, a brain‐specific homolog of the well‐known myosin V interactor calmodulin. While calmodulin is an essential activator of myosin V motor function, we found that caldendrin acts as an inhibitor of processive myosin V movement. In mouse and rat hippocampal neurons, caldendrin regulates spine apparatus localization to a subset of dendritic spines through a myosin V‐dependent pathway. We propose that caldendrin transforms myosin into a stationary F‐actin tether that enables the localization of ER tubules and formation of the spine apparatus in dendritic spines.     

Modeling activity-dependent synapse restructuring

The spread of electrical activity in a dendritic tree is shaped, in part, by its morphology. Conversely, experimental evidence is growing that electrical and chemical activity can slowly shape the morphology of the dendrite. In this theoretical study, the dendritic spines are dynamic elements, with biophysical properties that change in response to patterns of electrical activity. Recent experiments and diagrammatic models suggest that activity-dependent processes can regulate structural modifications in dendritic spines as well as their distribution along the dendrite. This study considers how local changes in spine structure (minutes to hours) can influence patterns of electrical activity along the dendrite; and how electrical activity due to synaptic events and excitable membrane dynamics can, over time, influence the morphology of the dendrite. The model presents a slow subsystem for structural synaptic plasticity associated with long-term potentiation. A perturbation problem evolves naturally when the spine stem shortens, since the ratio of spine stem resistance to input resistance is small. Hence, the difference between the spine head and dendritic potentials become negligible. This paper presents an asymptotic expansion of head potential in terms of dendritic potential. This leads to a reduced model for post-synaptic restructuring that captures the dynamics of the full model in a briefer computation period when the spines are well connected to the dendrite.     

Dendritic spine geometry: functional implication and regulation

Dendritic spines are tiny protrusions on dendritic shafts where most excitatory synapses are located. Recent advances in imaging technologies have given us great insight into the function of spines as biochemical compartments. Here we review recent evidence suggesting that the geometry of dendritic spines controls postsynaptic calcium signaling and is bidirectionally regulated during synaptic plasticity.     

Subtle re-location of dendritic spine branches containing membrane with fast spiking mechanisms can alter synaptic efficacy

The potential physiological impact of morphological changes in the active dendritic spines, which are believed to be associated with altered synaptic efficacy, was investigated in a computer simulation study using the NEURON package [1]. A compartmental model of a simplified neuron was built, which included 30 complex spines (neck, head, and active zone) and accommodating AMPA-type synaptic inputs with alpha-function conductances. Hodgkin-Huxley type excitable membranes were inserted into the spine heads. It was shown that arranging spines in dense clusters, as opposed to a uniformly random spine distribution, has a negligible effect on the synaptic signal transfer (other model conditions, including synaptic input and spine density, remained unchanged). However, if a proportion (e.g., 3–20%) of the spines partly fuse with their neighbors forming branched spines, this could increase dramatically the cell response to the unchanged synaptic input. Results of this pilot study provide the basis for a more detailed investigation of the relationship between the spine arrangement and synaptic function, considering dual-component synaptic currents and mechanisms controlling ion fluxes in the dendritic compartments.     

A Compartmental Model for Activity-Dependent Dendritic Spine Branching

Dendritic spines are small, mushroom-like protrusions from the arbor of a neuron in the central nervous system. Interdependent changes in the morphology, biochemistry, and activity of spines have been associated with learning and memory. Moreover, post-mortem cortices from patients with Alzheimer’s or Parkinson’s disease exhibit biochemical and physical alterations within their dendritic arbors and a reduction in the number of dendritic spines. For over a decade, experimentalists have observed perforations in postsynaptic densities on dendritic spines after induction of long-term potentiation, a sustained enhancement of response to a brief electrical or chemical stimulus, associated with learning and memory. In more recent work, some suggest that activity-dependent intraspine calcium may regulate the surface area of the spine head, and reorganization of postsynaptic densities on the surface. In this paper, we develop a model of a dendritic spine with the ability to partition its transmission and receptor zones, as well as the entire spine head. Simulations are initially performed with fixed parameters for morphology to study electrical properties and identify parameters that increase efficacy of the synaptic connection. Equations are then introduced to incorporate calcium as a second messenger in regulating continuous changes in morphology. In the model, activity affects compartmental calcium, which regulates spine head morphology. Conversely, spine head morphology affects the level of local activity, whether the spines are modeled with passive membrane properties, or excitable membrane using Hodgkin–Huxley kinetics. Results indicate that merely separating the postsynaptic receptors on the surface of the spine may add to the diversity of circuitry, but does not change the efficacy of the synapse. However, when the surface area of the spine is a dynamic variable, efficacy of the synapse may vary continuously over time.     

