Measuring F-actin properties in dendritic spines

Koskinen, Mikko; Hotulainen, Pirta

doi:10.3389/fnana.2014.00074

Cited by 40 publications

(39 citation statements)

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“…Conventional fluorescence microscopy in combination with photoactivatable probes or fluorescence recovery after photo-bleaching (FRAP) is frequently used to study protein localization or protein dynamics in spines89. However, due to the diffraction of light, it is not possible to resolve structures below approximately half of the wavelength used to excite the fluorophore, which is about 250 nm.…”

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confidence: 99%

Periodic F-actin structures shape the neck of dendritic spines

Bär

Kobler

Bommel

et al. 2016

Sci Rep

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Most of the excitatory synapses on principal neurons of the forebrain are located on specialized structures called dendritic spines. Their morphology, comprising a spine head connected to the dendritic branch via a thin neck, provides biochemical and electrical compartmentalization during signal transmission. Spine shape is defined and tightly controlled by the organization of the actin cytoskeleton. Alterations in synaptic strength correlate with changes in the morphological appearance of the spine head and neck. Therefore, it is important to get a better understanding of the nanoscale organization of the actin cytoskeleton in dendritic spines. A periodic organization of the actin/spectrin lattice was recently discovered in axons and a small fraction of dendrites using super-resolution microscopy. Here we use a small probe phalloidin-Atto647N, to label F-actin in mature hippocampal primary neurons and in living hippocampal slices. STED nanoscopy reveals that in contrast to β-II spectrin antibody labelling, phalloidin-Atto647N stains periodic actin structures in all dendrites and the neck of nearly all dendritic spines, including filopodia-like spines. These findings extend the current view on F-actin organization in dendritic spines and may provide new avenues for understanding the structural changes in the spine neck during induction of synaptic plasticity, active organelle transport or tethering.

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confidence: 99%

Periodic F-actin structures shape the neck of dendritic spines

Bär

Kobler

Bommel

et al. 2016

Sci Rep

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“…This was also confirmed by measuring the decay of photoactivated PAGFP‐actin [Koskinen et al, ]. Accordingly, F‐actin filaments are more tightly crosslinked in mature spines [Koskinen and Hotulainen, ]. Although the size of the stable F‐actin pool was larger in DIV21 neurons, the turnover rate of the dynamic F‐actin pool was also higher at this time point than in DIV14 neurons [Koskinen et al, ] (Fig.…”

Section: Actin Dynamics During Dendritic Spine Maturationmentioning

confidence: 55%

“…Additionally, this information could be used to improve the methods employed for the differentiation of the neurons derived from induced pluripotent cells. To the best of our knowledge, only three studies have addressed the changes in dendritic spine actin cytoskeleton during neuronal maturation so far [Star et al, ; Koskinen and Hotulainen, ; Koskinen et al, ]. Star et al [] used fluorescence recovery after photobleaching (FRAP) to study populations of dissociated hippocampal neurons cultured for 14 (DIV14) or 21 (DIV21) days in vitro and found no difference in the actin turnover rate.…”

Section: Actin Dynamics During Dendritic Spine Maturationmentioning

confidence: 99%

“…Myosin IIb stabilizes the stable F‐actin pool through actin crosslinking. Simultaneously, activation of myosin IIb contractility increases the treadmilling rate of the dynamic pool [Koskinen and Hotulainen, ; Koskinen et al, ]. We proposed a model for maturation, where myosin IIb stabilizes the actin filaments in the base of the dendritic spine heads by crosslinking actin filaments [Koskinen et al, ].…”

Section: Actin Dynamics During Dendritic Spine Maturationmentioning

confidence: 99%

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Dendritic spine actin dynamics in neuronal maturation and synaptic plasticity

