Time-lapse imaging of dendritic spines in vitro
Abstract
Dendritic spines are small protrusions present postsynaptically at approximately 90% of excitatory synapses in the brain. Spines undergo rapid spontaneous changes in shape that are thought to be important for alterations in synaptic connectivity underlying learning and memory. Visualization of these dynamic changes in spine morphology are especially challenging because of the small size of spines (approximately 1 microm). Here we describe a microscope system, based on a spinning-disk confocal microscope, suitable for imaging mature dendritic spines in brain slice preparations, with a time resolution of seconds. We discuss two commonly used in vitro brain slice preparations and methods for transfecting them. Preparation and transfection require approximately 1 d, after which slices must be cultured for at least 21 d to obtain spines of mature morphology. We also describe imaging and computer analysis routines for studying spine motility. These procedures require in the order of 2 to 4 h.
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Organotypic hippocampal slice cultures undergo key developmental steps similar to in vivo, including synapse maturation.
“Organotypic hippocampal slice cultures are known to undergo key developmental steps similar to those observed in vivo, including synapse maturation28–31, and have been previously used to study ramified microglia function32.”
From Microglia remodel synapses by presynaptic trogocytosis and spine head filopodia induction (2018) · cites this paper as [31]
Cites
A simple method for organotypic cultures of nervous tissue
Stoppini, L., Buchs, P., & Muller, D. (1991) · Journal of Neuroscience Methods · doi:10.1016/0165-0270(91)90128-m · Cited by 3,097 · see citing works
A guide to choosing fluorescent proteins
Shaner, N. C., Steinbach, P. A., & Tsien, R. Y. (2005) · Nature Methods · doi:10.1038/nmeth819 · Cited by 2,823 · see citing works
Structural basis of long-term potentiation in single dendritic spines
Matsuzaki, M., Honkura, N., Ellis-Davies, G. C. R., & Kasai, H. (2004) · Nature · doi:10.1038/nature02617 · Cited by 2,500 · see citing works
Heim, R., Cubitt, A. B., & Tsien, R. Y. (1995) · Nature · doi:10.1038/373663b0 · Cited by 1,792 · see citing works
Long-term dendritic spine stability in the adult cortex
Grutzendler, J., Kasthuri, N., & Gan, W.-B. (2002) · Nature · doi:10.1038/nature01276 · Cited by 1,198 · cited 1× · see citing works
“Computer-operated controller for motorized z-drive and xy-stage (16) MAC5000, Ludl Electronic Products, http://www.”
MethodRapid Actin-Based Plasticity in Dendritic Spines
Fischer, M., Kaech, S., Knutti, D., & Matus, A. (1998) · Neuron · doi:10.1016/s0896-6273(00)80467-5 · Cited by 905 · see citing works
Self-citationOrganotypic slice cultures: a technique has come of age
Gähwiler, B. H. (1997) · Trends in Neurosciences · doi:10.1016/s0166-2236(97)01122-3 · Cited by 884 · see citing works
Organotypic monolayer cultures of nervous tissue
Gähwiler, B. H. (1981) · Journal of Neuroscience Methods · doi:10.1016/0165-0270(81)90003-0 · Cited by 873 · see citing works
Okamoto, K.-I., Nagai, T., Miyawaki, A., & Hayashi, Y. (2004) · Nature Neuroscience · doi:10.1038/nn1311 · Cited by 854 · see citing works
Experience-dependent plasticity of dendritic spines in the developing rat barrel cortex in vivo
Lendvai, B., Stern, E. A., Chen, B., & Svoboda, K. (2000) · Nature · doi:10.1038/35009107 · Cited by 802 · see citing works
Synaptogenesis Via Dendritic Filopodia in Developing Hippocampal Area CA1
Fiala, J. C., Feinberg, M., Popov, V., & Harris, K. M. (1998) · The Journal of Neuroscience · doi:10.1523/jneurosci.18-21-08900.1998 · Cited by 758 · see citing works
Allison, D. W., Gelfand, V. I., Spector, I., & Craig, A. M. (1998) · The Journal of Neuroscience · doi:10.1523/jneurosci.18-07-02423.1998 · Cited by 556 · see citing works
Glutamate receptors regulate actin-based plasticity in dendritic spines
Fischer, M., Kaech, S., Wagner, U., Brinkhaus, H., & Matus, A. (2000) · Nature Neuroscience · doi:10.1038/78791 · Cited by 446 · see citing works
Self-citationElectron Microscopy of Synaptic Contacts on Dendrite Spines of the Cerebral Cortex
Gray, E. G. (1959) · Nature · doi:10.1038/1831592a0 · Cited by 443 · see citing works
Developmental regulation of spine motility in the mammalian central nervous system
Dunaevsky, A., Tashiro, A., Majewska, A., Mason, C., & Yuste, R. (1999) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.96.23.13438 · Cited by 412 · see citing works
High actin concentrations in brain dendritic spines and postsynaptic densities.
