SYNPLA: A synapse-specific method for identifying learning-induced synaptic plasticity loci

AbstractWhich neural circuits undergo synaptic changes when an animal learns? Although it is widely accepted that changes in synaptic strength underlie many forms of learning and memory, it remains challenging to connect changes in synaptic strength at specific neural pathways to specific behaviors and memories. Here we introduce SYNPLA (SYNaptic Proximity Ligation Assay), a synapse-specific, high-throughput and potentially brain-wide method capable of detecting circuit-specific learning-induced synaptic plasticity.

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SYNPLA, a method to identify synapses displaying plasticity after learning

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1919911117 ◽

2020 ◽

Vol 117 (6) ◽

pp. 3214-3219 ◽

Cited By ~ 4

Author(s):

Kim Dore ◽

Yvonne Pao ◽

Jose Soria Lopez ◽

Sage Aronson ◽

Huiqing Zhan ◽

...

Keyword(s):

Synaptic Plasticity ◽

Learning And Memory ◽

High Throughput ◽

Neural Circuits ◽

Synaptic Strength ◽

Proximity Ligation Assay ◽

Neural Pathways ◽

Proximity Ligation ◽

Synaptic Changes ◽

Specific Learning

Which neural circuits undergo synaptic changes when an animal learns? Although it is widely accepted that changes in synaptic strength underlie many forms of learning and memory, it remains challenging to connect changes in synaptic strength at specific neural pathways to specific behaviors and memories. Here we introduce SYNPLA (synaptic proximity ligation assay), a synapse-specific, high-throughput, and potentially brain-wide method capable of detecting circuit-specific learning-induced synaptic plasticity.

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Adult medial habenula neurons require GDNF receptor GFRα1 for synaptic stability and function

PLoS Biology ◽

10.1371/journal.pbio.3001350 ◽

2021 ◽

Vol 19 (11) ◽

pp. e3001350

Author(s):

Diana Fernández-Suárez ◽

Favio A. Krapacher ◽

Katarzyna Pietrajtis ◽

Annika Andersson ◽

Lilian Kisiswa ◽

...

Keyword(s):

Synaptic Plasticity ◽

Proximity Ligation Assay ◽

Glutamatergic Synapses ◽

Proximity Ligation ◽

Postsynaptic Currents ◽

Adult Mice ◽

Medial Habenula ◽

Factor Receptor Alpha ◽

And Function ◽

Anxiety And Fear

The medial habenula (mHb) is an understudied small brain nucleus linking forebrain and midbrain structures controlling anxiety and fear behaviors. The mechanisms that maintain the structural and functional integrity of mHb neurons and their synapses remain unknown. Using spatiotemporally controlled Cre-mediated recombination in adult mice, we found that the glial cell–derived neurotrophic factor receptor alpha 1 (GFRα1) is required in adult mHb neurons for synaptic stability and function. mHb neurons express some of the highest levels of GFRα1 in the mouse brain, and acute ablation of GFRα1 results in loss of septohabenular and habenulointerpeduncular glutamatergic synapses, with the remaining synapses displaying reduced numbers of presynaptic vesicles. Chemo- and optogenetic studies in mice lacking GFRα1 revealed impaired circuit connectivity, reduced AMPA receptor postsynaptic currents, and abnormally low rectification index (R.I.) of AMPARs, suggesting reduced Ca2+ permeability. Further biochemical and proximity ligation assay (PLA) studies defined the presence of GluA1/GluA2 (Ca2+ impermeable) as well as GluA1/GluA4 (Ca2+ permeable) AMPAR complexes in mHb neurons, as well as clear differences in the levels and association of AMPAR subunits with mHb neurons lacking GFRα1. Finally, acute loss of GFRα1 in adult mHb neurons reduced anxiety-like behavior and potentiated context-based fear responses, phenocopying the effects of lesions to septal projections to the mHb. These results uncover an unexpected function for GFRα1 in the maintenance and function of adult glutamatergic synapses and reveal a potential new mechanism for regulating synaptic plasticity in the septohabenulointerpeduncular pathway and attuning of anxiety and fear behaviors.