2016

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The majority of the postsynaptic terminals of excitatory synapses in the central nervous system exist on small bulbous structures on dendrites known as dendritic spines. The actin cytoskeleton is a structural element underlying the proper development and morphology of dendritic spines. Synaptic activity patterns rapidly change actin dynamics, leading to morphological changes in dendritic spines. In this mini-review, we will discuss recent findings on neuronal maturation and synaptic plasticityinduced changes in the dendritic spine actin cytoskeleton. We propose that actin dynamics in dendritic spines decrease through actin filament crosslinking during neuronal maturation. In long-term potentiation, we evaluate the model of fast breakdown of actin filaments through severing and rebuilding through polymerization and later stabilization through crosslinking. We will discuss the role of Ca 21 in long-term depression, and suggest that actin filaments are dissolved through actin filament severing. V C 2016 Wiley Periodicals, Inc.Key Words: dendritic spines; actin cytoskeleton; neuronal maturation; synaptic plasticity Introduction T he majority of postsynaptic terminals of excitatory synapses in the central nervous system exist on small bulbous structures on neuronal dendrites known as dendritic spines. Dendritic spines can be divided into three morphological categories: thin, consisting of a long thin neck and a small round head; stubby, with a large bulbous head and a short wide neck; and mushroom, characterized by a large bulbous head and a short narrow neck [Bourne and Harris, 2008]. Dendritic spine maturation is a process where the postsynaptic machinery is recruited to the newly formed spine and the spine acquires a more stable, usually mushroom-shaped morphology [Dailey and Smith, 1996;Dunaevsky et al., 1999]. The shape and size of the spine head and neck have been shown to influence the electrical properties and compartmentalization of the spine [Noguchi et al., 2005] and morphological changes were shown to account for changes in synaptic function [Yuste and Bonhoeffer, 2001]. Accordingly, synaptic activity is translated into structural changes in dendritic spines and functional changes in synaptic efficacy.The actin cytoskeleton is a structural element underlying the proper development and morphology of dendritic spines, where it controls the morphological and structural changes induced by synapse activation. Actin filaments are polar structures with one end growing more rapidly (the plus or "barbed" end) than the other (the minus or "pointed" end). The constant removal of actin subunits from the pointed ends and addition at the barbed ends is known as actin treadmilling. Filamentous (F-) actin structures range from a branched filament network to thick bundles of several crosslinked filaments [Blanchoin et al., 2014]. These structures have highly variable lifetimes. Filaments forming the actin mesh can change very rapidly whereas thick actin bundles can remain stable for a long time. Based on the turnover rate...

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“…Most of these applications are solution-based and performed with a spectrofluorimeter type of instrument; hence, the lack of spatial resolution is not relevant. However, with the development of advanced instrumentation, in particular confocal and two photon laser-scanning microscopy [2,3], fluorescence polarization imaging has become more and more popular over the last two decades [4,5]. Many polarization-based probes have been developed that can be used as biosensors in vitro as well as in living cells or tissues.…”

Section: Introductionmentioning

confidence: 99%

3D fluorescence anisotropy imaging using selective plane illumination microscopy

Hedde¹,

Ranjit²,

Gratton³

2015

Opt. Express

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Abstract:Fluorescence anisotropy imaging is a popular method to visualize changes in organization and conformation of biomolecules within cells and tissues. In such an experiment, depolarization effects resulting from differences in orientation, proximity and rotational mobility of fluorescently labeled molecules are probed with high spatial resolution. Fluorescence anisotropy is typically imaged using laser scanning and epifluorescencebased approaches. Unfortunately, those techniques are limited in either axial resolution, image acquisition speed, or by photobleaching. In the last decade, however, selective plane illumination microscopy has emerged as the preferred choice for three-dimensional time lapse imaging combining axial sectioning capability with fast, camera-based image acquisition, and minimal light exposure. We demonstrate how selective plane illumination microscopy can be utilized for three-dimensional fluorescence anisotropy imaging of live cells. We further examined the formation of focal adhesions by three-dimensional time lapse anisotropy imaging of CHO-K1 cells expressing an EGFP-paxillin fusion protein.

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Measuring F-actin properties in dendritic spines

Cited by 40 publications

References 57 publications

Periodic F-actin structures shape the neck of dendritic spines

Periodic F-actin structures shape the neck of dendritic spines

Dendritic spine actin dynamics in neuronal maturation and synaptic plasticity

3D fluorescence anisotropy imaging using selective plane illumination microscopy

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