Matus, A., Ackermann, M., Pehling, G., Byers, H. R., & Fujiwara, K. (1982) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.79.23.7590 · Cited by 401 · see citing works
Self-citationBook Review: Live Cell Imaging, A Laboratory Manual
Masters, B. R. (2006) · Journal of Biomedical Optics · doi:10.1117/1.2150471 · Cited by 355 · see citing works
Environmental-Dependent Acceleration of a Developmental Switch: The Floral Transition
Simpson, G. G., & Dean, C. (2000) · Science s STKE · doi:10.1126/stke.2000.18.pe1 · Cited by 318 · see citing works
Labeling Neural Cells Using Adenoviral Gene Transfer of Membrane-Targeted GFP
Moriyoshi, K., Richards, L. J., Akazawa, C., O’Leary, D. D. M., & Nakanishi, S. (1996) · Neuron · doi:10.1016/s0896-6273(00)80044-6 · Cited by 280 · see citing works
AMPA receptors regulate dynamic equilibrium of presynaptic terminals in mature hippocampal networks
Paola, V. D., Arber, S., & Caroni, P. (2003) · Nature Neuroscience · doi:10.1038/nn1046 · Cited by 236 · see citing works
Neuronal transfection in brain slices using particle-mediated gene transfer
Lo, D. C., McAllister, A. K., & Katz, L. C. (1994) · Neuron · doi:10.1016/0896-6273(94)90412-x · Cited by 230 · see citing works
Ehrengruber, M. U., Lundstrom, K. H., Schweitzer, C., Heuss, C., Schlesinger, S., & Gähwiler, B. H. (1999) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.96.12.7041 · Cited by 183 · see citing works
Cytoskeletal Plasticity in Cells Expressing Neuronal Microtubule-Associated Proteins
Kaech, S., Ludin, B., & Matus, A. I. (1996) · Neuron · doi:10.1016/s0896-6273(00)80249-4 · Cited by 163 · see citing works
Self-citationIsoform Specificity in the Relationship of Actin to Dendritic Spines
Kaech, S., Fischer, M., Doll, T., & Matus, A. (1997) · The Journal of Neuroscience · doi:10.1523/jneurosci.17-24-09565.1997 · Cited by 155 · see citing works
Self-citationBiolistic transfection of neuronal cultures using a hand-held gene gun
O’Brien, J., & Lummis, S. C. R. (2006) · Nature Protocols · doi:10.1038/nprot.2006.145 · Cited by 126 · see citing works
Cytoskeletal microdifferentiation: a mechanism for organizing morphological plasticity in dendrites
Kaech, S., Parmar, H., Roelandse, M., Bornmann, C., & Matus, A. (2001) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.111146798 · Cited by 124 · see citing works
Self-citationBiolistic Transfection of Neurons
McAllister, A. K. (2000) · Science s STKE · doi:10.1126/stke.2000.51.pl1 · Cited by 71 · see citing works
Hypothermia-Associated Loss of Dendritic Spines
Roelandse, M., & Matus, A. I. (2004) · Journal of Neuroscience · doi:10.1523/jneurosci.2872-04.2004 · Cited by 55 · see citing works
Self-citationFocal motility determines the geometry of dendritic spines☆
Roelandse, M., Welman, A., Wagner, U., Hagmann, J., & Matus, A. I. (2003) · Neuroscience · doi:10.1016/s0306-4522(03)00405-6 · Cited by 40 · see citing works
Self-citationBiolistic transfection of organotypic cultures of rat visual cortex using a handheld device
Wirth, M. J., & Wahle, P. (2003) · Journal of Neuroscience Methods · doi:10.1016/s0165-0270(03)00024-4 · Cited by 24 · see citing works
Griesbeck, O., Körte, M., Gravel, C., Bonhoeffer, T., & Thoenen, H. (1997) · Molecular Brain Research · doi:10.1016/s0169-328x(96)00246-x · Cited by 16 · see citing works
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Orlando, C., Ster, J., Gerber, U., Fawcett, J. W., & Raineteau, O. (2012) · The Journal of Neuroscience · doi:10.1523/jneurosci.2406-12.2012 · Cited by 153 · see citing works
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Enríquez-Barreto, L., Cuesto, G., Domıńguez-Iturza, N., Gavilán, E., Ruano, D., Sandi, C., Fernández‐Ruiz, A., Martín-Vázquez, G., Herreras, Ó., & Morales, M. (2014) · Frontiers in Molecular Neuroscience · doi:10.3389/fnmol.2013.00054 · Cited by 33 · see citing works
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Falougy, H. E., Filová, B., Ostatníková, D., Bačová, Z., & Bakoš, J. (2019) · Endocrine Regulations · doi:10.2478/enr-2019-0006 · Cited by 22 · see citing works
Kiiski, H., Äänismaa, R., Tenhunen, J. J., Hagman, S., Ylä‐Outinen, L., Aho, A., Yli‐Hankala, A., Bendel, S., Skottman, H., & Narkilahti, S. (2013) · Biology Open · doi:10.1242/bio.20134648 · Cited by 15 · see citing works
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- Authors
- Verkuyl, J. M., & Matus, A.
- Affiliations
- Friedrich Miescher Institute (CH)
- Corresponding author
- Matus, Andrew
- ORCID
- Verkuyl, J Martin
- Venue
- Nature Protocols
- Published
- 2006-12
- Volume
- 1
- Issue
- 5
- Pages
- 2399-2405
- ISSN
- 1750-2799 (ISSN-L)
- 1754-2189
- OpenAlex
- W1990142376
- Cited by
- 13
- References
- 31
- Citations by year
- 2024: 1 · 2020: 1 · 2019: 3 · 2018: 2 · 2015: 1 · 2014: 3 · 2013: 1 · 2012: 1
- FWCI
- 0.00
- Type
- journal article
- Language
- en
- Open access
- closed
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Protocols (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Cell Image Analysis Techniques
- Keywords
- dendritic spines, spine motility, dendritic spine morphology, time-lapse imaging, spinning disk confocal microscopy, brain slice preparation, in vitro transfection, excitatory synapses, synaptic plasticity, learning and memory
- MeSH terms
- Electrophysiology, Hippocampus, Time Factors, Microscopy, Confocal, Tissue Culture Techniques, Dendritic Spines
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