Structural and mechanistic insights into gating and allosteric modulation of GluN1-GluN3A NMDA receptors.
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The authors' code
Python · 291 lines · 8.5 KB · CC-BY-4.0
- #!/opt/anaconda3/bin/python3
- import streamlit as st
- import glob
- import os
- import read_lif
- import numpy as np
- import cv2
- import matplotlib.pyplot as plt
- import math
- import plotly.graph_objects as go
- import SessionState
- from plotly.offline import plot
- from skimage.measure import label, regionprops
- from skimage import data
- from skimage import color
- from skimage.morphology import extrema
- from skimage import exposure
- from scipy import spatial
- def bytescale(data, in_min, in_max):
- data = np.clip(data, in_min, in_max)
- data = (data - float(in_min)) / float(in_max - in_min)
- return np.array(data * 255, dtype=np.uint8)
- def pixel_list_by_radius(radius):
- pixel_list = []
- max_pix = math.ceil(radius)
- for x in range(max_pix+1):
- for y in range(max_pix+1):
- if math.sqrt(x*x+y*y) < radius:
- pixel_list.append((x,y))
- if x > 0:
- pixel_list.append((-x,y))
- if y > 0:
- pixel_list.append((x,-y))
- if x >0 and y > 0:
- pixel_list.append((-x,-y))
- return(pixel_list)
- @st.cache
- def load_zero_frame(filename,sel_series,sel_channel):
- reader = read_lif.Reader(filename)
- series = reader.getSeries()
- image = series[sel_series].getFrame2D(T=0, channel = sel_channel, dtype=np.uint16)
- image_b = bytescale(image,np.min(image),np.max(image))
- return image_b
- @st.cache
- def load_final_frame(filename,sel_series,sel_channel):
- reader = read_lif.Reader(filename)
- series = reader.getSeries()
- num_timepoints = series[sel_series].getNbFrames()
- image = series[sel_series].getFrame2D(T=num_timepoints-1, channel = sel_channel, dtype=np.uint16)
- image_b = bytescale(image,np.min(image),np.max(image))
- return image_b
- @st.cache
- def load_time_series(filename,sel_series,sel_channel):
- reader = read_lif.Reader(filename)
- series = reader.getSeries()
- num_timepoints = series[sel_series].getNbFrames()
- shape = series[sel_series].get2DShape()
- data = np.zeros((shape[0],shape[1],num_timepoints))
- for tp in range(num_timepoints):
- im = series[sel_series].getFrame2D(T=tp, channel=sel_channel,dtype=np.uint16)
- data[:,:,tp] = im
- return data
- @st.cache
- def get_peaks_positions(blur, percent):
- h_maxima = extrema.h_maxima(blur, percent)
- label_h_maxima = label(h_maxima)
- return [(x.centroid[1], x.centroid[0]) for x in regionprops(label_h_maxima)]
- st.sidebar.title("Input selection")
- filename = st.sidebar.selectbox(
- 'Input File',
- glob.glob("*.lif"))
- reader = read_lif.Reader(filename)
- series = reader.getSeriesHeaders()
- sel_series_i = st.sidebar.selectbox(
- 'Series',
- list(range(len(series))),format_func=lambda x:series[x].getName())
- channels = series[sel_series_i].getChannels()
- sel_channel_i = st.sidebar.selectbox(
- 'Channel',
- list(range(len(channels))),format_func=lambda x:channels[x].getAttribute('ChannelTag'))
- #Load timepoint 0 of desired image
- image_b = load_zero_frame(filename,sel_series_i,sel_channel_i)
- image_b_final = load_final_frame(filename,sel_series_i,sel_channel_i)
- # Blur image
- st.sidebar.title("Peak picking parameters")
- blurring = st.sidebar.slider("Blurring", min_value=1, max_value=21, value=5)
- blur = cv2.GaussianBlur(image_b,(blurring,blurring),0)