The Phosphodiesterase 9 Inhibitor PF‐04449613 Promotes Dendritic Spine Formation and Performance Improvement after Motor Learning

The cyclic nucleotide cGMP is an intracellular second messenger with important roles in neuronal functions and animals' behaviors. The phosphodiesterases (PDEs) are a family of enzymes that hydrolyze the second messengers cGMP and cAMP. Inhibition of phosphodiesterase 9 (PDE9), a main isoform of PDEs hydrolyzing cGMP, has been shown to improve learning and memory as well as cognitive function in rodents. However, the role of PDE9 in regulating neuronal structure and function in vivo remains unclear. Here we used in vivo two‐photon microscopy to investigate the effect of a selective PDE9 inhibitor PF‐04449613 on the activity and plasticity of dendritic spines of layer V pyramidal neurons in the mouse primary motor cortex. We found that administration of PF‐04449613 increased calcium activity of dendrites and dendritic spines of layer V pyramidal neurons in mice under resting and running conditions. Chronic treatment of PF‐04449613 over weeks increased dendritic spine formation and elimination under basal conditions. Furthermore, PF‐04449613 treatment over 1–7 days increased the formation and survival of new spines as well as performance improvement after rotarod motor training. Taken together, our studies suggest that elevating the level of cGMP with the PDE9 inhibitor PF‐04449613 increases synaptic calcium activity and learning‐dependent synaptic plasticity, thereby contributing to performance improvement after learning. © 2018 Wiley Periodicals, Inc. Develop Neurobiol 00: 000–000, 2018     

Three-dimentional organization of synapses and astroglia in the hippocampus of rats and ground squirrels: new structural and functional paradigms of the synapse function

The literature data and our own data on the synaptic plasticity and remodeling of synaptic organelles in the central nervous system are reviewed. Modern techniques of laser scanning confocal microscopy and serial thin sectioning for in vivo and in vitro studies of dendritic spines, including the relationship between morphological changes and the efficacy of synaptic transmission, are discussed using, in particular, a model of long-term potentiation. The organization of dendritic spines and postsynaptic densities of different categories as well as the role of filopodia in spine genesis were analyzed. It was shown that the method of serial ultrathin sections is the most effective for unbiased quantitative stereological analysis and 3D reconstructions. By using the refined method of serial ultrathin sections with subsequent three-dimensional reconstructions, the presence of giant mitochondria in hippocampal neuronal dendrites was demonstrated. It was shown that smooth endoplasmic reticulum forms a unified continuum with the outer membrane of the mitochondrial envelope within dendrites. It was suggested that this continuum provides calcium tunneling, which makes possible intracellular signal transduction during synaptic transmission. Evidence is presented indicating the presence of gap junctions ("electrical synapses") in the synapses of mammalian brain, as well as between glial processes, and between glial cells and neurons. Our data and the data of other authors show that glial cell processes form a structural and functional glial network, which modulates the functioning of the neuronal network. The connection of dendritic spines with the glial network is shown on 3D reconstructions by analyzing the neuropil volume in CA1 hippocampal area of ground squirrels in three functional states: normothermia, provoked arousal, and hibernation when brain temperature falls below 6 degrees C. The own data of the authors are discussed indicating the formation of more than five presynaptic boutons (multiple synapses) on both CA1 mushroom-like dendritic spines and CA3 thorny excrescences. On the basis of the analysis, new ideas of the organization and functioning of synapses were suggested.     

The RNA binding protein TLS is translocated to dendritic spines by mGluR5 activation and regulates spine morphology

Neuronal dendrites, together with dendritic spines, exhibit enormously diverse structure. Selective targeting and local translation of mRNAs in dendritic spines have been implicated in synapse remodeling or synaptic plasticity. The mechanism of mRNA transport to the postsynaptic site is a fundamental question in local dendritic translation. TLS (translocated in liposarcoma), previously identified as a component of hnRNP complexes, unexpectedly showed somatodendritic localization in mature hippocampal pyramidal neurons. In the present study, TLS was translocated to dendrites and was recruited to dendrites not only via microtubules but also via actin filaments. In mature hippocampal pyramidal neurons, TLS accumulated in the spines at excitatory postsynapses upon mGluR5 activation, which was accompanied by an increased RNA content in dendrites. Consistent with the in vitro studies, TLS-null hippocampal pyramidal neurons exhibited abnormal spine morphology and lower spine density. Our results indicate that TLS participates in mRNA sorting to the dendritic spines induced by mGluR5 activation and regulates spine morphology to stabilize the synaptic structure.     