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The Proximity Ligation Assay: A High Throughput Technique for Protein Analysis in Neuroscience

Neural Stem Cell Assays ◽

10.1002/9781118308295.ch23 ◽

2015 ◽

pp. 231-240

Author(s):

Michela Zaltieri ◽

PierFranco Spano ◽

Cristina Missale ◽

Arianna Bellucci

Keyword(s):

High Throughput ◽

Protein Analysis ◽

Proximity Ligation Assay ◽

Proximity Ligation

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HiPLA: High-throughput imaging Proximity Ligation Assay

10.1101/371062 ◽

2018 ◽

Author(s):

Leonid A. Serebryannyy ◽

Tom Misteli

Keyword(s):

High Throughput ◽

Protein Interactions ◽

Large Scale ◽

Protein Complexes ◽

Nuclear Lamina ◽

Cellular Protein ◽

Premature Aging ◽

Proximity Ligation Assay ◽

Proximity Ligation ◽

High Throughput Imaging

AbstractProtein-protein interactions are essential for cellular structure and function. To delineate how the intricate assembly of protein interactions contribute to cellular processes in health and disease, new methodologies that are both highly sensitive and can be applied at large scale are needed. Here, we develop HiPLA (high-throughput imaging proximity ligation assay), a method that employs the antibody-based proximity ligation assay in a high-throughput imaging screening format to systematically probe protein interactomes. Using HiPLA, we probe the interaction of 60 proteins and associated PTMs with the nuclear lamina in a model of the premature aging disorder Hutchinson-Gilford progeria syndrome (HGPS). We identify a subset of proteins that differentially interact with the nuclear lamina in HGPS. In combination with quantitative indirect immunofluorescence, we find that the majority of differential interactions were accompanied by corresponding changes in expression of the interacting protein. Taken together, HiPLA offers a novel approach to probe cellular protein-protein interaction at a large scale and reveals mechanistic insights into the assembly of protein complexes.

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Adult medial habenula neurons require GDNF receptor GFRalpha1 for synaptic stability and function

10.1101/2021.07.07.451421 ◽

2021 ◽

Author(s):

Diana Fernandez-Suarez ◽

Favio Krapacher ◽

Katarzyna Pietrajtis ◽

Annika Andersson ◽

Lilian Kisiswa ◽

...

Keyword(s):

Synaptic Plasticity ◽

Proximity Ligation Assay ◽

Glutamatergic Synapses ◽

Genetic Studies ◽

Proximity Ligation ◽

Postsynaptic Currents ◽

Adult Mice ◽

Medial Habenula ◽

And Function ◽

Anxiety And Fear

The medial habenula (mHb) is an understudied small brain nucleus linking forebrain and midbrain structures controlling anxiety and fear behaviors. The mechanisms that maintain the structural and functional integrity of mHb neurons and their synapses remain unknown. Using spatio-temporally controlled Cre-mediated recombination in adult mice, we found that the GDNF receptor alpha 1 (GFRα1) is required in adult mHb neurons for synaptic stability and function. mHb neurons express some of the highest levels of GFRα1 in the mouse brain, and acute ablation of GFRα1 results in loss of septo-habenular and habenulo-interpeduncular glutamatergic synapses, with the remaining synapses displaying reduced numbers of presynaptic vesicles. Chemo- and opto-genetic studies in mice lacking GFRα1 revealed impaired circuit connectivity, reduced AMPA receptor postsynaptic currents, and abnormally low rectification index of AMPARs, suggesting reduced Ca 2+ -permeability. Further biochemical and proximity ligation assay studies defined the presence of GluA1/GluA2 (Ca 2+ -impermeable) as well as GluA1/GluA4 (Ca 2+ -permeable) AMPAR complexes in mHb neurons, as well as clear differences in the levels and association of AMPAR subunits in mHb neurons lacking GFRα1. Finally, acute loss of GFRα1 in adult mHb neurons reduced anxiety-like behavior and potentiated context-based fear responses, phenocopying the effects of lesions to septal projections to the mHb. These results uncover an unexpected function for GFRα1 in the maintenance and function of adult glutamatergic synapses, and reveal a potential new mechanism for regulating synaptic plasticity in the septo-habenulo-interpeduncular pathway and attuning of anxiety and fear behaviors.