- blur_final = cv2.GaussianBlur(image_b_final,(blurring,blurring),0)
- #st.image(blur)
- # Threshold image based on percentage
- percent = st.sidebar.slider("H-height", min_value=1., max_value=100., value=10.,step=0.1,format="%.3f")
- percent_final = st.sidebar.slider("Exclude based on final H-height", min_value=1., max_value=100., value=10.,step=0.1,format="%.3f")
- coor_list_start = get_peaks_positions(blur, percent)
- coor_list_final = get_peaks_positions(blur_final, percent_final)
- #st.write(len(coor_list_start))
- #st.write(len(coor_list_final))
- final_tree = spatial.KDTree(coor_list_final)
- coor_list = [ x for x in coor_list_start if final_tree.query(x,distance_upper_bound=3)[1] == len(coor_list_final)]
- #st.write(len(coor_list))
- skip = st.button('skip')
- step1 = st.button('1 step')
- step2 = st.button('2 step')
- step3 = st.button('3 step')
- step4 = st.button('4 step')
- step5 = st.button('5 step')
- ss = SessionState.get(position=1, count0=0, count1=0, count2=0, count3=0, count4=0, count5=0)
- #ss1 = SessionState.get(count=1)
- widget = st.empty()
- #ss1 = SessionState.get(count1=1)
- if skip:
- ss.position = ss.position + 1
- ss.count0 = ss.count0 + 1
- if step1:
- ss.position = ss.position + 1
- ss.count1 = ss.count1 + 1
- if step2:
- ss.position = ss.position + 1
- ss.count2 = ss.count2 + 1
- if step3:
- ss.position = ss.position + 1
- ss.count3 = ss.count3 + 1
- if step4:
- ss.position = ss.position + 1
- ss.count4 = ss.count4 + 1
- if step5:
- ss.position = ss.position + 1
- ss.count5 = ss.count5 + 1
- st.write('skip=',ss.count0,'1 step=',ss.count1, '2 step=',ss.count2, '3 step=',ss.count3, '4 step=',ss.count4, '5 step=',ss.count5)
- max=len(coor_list)
- ss.position = widget.slider('Peak', 0, max, ss.position)
- #ss1.count1 = st.write("", ss1.count1)
- #selected_keypoint_i = st.slider("Select peak", min_value=0, max_value=len(coor_list), value=0,step=1)
- selected_keypoint_i=ss.position
- st.sidebar.title("Trace calculation parameters")
- extraction_radius = st.sidebar.slider("Extraction radius", min_value=0.0, max_value=20.0, value=3.0,step=0.1)
- time_series_images = load_time_series(filename,sel_series_i,sel_channel_i)
- coord = (int(round(coor_list[selected_keypoint_i][0])),int(round(coor_list[selected_keypoint_i][1])))
- #st.write(coord)
- pixel_list = pixel_list_by_radius(extraction_radius)
- trace = []
- for tp in range(time_series_images.shape[2]):
- summation = 0
- for pix in pixel_list:
- ex_pix = (coord[0] + pix[0], coord[1] + pix[1])
- summation += time_series_images[ex_pix[1],ex_pix[0],tp]
- trace.append(summation)
- st.line_chart(trace)
- min_v = np.min(time_series_images)
- max_v = np.max(time_series_images)
- plot_points = np.linspace(0,time_series_images.shape[2]-1,num = 10,dtype=int)
- image_to = np.zeros((30,31*10))
- for i, timepoint in enumerate(plot_points):
- image = bytescale(time_series_images[coord[1]-15:coord[1]+15,coord[0]-15:coord[0]+15,timepoint],min_v,max_v)
- image_to[0:30,31*i:31 * (i+1) -1] = image
- #plt.imshow(image_to)
- #st.pyplot()
- plt.clf()
- st.write(coor_list[selected_keypoint_i])
- # Show image with picked peaks
- show_image=st.button('SHOW IMAGE')
- if show_image:
- # Create figure
- fig = go.Figure()
- # Constants
- img_width = blur.shape[0]
- img_height = blur.shape[1]
- scale_factor = 1.0
- # Add invisible scatter trace.
- # This trace is added to help the autoresize logic work.
