Data for: A synapse-specific refractory period for plasticity at individual dendritic spines
Data files
Feb 18, 2025 version files 10.46 GB
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FloresEtAl_Analysis.zip
13.14 MB
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FloresEtAl_RawData.zip
10.44 GB
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README.md
3.94 KB
Abstract
How newly formed memories are preserved while brain plasticity is ongoing has been a source of debate. One idea is that synapses which experienced recent plasticity become resistant to further plasticity, a type of metaplasticity often referred to as saturation. Here, we probe the local dendritic mechanisms that limit plasticity at recently potentiated synapses. We show that recently potentiated individual synapses exhibit a synapse-specific refractory period for further potentiation. We further found that the refractory period is associated with reduced postsynaptic CaMKII signaling; however, stronger synaptic activation fully restored CaMKII signaling but only partially restored the ability for further plasticity. Importantly, the refractory period is released after one hour, a timing that coincides with the enrichment of several postsynaptic proteins to pre-plasticity levels. Notably, increasing the level of the postsynaptic scaffolding protein, PSD95, but not of PSD93, overcomes the refractory period. Our results support a model in which potentiation at a single synapse is sufficient to initiate a synapse-specific refractory period that persists until key postsynaptic proteins regain their steady-state synaptic levels.
Overview
Fluorescence data was collected using a custom-built 2-photon microscope. Green fluorescence from Green Fluorescent Protein (GFP) or Super Ecliptic Phluorin (SEP) were acquired in channel 1 and red fluoroescence from td-DsRed or DsRedExpress were acquired in channel 2. Analysis was performed using custom MATLAB software.
Forster Resonance Energy Transfer-Fluorescence Lifetime Imaging Microscopy (FRET-FLIM) data was collected using a custom-built 2-photon microscope with time correlated single photon counting (TCSPC). Lifetime data is stored in channel 1 and the fluorescence intensity of a red cell fill is stored in channel 2. Data was analyzed using custom MATLAB software.
Description of the data and file structure
We have provided our raw data in the “FloresEtAl_2024_RawData.zip” file and our analysis files in the “FloresEtAl_2024_Analysis.zip” file. These two folders are sub-divided into files for each individual figure.
Raw data:
Folders for Figures 1, 2, 4, 5 & 6 contain date from 2-photon imaging experiments. These Figure folders are separated into Figure panels folders containing individual cells folders. Cell folders are named with the following convention: [initials, cell number]. Each cell folder contains all of the images for each cell in the experiment with the image name convention: [initials, cell number, dendrite identifier (a or b)]. Z-stacks and maximum projection images are both included and stored in .tif format. A zoomed out image of the whole cell is included in .tif format with the name ‘whole’ added.
Folders for Figures 3 & 4 contain date from 2-photon FRET-FLIM experiments. These Figure folders are separated into Figure panels folders containing individual cells folders with the naming convention: [initials, cell number]. The cell folder contains the raw fluorescence data from channel 2 (red) and a sub-folder named “spc” which contains the raw single photon counting data per pixel. Both are stored in .tif format and use the naming convention [initials, cell number, dendrite identifier (a or b)]. The “spc” folders also include .mat files used to determine whether enough photons were collected.
Analysis:
Folders for Figures 1, 2, 4, 5 & 6 contain analysis files for 2-photon imaging experiments. These Figure folders are separated into Figure panels folders containing .ann files and .xls files. The .ann files store integrated fluorescence data for green and red fluorescence (channel 1 and channel 2, respectively) and the positions of the analyzed ROIs. These have the naming convention [initials, cell number, dendrite identifier (a or b)]. The .xls files store the integrated background subtracted fluorescence for the analyzed spines with the naming convention [initials, cell number, dendrite identifier (a or b), channel (red or green)].
Folders for Figures 3 & 4 contain analysis files for FRET-FLIM experiments. These Figure folders are separated into Figure panels folders containing .MAT files and .xls files. The .MAT files store data detailing the ROI position, the average lifetime (tau_m) and the red fluorescence intensity (int_int2). These .MAT files are named with the following convention:[“A_”, initials, cell number, dendrite identifier (a or b), “_ROI2”]. The .xls files store the raw lifetime and fluorescence intensities for each cell with the convention: [initials, cell number, dendrite identifier (a or b)].
All Figure folders contain summary .xlsx files where normalization and averaging of spine size or average lifetime across cells were performed.
Usage Notes
.tif images can be opened using most image analysis softwares (e.g. ImageJ)
.MAT and .ann files can be opened using MATLAB
.xls and .xlsx files can be opened using Microsoft Excel