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HiPLA: High-throughput imaging proximity ligation assay

Methods ◽

10.1016/j.ymeth.2018.11.004 ◽

2019 ◽

Vol 157 ◽

pp. 80-87 ◽

Cited By ~ 3

Author(s):

Leonid A. Serebryannyy ◽

Tom Misteli

Keyword(s):

High Throughput ◽

Proximity Ligation Assay ◽

Proximity Ligation ◽

High Throughput Imaging

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How rhythms of the sleeping brain tune memory and synaptic plasticity

SLEEP ◽

10.1093/sleep/zsz095 ◽

2019 ◽

Vol 42 (7) ◽

Cited By ~ 13

Author(s):

Carlos Puentes-Mestril ◽

James Roach ◽

Niels Niethard ◽

Michal Zochowski ◽

Sara J Aton

Keyword(s):

Synaptic Plasticity ◽

Eye Movement ◽

Neural Circuits ◽

Declarative Memory ◽

Rapid Eye Movement ◽

Brain Areas ◽

Brain Circuits ◽

New Information ◽

Procedural Task ◽

Synaptic Changes

Abstract Decades of neurobehavioral research has linked sleep-associated rhythms in various brain areas to improvements in cognitive performance. However, it remains unclear what synaptic changes might underlie sleep-dependent declarative memory consolidation and procedural task improvement, and why these same changes appear not to occur across a similar interval of wake. Here we describe recent research on how one specific feature of sleep—network rhythms characteristic of rapid eye movement and non-rapid eye movement—could drive synaptic strengthening or weakening in specific brain circuits. We provide an overview of how these rhythms could affect synaptic plasticity individually and in concert. We also present an overarching hypothesis for how all network rhythms occurring across the sleeping brain could aid in encoding new information in neural circuits.

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Environmental enrichment enhances synaptic plasticity by internalization of striatal dopamine transporters

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x15613525 ◽

2016 ◽

Vol 36 (12) ◽

pp. 2122-2133 ◽

Cited By ~ 15

Author(s):

Myung-Sun Kim ◽

Ji Hea Yu ◽

Chul Hoon Kim ◽

Jae Yong Choi ◽

Jung Hwa Seo ◽

...

Keyword(s):

Synaptic Plasticity ◽

Environmental Enrichment ◽

Functional Improvement ◽

Proximity Ligation Assay ◽

Proximity Ligation ◽

Treatment Benefits ◽

Positron Emission ◽

Walking Tests ◽

Relationship Of

Environmental enrichment (EE) with a complex combination of physical, cognitive and social stimulations enhances synaptic plasticity and behavioral function. However, the mechanism remains to be elucidated in detail. We aimed to investigate dopamine-related synaptic plasticity underlying functional improvement after EE. For this, six-week-old CD-1 mice were randomly allocated to EE or standard conditions for two months. EE significantly enhanced behavioral functions such as rotarod and ladder walking tests. In a [18F]FPCIT positron emission tomography scan, binding values of striatal DAT were significantly decreased approximately 18% in the EE mice relative to the control mice. DAT inhibitor administrated to establish the relationship of the DAT down-regulation to the treatment effects also improved rotarod performances, suggesting that DAT inhibition recapitulated EE-mediated treatment benefits. Next, EE-induced internalization of DAT was confirmed using a surface biotinylation assay. In situ proximity ligation assay and immunoprecipitation demonstrated that EE significantly increased the phosphorylation of striatal DAT as well as the levels of DAT bound with protein kinase C (PKC). In conclusion, we suggest that EE enables phosphorylation of striatal DAT via a PKC-mediated pathway and causes DAT internalization. This is the first report to suggest an EE-mediated mechanism of synaptic plasticity by internalization of striatal DAT.

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