- fig.add_trace(
- go.Scatter(
- x=[0, img_width * scale_factor],
- y=[0, img_height * scale_factor],
- mode="markers",
- marker_opacity=0
- )
- )
- # Configure axes
- fig.update_xaxes(
- visible=False,
- range=[0, img_width * scale_factor]
- )
- fig.update_yaxes(
- visible=False,
- range=[0, img_height * scale_factor],
- # the scaleanchor attribute ensures that the aspect ratio stays constant
- scaleanchor="x"
- )
- fig.add_trace(
- go.Heatmap(
- z=blur,
- showscale=False,
- colorscale='Greys',
- reversescale=True
- )
- )
- # Add image
- #fig.update_layout(
- # images=[go.layout.Image(
- # x=0,
- # sizex=img_width * scale_factor,
- # y=img_height * scale_factor,
- # sizey=img_height * scale_factor,
- # xref="x",
- # yref="y",
- # opacity=1.0,
- # layer="below",
- # sizing="stretch",
- # source=blur)]
- #)
- # Configure other layout
- fig.update_layout(
- width=img_width * scale_factor,
- height=img_height * scale_factor,
- margin={"l": 0, "r": 0, "t": 0, "b": 0},
- )
- shapes = [ go.layout.Shape(
- type="rect",
- xref="x",
- yref="y",
- x0=point[0]-2,
- y0=point[1]-2,
- x1=point[0]+2,
- y1=point[1]+2,
- line=dict(
- color="yellow",
- width=3,
- )
- ) for point in coor_list]
- shapes[selected_keypoint_i]["line"]["color"] = "red"
- fig.update_layout(
- shapes=shapes)
- # Disable the autosize on double click because it adds unwanted margins around the image
- # More detail: https://plot.ly/python/configuration-options/
- st.plotly_chart(fig)
- #fig.show(config={'doubleClick': 'reset'})
BleachProcess.py, under CC-BY-4.0 · at the source
Overview
- Vollum Institute, Oregon Health and Science University, Portland, OR USA
- Center for Structural and Functional Neuroscience, Center for Biomolecular Structure and Dynamics, Division of Biological and Biomedical Sciences, University of Montana, Missoula, MT USA
- Howard Hughes Medical Institute, Oregon Health and Science University, Portland, OR USA
Abstract
N-methyl-d-aspartate receptors (NMDARs) mediate excitatory signaling essential for synaptic plasticity and memory. Unlike GluN2-containing NMDARs, GluN3-containing receptors are activated solely by glycine and exhibit profound desensitization and paradoxical potentiation by GluN1-selective antagonists, including CGP-78608 (CGP). Although GluN3A-containing NMDARs regulate synapse pruning and excitotoxicity, and are associated with schizophrenia, autism and stroke, their native stoichiometry and gating mechanism are poorly defined. Here, using single-molecule pull-down analysis, we show that native GluN3A-containing receptors are diheteromeric assemblies. Cryogenic-electron microscopy analysis of GluN1–GluN3A receptors in antagonist-bound, preactive, active and desensitized states, augmented by electrophysiology and pharmacology experiments, show how glycine activates the receptor solely via GluN3A-dependent conformational changes, opening the gate with two-fold symmetry, and induces a roughly four-fold symmetric desensitized state. CGP-bound GluN1 restricts GluN3A rotation, promoting glycine-induced activation by blocking desensitization. These findings illuminate how CGP potentiates GluN1–GluN3A receptor activity, place the receptor gating mechanism on a structural foundation and define the molecular basis for pharmacological modulation.
Reproduced under the paper's license (CC BY), from the paper cited above.
Repository
Its files are read in the Code ↔ Paper reader above.
Zenodo 8161179
Availability: 1 check, the latest on 27 September 2026: the link answers (HTTP 200)
- 27 September 2026: the link answers (HTTP 200)
2 files
- BleachProcess.py, Python, 291 lines
- SessionState.py, Python, 117 lines
Code availability
Custom code used for analyzing single-molecule photobleaching trajectories in this study is available via Zenodo at https://
Reproduced under the paper's license (CC BY), from the paper cited above.
Tracing map
Proposed by the machine: these links were found in the paper and verified at the source, without human review. The map will receive a Zenodo DOI once one of the paper's authors has validated it with their ORCID.