High speed two-photon imaging of calcium dynamics in dendritic spines: consequences for spine calcium kinetics and buffer capacity

Rapid calcium concentration changes in postsynaptic structures are crucial for synaptic plasticity. Thus far, the determinants of postsynaptic calcium dynamics have been studied predominantly based on the decay kinetics of calcium transients. Calcium rise times in spines in response to single action potentials (AP) are almost never measured due to technical limitations, but they could be crucial for synaptic plasticity. With high-speed, precisely-targeted, two-photon point imaging we measured both calcium rise and decay kinetics in spines and secondary dendrites in neocortical pyramidal neurons. We found that both rise and decay kinetics of changes in calcium-indicator fluorescence are about twice as fast in spines. During AP trains, spine calcium changes follow each AP, but not in dendrites. Apart from the higher surface-to-volume ratio (SVR), we observed that neocortical dendritic spines have a markedly smaller endogenous buffer capacity with respect to their parental dendrites. Calcium influx time course and calcium extrusion rate were both in the same range for spines and dendrites when fitted with a dynamic multi-compartment model that included calcium binding kinetics and diffusion. In a subsequent analysis we used this model to investigate which parameters are critical determinants in spine calcium dynamics. The model confirmed the experimental findings: a higher SVR is not sufficient by itself to explain the faster rise time kinetics in spines, but only when paired with a lower buffer capacity in spines. Simulations at zero calcium-dye conditions show that calmodulin is more efficiently activated in spines, which indicates that spine morphology and buffering conditions in neocortical spines favor synaptic plasticity.     

Convergent CaMK and RacGEF signals control dendritic structure and function

Structural plasticity of excitatory synapses is a vital component of neuronal development, synaptic plasticity and behavior, and its malfunction underlies many neurodevelopmental and psychiatric disorders. However, the molecular mechanisms that control dendritic spine morphogenesis have only recently emerged. We summarize recent work that has revealed an important connection between calcium/calmodulin-dependent kinases (CaMKs) and guanine-nucleotide-exchange factors (GEFs) that activate the small GTPase Rac (RacGEFs) in controlling dendritic spine morphogenesis. These two groups of molecules function in neurons as a unique signaling cassette that transduces calcium influx into small GTPase activity and, thence, actin reorganization and spine morphogenesis. Through this pathway, CaMKs and RacGEFs amplify calcium signals and translate them into spatially and temporally regulated structural remodeling of dendritic spines.     

A possible mechanism of morphometric changes in dendritic spines induced by stimulation

A number of experimental procedures which induce increased electrical activity (including long-term potentiation) were shown to be accompanied by morphometric changes in dendritic spines. These changes include an enlargement of the spine head, shortening and widening of the spine stalk, and an increase in the length of synaptic apposition. A possible mechanism is suggested which takes into account specific cytological features of the spine and the existence of contractile proteins in neurons. Dendritic spines are defined as special domains of the neuron which have a unique organization of the cytoplasm. Actin filaments form a very dense network in the spine head, and they are longitudinally organized within the spine stalk. Spines were also shown to contain myosin and other actin-regulatory proteins. The high density of the actin network could explain the characteristic absence of the cytoplasmic organelles from dendritic spines. In analogy with other cells, such an actin organization indicates low levels of free cytosolic calcium. Even in the resting state, calcium levels may be unevenly distributed through the neuron, being lowest within the subplasmalemmal region. Due to the high surface-to-volume ratio in spines, the cytoplasm is formed mostly by the subplasmalemmal region. The spine apparatus or the smooth endoplasmic reticulum, which is recognized as a calcium-sequestering site in spines, may also contribute to the low calcium levels there. However, when in the stimulated spine the voltage-dependent calcium channels open, then, given the spine's high surface-to-volume ratio, the concentration of calcium may very quickly attain levels that will activate the actin-regulatory proteins and myosin and thus trigger the chain of events leading to the enlargement of the spine head and to the contraction (i.e., widening and shortening) of the spine stalk. The increased free cytosolic calcium may also activate the protein-producing system localized at the base of the spine, which, under certain conditions, could stabilize the morphometric changes of the spine.     

Spine-neck geometry determines NMDA receptor-dependent Ca2+ signaling in dendrites

Increases in cytosolic Ca2+ concentration ([Ca2+]i) mediated by NMDA-sensitive glutamate receptors (NMDARs) are important for synaptic plasticity. We studied a wide variety of dendritic spines on rat CA1 pyramidal neurons in acute hippocampal slices. Two-photon uncaging and Ca2+ imaging revealed that NMDAR-mediated currents increased with spine-head volume and that even the smallest spines contained a significant number of NMDARs. The fate of Ca2+ that entered spine heads through NMDARs was governed by the shape (length and radius) of the spine neck. Larger spines had necks that permitted greater efflux of Ca2+ into the dendritic shaft, whereas smaller spines manifested a larger increase in [Ca2+]i within the spine compartment as a result of a smaller Ca2+ flux through the neck. Spine-neck geometry is thus an important determinant of spine Ca2+ signaling, allowing small spines to be the preferential sites for isolated induction of long-term potentiation.