What the map holds:
- 1 repository of the authors' code, each at its verified commit, with its license and how the link was found in the paper;
- 2 scripts, each with its path and the digest of its content;
- no match between paragraphs and code yet;
- neither the text of the paper nor the code itself.
Its JSON (tracing-map.json) is deposited on Zenodo with its DOI once the map is validated.
Data
Datasets cited
- ebi.ac.uk/
pdbe/ , at EMBL-EBI; found in “Data availability”entry - figshare:31004269, at figshare; found in DataCite
- uniprot.org/
uniprot/ , at UniProt; found in the text, “Construct design”p35439 - uniprot.org/
uniprot/ , at UniProt; found in the text, “Construct design”q9r1m7
Data Availability Statement
Data and materials can be obtained from the corresponding authors on request and are available from the following repositories: cryo-EM maps and coordinates have been deposited in the Electron Microscopy Data Bank and in the PDB under accession codes EMD-70399 (http://
Custom code used for analyzing single-molecule photobleaching trajectories in this study is available via Zenodo at https://
Reproduced under the paper's license (CC BY), from the paper cited above.
Versions
The history of this record: each version stored by the harvester or made by a correction of its authors or of the maintainers of its code, and what changed in its facts. The texts of the paper (its abstract, its availability statements) are not part of it; versions that changed only those are not listed.
Version 2, 28 September 2026
- Publisher: n/a → Nature Portfolio
Version 1, 27 September 2026: the first record
Recorded: type, language, journal, volume, issue, pages, dates, 6 authors, 4 keywords, 9 MeSH terms, 3 funders, 82 references.
Cite
This paper
Kim, J., Benton, A. J., Lotti, J. S., Rouzbeh, N., Hansen, K. B., & Gouaux, E. (2026). Structural and mechanistic insights into gating and allosteric modulation of GluN1-GluN3A NMDA receptors. Nature structural & molecular biology, 33(9), 1299-1310. https://
BibTeX
@article{kim2026structur
author = {Kim, Junhoe and Benton, Avery J and Lotti, James S and Rouzbeh, Nirvan and Hansen, Kasper B and Gouaux, Eric},
title = {{Structural and mechanistic insights into gating and allosteric modulation of GluN1-GluN3A NMDA receptors}},
journal = {Nature structural \& molecular biology},
year = {2026},
month = aug,
volume = {33},
number = {9},
pages = {1299--1310},
publisher = {Nature Portfolio},
issn = {1545-9993},
doi = {10.1038/
url = {https://
pmid = {42665662},
pmcid = {PMC13561882}
}
RIS
TY - JOUR
AU - Kim, Junhoe
AU - Benton, Avery J
AU - Lotti, James S
AU - Rouzbeh, Nirvan
AU - Hansen, Kasper B
AU - Gouaux, Eric
TI - Structural and mechanistic insights into gating and allosteric modulation of GluN1-GluN3A NMDA receptors
T2 - Nature structural & molecular biology
J2 - Nat Struct Mol Biol
PY - 2026
DA - 2026/
VL - 33
IS - 9
SP - 1299
EP - 1310
SN - 1545-9993
PB - Nature Portfolio
DO - 10.1038/
UR - https://
LA - en
ER -
CSL-JSON
{
"id": "10.1038/
"type": "article-journal",
"title": "Structural and mechanistic insights into gating and allosteric modulation of GluN1-GluN3A NMDA receptors",
"container-title": "Nature structural & molecular biology",
"author": [
{
"family": "Kim",
"given": "Junhoe"
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{
"family": "Benton",
"given": "Avery J"
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{
"family": "Lotti",
"given": "James S"
},
{
"family": "Rouzbeh",
"given": "Nirvan"
},
{
"family": "Hansen",
"given": "Kasper B"
},
{
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"given": "Eric"
}
],
"container-title-short":
"volume": "33",
"issue": "9",
"page": "1299-1310",
"DOI": "10.1038/
"PMID": "42665662",
"PMCID": "PMC13561882",
"ISSN": "1545-9993",
"publisher": "Nature Portfolio",
"URL": "https://
"language": "en",
"issued": {
"date-parts": [
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2026,
8,
28
]
]
}
}
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